Traditional Chinese medicine composition for treating hepatic cirrhosis portal hypertension and application thereof

Abstract

The invention relates to a traditional Chinese medicine composition for treating hepatic cirrhosis portal hypertension and application thereof. The traditional Chinese medicine composition comprises astragalus membranaceus, codonopsis pilosula, atractylodes macrocephala koidz and poria cocos. The invention explores the effects of the compound 215 on portal vein pressure, inflammatory response, hepatic sinus capillary vascularization, hepatic sinus microcirculation and the like of cirrhosis rats. Results show that the compound 215 can effectively reduce portal vein pressure of liver cirrhosis portal hypertension model rats. A brand new mechanism for treating cirrhosis portal hypertension is explored, that is, the compound 215 controls blood flow of mesenteric vessels by adjusting expression of V1aR, expression of V1a receptors in mesenteric artery smooth muscles of cirrhosis portal hypertension rats can be increased, then mesenteric arteriole constriction is caused, blood flow flowing back to portal veins is reduced, and the liver cirrhosis portal hypertension treatment effect is improved. Therefore, portal vein pressure is reduced. The compound 215 disclosed by the invention provides a brand new treatment direction in the aspects of maintaining liver health and overall hemodynamics, and is beneficial to developing a new treatment strategy.

Description

Technical Field

[0001] The present invention relates to the technical field of traditional Chinese medicines, and in particular to a traditional Chinese medicine composition for treating portal hypertension caused by liver cirrhosis and an application thereof. Background Art

[0002] Cirrhosis is a common chronic liver disease caused by long-term liver damage from one or more etiologies, resulting in progressive, diffuse, and fibrotic liver lesions. Cirrhosis is the terminal stage of most chronic liver diseases. Liver histopathology reveals extensive hepatocyte necrosis, nodular regeneration of residual hepatocytes, connective tissue proliferation, and the formation of fibrous septa, leading to structural disruption of the hepatic lobule and the formation of pseudolobules. The liver gradually deforms and hardens, progressing to cirrhosis. The natural course of cirrhosis is a progressive and dynamic process, transitioning from the relatively stable state of compensated cirrhosis to the advanced stage of decompensated cirrhosis. Clinically, cirrhosis is divided into compensated and decompensated stages. The core of this transition is the degree of portal hypertension (PH), which is the primary driver of complications such as variceal bleeding, ascites, renal insufficiency, hepatic encephalopathy (HE), hyperdynamic circulation, and cardiomyopathy.

[0003] At the core of PH is resistance at any point in the portal venous system, resulting in a pressure gradient. In patients with cirrhosis, this resistance is at the level of the hepatic sinusoids, driven by a combination of structural and functional alterations. Sinusoidal structural changes are driven by interactions between hepatic stellate cells (HSCs) and liver sinusoidal endothelial cells (LSECs). In response to injury, HSCs are activated and induce fibrosis, while LSECs undergo phenotypic remodeling, leading to sinusoidal capillarization and increased intrahepatic resistance. Simultaneously, dynamic components of myofibroblast contraction and reduced production of vasodilators (such as nitric oxide) further exacerbate PH resistance. These two fundamental mechanisms contribute to the progressive development of PH, ultimately leading to splanchnic vasodilation, neurohormonal disturbances, systemic vasodilation, decreased mean arterial pressure (MAP), and a hyperdynamic circulatory state.

[0004] Currently, pharmacological treatment for PH is limited to non-selective beta-blockers (NSBBs). NSBBs are the first-line treatment for the prevention and treatment of portal hypertension complications in patients with cirrhosis. NSBBs are the preferred treatment for reducing portal pressure and preventing variceal bleeding, effectively preventing variceal bleeding and reducing bleeding-related mortality. Studies have shown that carvedilol and propranolol, representative NSBBs, reduce the hepatic venous pressure gradient (HVPG) by more than 20% compared to baseline in 64% and 14% of patients with cirrhosis, respectively. Carvedilol has been shown to be more effective than propranolol in reducing portal pressure.

[0005] For patients with clinically significant portal hypertension (CSPH), nonsteroidal anti-inflammatory drugs (NSBBs) have been shown to prevent decompensated cirrhosis events other than variceal bleeding, such as ascites and hepatic encephalopathy. However, current research evidence does not support the use of NSBBs in compensated patients without CSPH. Although there are reliable data supporting their use in end-stage cirrhosis, safety concerns remain in some patients with refractory ascites and spontaneous peritonitis, and their dosage and timing remain to be determined. NSBBs, particularly carvedilol, may be harmful in the treatment of advanced cirrhosis (such as refractory ascites). On the other hand, Traditional Chinese Medicine (TCM) posits that the etiology and pathogenesis of cirrhosis are caused by the invasion and lingering effects of pathogenic factors, leading to the accumulation of pathogenic factors, persistent dampness and heat, liver depression, and spleen qi and blood deficiency. Spleen deficiency is the key factor, and treatment should prioritize strengthening the spleen. The spleen governs blood, which refers to its function of regulating and controlling the normal flow of blood within the vasculature, preventing it from overflowing. This blood regulation function of the spleen is entirely dependent on the consolidation of Qi. A healthy spleen leads to active Qi and blood production, and the Qi's retaining function is robust, preventing blood from spilling out of the vessels. Ascites is a primary symptom of decompensated cirrhosis, classified in Traditional Chinese Medicine as "bloating." Clinically, it is characterized by a distended, drum-like abdomen, sallow complexion, and exposed veins. The pathogenesis of ascites is generally attributed to damage to the liver, spleen, and kidneys, leading to Qi stagnation, blood stasis, and water retention in the abdomen, resulting in bloating. When ascites develops in cirrhosis, clinical practice focuses on regulating the Spleen and Earth to aid the functioning of the Central Palace, promote Yang Qi, distribute body fluids, and dispel water. In the existing technology of liver cirrhosis treatment research, Hou Yixin et al. compared the comprehensive treatment of Western medicine and the method of regulating qi and strengthening spleen in traditional Chinese medicine. By observing the bleeding rate within 1 year and the bleeding-free time within 1 year in patients with liver cirrhosis and esophageal varices, the effect of the Qi-regulating and spleen-strengthening prescription on patients with liver cirrhosis was determined. Among them, the Qi-regulating and spleen-strengthening prescription treatment group of this study was treated with the Qi-regulating and spleen-strengthening prescription (15g of Bupleurum chinense, 15g of White Peony Root, 15g of Angelica Sinensis, 15g of Citrus aurantium, 15g of Codonopsis pilosula, 15g of Astragalus membranaceus, 15g of Atractylodes macrocephala, 15g of Poria cocos, 10g of Tangerine peel, 15g of Chuanxiong on the basis of comprehensive treatment of Western medicine. Add or subtract according to symptoms: if combined with ascites, add 30g of Imperata cylindrica and 30g of Corn Silk; if combined with abdominal distension, add 30g of Citrus aurantium and 15g of Magnolia officinalis; if combined with abdominal distension and loose stools, add 15g of Chinese yam and 20g of stir-fried Coix seed; if combined with poor appetite, add 15g of Chicken Gizzard Stone and 15g each of Jiao Sanxian).The results of this study showed that the risk of EGVB in the Liqi Jianpi group was significantly lower than that in the control group at baseline, and the Liqi Jianpi group was able to delay the bleeding time of patients (Effects of Adjuvant Chinese Patent Medicine Therapy on Prevention of Variceal Rebleeding: A Retrospective Cohort Study. Chin J Integr Med. 2021Aug; 27(8): 589-596. Effect of Liqi Jianpi on Esophageal Variceal Bleeding in Patients with Cirrhosis [J]. Beijing Traditional Chinese Medicine, 2018, 37(10): 959-963+966). However, the aforementioned study did not evaluate the efficacy in patients with cirrhosis and portal hypertension.

[0006] CN114272281A discloses a traditional Chinese medicine compound medicine for treating early-stage liver cirrhosis. The medicine composition comprises astragalus, angelica, safflower, millettia reticulata and gentian. The medicine composition is characterized in that, by weight, the ratio of astragalus: angelica: safflower: millettia reticulata: gentian is (1-5): (1-2): (1-2): (1-8): (1-4), and the medicine can only exert a certain effect on liver tissue of early-stage liver cirrhosis.

[0007] CN 117414406A discloses a traditional Chinese medicine compound composition for treating liver fibrosis and its application. The composition comprises the following components: 8-12 parts of turtle shell, 4-8 parts of leech, 8-12 parts of earthworm, 8-12 parts of triangularis, 8-12 parts of zedoaria, 10-20 parts of bupleurum, 10-20 parts of scutellaria, 6-12 parts of pinellia, 10-20 parts of codonopsis, 10-20 parts of angelica, 10-20 parts of ligusticum, 10-20 parts of white peony root, 10-20 parts of red peony root, 10-20 parts of stir-fried atractylodes, 10-20 parts of tuckahoe, 10-20 parts of oriental rhizome, 20-40 parts of wormwood, 10-20 parts of polygonum cuspidatum, 15-25 parts of astragalus, and 8-12 parts of liquorice. The traditional Chinese medicine compound composition has complex ingredients and is mainly used to treat liver fibrosis.

[0008] Existing medications and traditional Chinese medicine formulas for treating portal hypertension in cirrhosis primarily target anti-fibrotic, multi-component, and multi-target approaches to improve PH in cirrhosis. However, these treatments are complex and ineffective, and research on alternative therapeutic mechanisms for portal hypertension in cirrhosis is limited. Therefore, developing new medications to treat portal hypertension has significant clinical and scientific value.

[0009] In addition, on the one hand, there are differences in understanding among those skilled in the art; on the other hand, the applicant studied a large number of documents and patents when making the present invention, but due to space limitations, not all details and contents are listed in detail. However, this does not mean that the present invention does not have the characteristics of these prior arts. On the contrary, the present invention already has all the characteristics of the prior art, and the applicant reserves the right to add relevant prior art to the background technology. Summary of the Invention

[0010] Driven by the severity of PH, cirrhosis progresses through various stages of progression, from compensation to decompensation. The downstream effects of worsening PH lead to multiple pathophysiological pathways, contributing to the major complications of cirrhosis, including ascites, variceal bleeding, and hepatic encephalopathy. The severity of PH is also a major driver of further late complications, such as hyperdynamic circulation, hepatorenal syndrome, and cirrhotic cardiomyopathy. Currently, there are few effective pharmacological treatment options for PH in clinical practice. Current treatments for PH primarily include medication, endoscopic therapy, surgery, and interventional therapy. Pharmacological therapy aims to control the various complications of PH in cirrhosis and can be used for primary and secondary prevention of esophageal and gastric variceal bleeding, as well as for the treatment of acute bleeding. Nonsteroidal anti-inflammatory drugs (NSBBs) primarily reduce portal venous pressure by blocking β-1 adrenergic receptors, resulting in a decrease in cardiac output and splanchnic blood flow. Carvedilol improves intrahepatic vascular resistance by antagonizing α-adrenergic activity and increasing intrahepatic nitric oxide release.

[0011] The second class of medications for treating portal hypertension in cirrhosis is vasopressin, such as terlipressin, which reduces portal vein pressure and improves splanchnic vasodilation. Terlipressin is highly effective in improving renal function in patients with hepatorenal syndrome. Compared with other vasoactive drugs (including midodrine, octreotide, and norepinephrine), terlipressin is more effective in reversing hepatorenal syndrome and improving short-term survival. However, because terlipressin also activates V2 receptors in the renal collecting ducts, causing water retention, it partially attenuates terlipressin-mediated fluid excretion via V1a receptors. Terlipressin is also associated with some serious adverse events, such as respiratory failure. Terlipressin use in patients with a high end-stage liver disease score is known to be associated with potentially life-threatening ischemic adverse events. In addition, there are some drugs such as sodium-glucose co-transporter 2 inhibitors and statins. The former can treat patients with refractory ascites through natriuresis, osmotic diuresis and inhibition of the renin-angiotensin-aldosterone system, and the latter can reduce portal vein pressure in rats by improving hepatic hemodynamics and sinusoidal endothelial dysfunction in the liver microvascular system. However, the therapeutic effect and prognosis are still poor.

[0012] The use of NSBBs in PH has been well studied and has pleiotropic mechanisms, such as preventing bacterial translocation, antioxidant properties, and improving survival in acute-on-chronic liver failure. However, not all patients respond hemodynamically to NSBBs, and in advanced cirrhosis (such as with refractory ascites), NSBB therapy may be harmful. Studies have shown that carvedilol's vasodilatory effects may enhance systemic hypotension and lead to reduced renal perfusion, thus limiting its clinical application, particularly in patients with cirrhosis and refractory ascites and those with advanced decompensated cirrhosis. The clinical risk-benefit ratio of NSBBs worsens in the advanced decompensated stage, particularly in patients with refractory ascites, hypotension, and renal impairment, who have an increased risk of medication harms, clinically defining a closed therapeutic window. Therefore, the development of novel agents to reduce portal venous pressure is urgently needed for PH in cirrhosis.

[0013] To address the shortcomings of the existing technology, the present invention provides a traditional Chinese medicine composition for treating portal hypertension in cirrhosis. The composition comprises: astragalus root, codonopsis root, atractylodes macrocephala, and poria, wherein the weight ratio of astragalus root: codonopsis root: atractylodes macrocephala: poria is 1-2:1:1:1. Hereinafter, the composition is referred to as Compound 215.

[0014] According to a preferred embodiment, in the traditional Chinese medicine composition, the weights of Astragalus, Codonopsis, Atractylodes and Poria are 30 grams, 15 grams, 15 grams and 15 grams respectively.

[0015] Preferably, in the traditional Chinese medicine composition, the astragalus is raw astragalus.

[0016] Preferably, in the traditional Chinese medicine composition, the Atractylodes macrocephala is stir-fried Atractylodes macrocephala.

[0017] The second aspect of the present invention provides a pharmaceutical preparation, which comprises the traditional Chinese medicine composition provided by the first aspect of the present invention as an active ingredient and any one or more pharmaceutically acceptable carriers.

[0018] According to a preferred embodiment, the pharmaceutical preparation can be in any pharmaceutically acceptable dosage form, preferably a tablet, capsule, granule or liquid preparation.

[0019] The third aspect of the present invention provides the use of the traditional Chinese medicine composition provided by the first aspect of the present invention or the pharmaceutical preparation provided by the second aspect of the present invention in reducing portal hypertension in liver cirrhosis and corresponding pharmaceutical applications.

[0020] The fourth aspect of the present invention provides a method for preparing a traditional Chinese medicine preparation based on the traditional Chinese medicine composition provided by the first aspect of the present invention, the method comprising: mixing the ingredients astragalus, codonopsis, atractylodes and poria in proportion to obtain a mixture; soaking, decocting or extracting the mixture to obtain a traditional Chinese medicine composition; and filtering, concentrating and other steps to obtain a final preparation that can be used clinically.

[0021] According to a preferred embodiment, the method for preparing the Chinese medicine composition includes: weighing appropriate amounts of Astragalus, Codonopsis, Atractylodes and Poria in proportion; heating and refluxing or ultrasonic extraction to obtain an extract; filtering and concentrating to obtain a concentrate; and drying the concentrate to obtain the Chinese medicine composition.

[0022] The fifth aspect of the present invention provides a traditional Chinese medicine preparation prepared by the method provided by the fourth aspect of the present invention.

[0023] The sixth aspect of the present invention provides the use of the traditional Chinese medicine composition provided by the first aspect of the present invention, the pharmaceutical preparation provided by the second aspect of the present invention, or the traditional Chinese medicine preparation provided by the fifth aspect of the present invention in reducing portal hypertension in cirrhosis by upregulating the expression of vasopressin 1a receptors and corresponding pharmaceutical applications.

[0024] The seventh aspect of the present invention provides the use of the traditional Chinese medicine composition provided by the first aspect of the present invention, the pharmaceutical preparation provided by the second aspect of the present invention, or the traditional Chinese medicine preparation provided by the fifth aspect of the present invention in reducing portal hypertension in cirrhosis by increasing mesenteric vasoconstriction and reducing portal vein reflux, and corresponding pharmaceutical applications.

[0025] The eighth aspect of the present invention provides the use of the traditional Chinese medicine composition provided by the first aspect of the present invention, the pharmaceutical preparation provided by the second aspect of the present invention, or the traditional Chinese medicine preparation provided by the fifth aspect of the present invention in maintaining systemic hemodynamic stability and corresponding pharmaceutical applications.

[0026] According to a preferred embodiment, maintaining systemic hemodynamic stability includes maintaining stability of mean arterial pressure and heart rate.

[0027] The ninth aspect of the present invention provides the use of the traditional Chinese medicine composition provided in the first aspect of the present invention, the pharmaceutical preparation provided in the second aspect of the present invention, or the traditional Chinese medicine preparation provided in the fifth aspect of the present invention in reducing inflammatory response and corresponding pharmaceutical applications.

[0028] According to a preferred embodiment, reducing the inflammatory response includes reducing the gene expression levels of IL-6 (interleukin-6) and TNF-α (tumor necrosis factor-α).

[0029] The tenth aspect of the present invention provides the use of the traditional Chinese medicine composition provided in the first aspect of the present invention, the pharmaceutical preparation provided in the second aspect of the present invention, or the traditional Chinese medicine preparation provided in the fifth aspect of the present invention in improving hepatic sinusoidal remodeling and inhibiting pathological angiogenesis to reduce portal hypertension in cirrhosis, and corresponding pharmaceutical applications.

[0030] According to a preferred embodiment, the traditional Chinese medicine composition provided by the first aspect of the present invention, the pharmaceutical preparation provided by the second aspect of the present invention, or the traditional Chinese medicine group preparation provided by the fifth aspect of the present invention reduces portal hypertension in cirrhosis by inhibiting the gene expression level and protein expression level of VEGF and / or CD31.

[0031] The eleventh aspect of the present invention provides the use of the traditional Chinese medicine composition provided by the first aspect of the present invention, the pharmaceutical preparation provided by the second aspect of the present invention, or the traditional Chinese medicine preparation provided by the fifth aspect of the present invention in inhibiting the activation and aggregation of macrophages and corresponding pharmaceutical applications.

[0032] According to a preferred embodiment, the Chinese medicine composition provided by the first aspect of the present invention, the pharmaceutical preparation provided by the second aspect of the present invention, or the Chinese medicine preparation provided by the fifth aspect of the present invention reduces portal hypertension in cirrhosis by inhibiting the number of CD68-positive cells in the liver.

[0033] The twelfth aspect of the present invention provides the use of the traditional Chinese medicine composition provided by the first aspect of the present invention, the pharmaceutical preparation provided by the second aspect of the present invention, or the traditional Chinese medicine preparation provided by the fifth aspect of the present invention in reducing portal hypertension in cirrhosis by improving hepatic sinusoidal capillarization and hepatic sinusoidal microcirculation, and corresponding pharmaceutical applications.

[0034] The thirteenth aspect of the present invention provides the use of the traditional Chinese medicine composition provided by the first aspect of the present invention, the pharmaceutical preparation provided by the second aspect of the present invention, or the traditional Chinese medicine preparation provided by the fifth aspect of the present invention in reducing portal hypertension in cirrhosis by increasing the fenestrae of hepatic sinusoidal endothelial cells, and corresponding pharmaceutical applications.

[0035] The fourteenth aspect of the present invention provides the use of the traditional Chinese medicine composition provided by the first aspect of the present invention, the pharmaceutical preparation provided by the second aspect of the present invention, or the traditional Chinese medicine preparation provided by the fifth aspect of the present invention in reducing the bleeding rate and / or rebleeding rate of esophageal variceal rupture in cirrhosis of the liver and corresponding pharmaceutical applications.

[0036] In a fifteenth aspect, the present invention provides the use of the Chinese medicine composition provided in the first aspect, the pharmaceutical preparation provided in the second aspect, or the Chinese medicine preparation provided in the fifth aspect for improving ALT (aspartate aminotransferase, also known as alanine aminotransferase) and AST (aspartate aminotransferase) levels in subjects with cirrhosis and portal hypertension, and corresponding pharmaceutical applications. Preferably, the subject can be a mammal, such as a rat or a human.

[0037] A sixteenth aspect of the present invention further provides a method for treating portal hypertension in cirrhosis, comprising: administering the Chinese medicine composition provided by the first aspect of the present invention, the pharmaceutical preparation provided by the second aspect of the present invention, or the Chinese medicine preparation provided by the fifth aspect of the present invention to a subject by oral gavage to significantly reduce portal pressure and improve clinical symptoms. A further method of treating portal hypertension in cirrhosis of the present invention further provides a method for treating portal hypertension in cirrhosis of the present invention, comprising: administering the Chinese medicine composition provided by the first aspect of the present invention, the pharmaceutical preparation provided by the second aspect of the present invention, or the Chinese medicine preparation provided by the fifth aspect of the present invention to a subject by oral administration to reduce portal pressure and improve clinical symptoms.

[0038] The seventeenth aspect of the present invention also provides a method for treating portal hypertension caused by cirrhosis, comprising: jointly using the traditional Chinese medicine composition provided by the first aspect of the present invention, the pharmaceutical preparation provided by the second aspect of the present invention, or the traditional Chinese medicine preparation provided by the fifth aspect of the present invention and other drugs for controlling portal hypertension caused by cirrhosis to enhance the effect of reducing portal pressure.

[0039] Technical Effects: This study investigated the effects of Compound 215 on portal pressure, liver inflammation, sinusoidal capillarization, sinusoidal microcirculation, and the fenestration rate of sinusoidal endothelial cells in rats with cirrhosis using a CCL4 rat model of portal hypertension. The results demonstrated that Compound 215 effectively reduced portal pressure in rats with cirrhosis and portal hypertension, while maintaining the stability of mean arterial pressure and heart rate. Compound 215 significantly reduced liver inflammation in rats with cirrhosis and portal hypertension. Compound 215 inhibited abnormal angiogenesis in the liver, helping to stabilize vascular function and reduce vascular leakage and abnormal permeability. Compound 215 improved sinusoidal capillarization and sinusoidal microcirculation in rats with cirrhosis and portal hypertension. Treatment with Compound 215 significantly increased the fenestration rate of sinusoidal endothelial cells, enhancing the exchange of substances between hepatocytes and plasma and alleviating hepatocyte damage. The reduction in portal pressure in cirrhosis is primarily due to improved sinusoidal microcirculation and reduced intrahepatic vascular resistance.

[0040] Unexpectedly, the present invention found that the liver collagen content in rat liver slices after intervention treatment with Compound 215 did not significantly decrease, and did not significantly improve the level of liver fibrosis in rats with cirrhosis and portal hypertension. The path of Compound 215 in reducing portal pressure in cirrhosis is to regulate vasoactive substances (ET-1, eNOS, NO) in intrahepatic blood vessels and simultaneously reduce the expression of VEGF to inhibit intrahepatic vasoconstriction, thereby reducing intrahepatic vascular resistance. It also reduces damage to hepatocytes and hepatic sinusoidal endothelial cells by inhibiting inflammatory reactions. In addition, it is also manifested in improving hepatic sinusoidal capillarization and increasing the fenestrae of hepatic sinusoidal endothelial cells. Existing Chinese herbal compound prescriptions such as Fuzheng Huayu Capsule, Compound 861, and Kangxian Ruangan Granule all treat cirrhosis and portal hypertension through mechanisms such as anti-liver fibrosis, improving hemodynamics, regulating vasoactive substances, and inhibiting inflammatory reactions. Another novel mechanism of this invention is that Compound 215 controls mesenteric blood flow by regulating the expression of Via receptors. Compound 215 increases the expression of Via receptors in the smooth muscle of the mesenteric arteries of rats with cirrhosis and portal hypertension, thereby causing constriction of the mesenteric arterioles, reducing blood flow back to the portal vein and thereby lowering portal pressure. This process is particularly important in patients with cirrhosis, as portal hypertension caused by cirrhosis can lead to a series of complications, such as esophageal varices and ascites. By reducing portal pressure, Compound 215 can effectively mitigate the occurrence of these complications. This mechanism not only helps improve the symptoms of patients with cirrhosis but also provides a theoretical and practical basis for Compound 215's use in the treatment of related diseases. Through its comprehensive regulatory effects, Compound 215 shows broad application prospects in maintaining liver health and overall hemodynamics.

[0041] The research results of the present invention provide clinical basis and effective treatment strategy for patients with portal hypertension due to cirrhosis, and effectively improve the quality of life and disease prognosis of patients with cirrhosis.

[0042] According to a preferred embodiment, a traditional Chinese medicine composition comprises astragalus root, codonopsis root, atractylodes macrocephala, poria cocos, salvia miltiorrhiza root, and panax notoginseng. This composition is suitable for treating patients with portal hypertension due to cirrhosis and blood stasis. Preferably, the ratio of astragalus root, codonopsis root, atractylodes macrocephala, poria cocos, salvia miltiorrhiza root, and panax notoginseng is 1-2:1:1:1:1:1-2:1. Salvia miltiorrhiza root not only promotes blood circulation and improves liver blood circulation, but also reduces the blood pressure elevation caused by astragalus root and the symptoms of dysmenorrhea in women.

[0043] According to a preferred embodiment, a traditional Chinese medicine composition comprises astragalus, codonopsis, atractylodes, poria, bupleurum, and white peony root. This composition is suitable for treating patients with cirrhosis and portal hypertension with symptoms of liver depression. Preferably, the ratio of astragalus, codonopsis, atractylodes, poria, bupleurum, and white peony root is 1-2:1:1:1:0.5:0.5-1. White peony root not only nourishes the blood and softens the liver, but also reduces the side effects of internal heat and allergies that can be caused by codonopsis and atractylodes. BRIEF DESCRIPTION OF THE DRAWINGS

[0044] Figure 1 The statistical results of the therapeutic effect of Compound 215 on portal pressure, mean arterial pressure and heart rate in rats with cirrhosis and portal hypertension are shown;

[0045] Figure 2 This is the result of the effect of Compound 215 on liver inflammation in rats with cirrhosis and portal hypertension;

[0046] Figure 3 This is the result of the effect of Compound 215 on fibrosis in rats with cirrhosis and portal hypertension;

[0047] Figure 4 The results are as follows: Effects of Compound 215 on hepatic sinusoidal remodeling and pathological angiogenesis in rats with cirrhosis and portal hypertension:

[0048] Figure 5 This is the result of the effect of Compound 215 on vasoactive molecules in rats with cirrhosis and portal hypertension;

[0049] Figure 6 This is the result of the effect of Compound 215 on the function of liver sinusoidal endothelial cells;

[0050] Figure 7 This is the result of the effect of Compound 215 on the extrahepatic blood vessels of cirrhotic rats. DETAILED DESCRIPTION

[0051] The following is a detailed description with reference to the accompanying drawings.

[0052] The experimental methods used in the following examples, unless otherwise specified, are conventional methods. Materials, reagents, etc. used in the following examples, unless otherwise specified, are commercially available. In the figures of the present invention, NS represents no statistical significance, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Via receptor is equivalent to ViaR. In the present invention, portal pressure is also referred to as portal vein pressure and is used interchangeably; portal hypertension is also referred to as portal hypertension and is used interchangeably.

[0053] Example

[0054] This example investigates the effects of Compound 215 on portal vein pressure, liver function, liver fibrosis, sinusoidal capillarization, and fenestration rate of sinusoidal endothelial cells in rats with cirrhosis. Statistical data in this example are presented as mean ± standard deviation. The model group corresponds to the CCL4 rat model; the treatment group corresponds to the Compound 215-treated CCL4 rat model.

[0055] 1. Preparation of CCL4-induced Portal Hypertension Model in Rats

[0056] Forty-eight rats weighing 195-210 g were obtained from the Experimental Animal Center of Capital Medical University and divided into three groups: control group, model group, and treatment group (Compound 215 treatment group). All experimental procedures for the care and use of rats were consistent with those of the Capital Medical University Ethics Committee.

[0057] To establish a rat model of liver cirrhosis and portal hypertension, 40% CCL4 (CCL4 dissolved in olive oil) was administered intraperitoneally twice weekly at 2 ml / kg for 10 weeks. Starting from the fifth week of modeling, Compound 215 granules were administered orally at an equivalent dose of 1.4 mg / kg, 6.3 times the human dose, once daily for 6 weeks. The rat model of liver cirrhosis and portal hypertension was established over a total of 10 weeks. A control group received the same amount of saline.

[0058] The relevant indicators of rats in the control group, model group and intervention group were examined.

[0059] 2. Animal Dissection and Handling

[0060] According to the preliminary experimental design, rats from the corresponding groups were taken for experimental operations at the dissection time point. After anesthesia, the animals were fixed in the supine position, the abdominal cavity was opened, the gross morphology of the liver and the ascites were observed and recorded, and the portal pressure was measured immediately. After the pressure measurement was completed, the puncture needle and the pressure transducer were disconnected, and the syringe was immediately connected directly to the end of the puncture needle hose. About 4 to 5 mL of portal vein blood was slowly drawn, and the supernatant was separated within 24 hours (packed, stored in the dark at 4°C, allowed to stand for 3 hours, 3000 rpm / min×15 minutes, and the supernatant was taken), and placed in a -80°C refrigerator for testing. After the animals were sacrificed by blood sampling, all liver tissues were removed, and liver specimens from the same part were taken and immersed in neutral formalin solution for fixation. After 24 hours, they were dehydrated in graded alcohol, transparentized with xylene, embedded in paraffin at 56°C, and sliced 4 μm thick for hematoxylin-eosin (HE) staining of liver and intestinal tissues. The remaining liver tissue was stored at -80°C for future use.

[0061] 3. Project testing

[0062] (1) Portal pressure in rats with cirrhosis and venous hypertension. Figure 1 Show, among them, Figure 1 A is the test result of rat portal vein pressure; Figure 1 B is the test result of mean arterial pressure of rats; Figure 1 C is the detection result of rat heart rate.

[0063] After 5 weeks of treatment, portal vein pressure values of rats in the control group, model group and treatment group were measured using portal vein direct puncture method. Figure 1 In Figure A, the horizontal axis represents the different treatment groups, and the vertical axis represents the portal vein pressure (mmHg). Starting from the 5th week of modeling, this experiment began to use Compound 215 intervention on some model rats (treatment group). After 6 weeks of administration, the portal vein pressure of the model group rats increased significantly (P<0.0001), while the portal vein pressure of the treatment group rats was significantly reduced compared with the model group during the same period (P<0.01), indicating that with the continuous intervention of CCL4, the portal vein pressure of the model group rats continued to increase, and Compound 215 can effectively reduce the portal vein pressure of rats with cirrhosis and portal hypertension model. Specifically, Compound 215 may reduce the vascular resistance of the liver by improving the microvascular circulation of the liver, thereby reducing the retention of blood in the portal vein system and effectively reducing the portal vein pressure.

[0064] Figure 1 In Figure B, the horizontal axis represents the different treatment groups, and the vertical axis represents mean arterial pressure (mmHg). Compared with the control group, the mean arterial pressure of the model group rats decreased significantly. After the model group rats were treated with Compound 215, the mean arterial pressure of the treatment group rats did not differ significantly from that of the model group rats. Figure 1 In Figure C, the horizontal axis represents the different treatment groups, and the vertical axis represents heart rate (beats / min). Compared with the control group, the heart rate of the model group rats did not decrease significantly. After the model group rats were treated with Compound 215, the heart rate of the treated rats did not differ significantly from that of the model group rats. Therefore, Compound 215 reduced portal pressure in rats with cirrhosis and portal hypertension without reducing systemic hemodynamics, including heart rate and mean arterial pressure. Compound 215 may selectively act on the portal venous system without affecting the systemic vascular system and hemodynamics. This selective effect helps to specifically reduce portal pressure without negatively affecting the systemic circulation. Compound 215 may help improve endothelial function, promote vasodilation, and reduce portal vein resistance, while having less impact on the systemic vascular system. Since heart rate and mean arterial pressure did not decrease, it indicates that Compound 215 may not have a negative impact on the heart's contractile function or heart rate, thereby protecting the normal function of the heart. For patients with cirrhosis, they may already have problems regulating blood pressure. After administering Compound 215, there was no significant change in mean arterial pressure and heart rate, which means that while reducing portal hypertension, Compound 215 can maintain the stability of systemic blood pressure, which is very important for patients with cirrhosis.

[0065] (2) Hepatic inflammation in rats with cirrhosis and portal hypertension. Figure 2 shown.

[0066] Figure 2 The results of the effect of Compound 215 on liver inflammation in rats with cirrhosis and portal hypertension are shown in the table below. Figure 2 A is the HE staining result of liver tissue, scale bar = 100 μm; Figure 2 B is the result of Compound 215 improving the gene expression level of inflammatory factors. The horizontal axis represents the different groups, and the vertical axis represents the relative mRNA expression levels of IL-6 (interleukin-6) and TNF-α (tumor necrosis factor-α); Figure 2 C is the immunofluorescence staining results of CD31 and CD68 in different groups.

[0067] Figure 2 In Figure A, compared to the model group, the liver tissue treated with Compound 215 displayed clear cell outlines and distinct boundaries, indicating that Compound 215 promotes the integrity of hepatocyte membranes and normalizes cell morphology. Furthermore, HE staining of the liver tissue following Compound 215 treatment revealed normal hepatic plate arrangement and sinusoidal structure, demonstrating that Compound 215 helps restore the liver's microstructure. HE staining also revealed neat and orderly arrangement of hepatic cords and reduced inflammatory cell infiltration following Compound 215 treatment, reflecting the repair of liver tissue structure and the restoration of liver function. Compound 215 promotes the overall health of liver tissue.

[0068] Figure 2In Figure B, the relative mRNA expression of IL-6 in the model group rats was significantly increased compared with the control group (P < 0.0001). After 6 weeks of treatment with Compound 215, the relative mRNA expression of IL-6 in the Compound 215-treated group was significantly decreased compared with the model group (P < 0.01). IL-6 is involved in immune responses, inflammatory responses, and tissue repair. In liver diseases, elevated IL-6 levels are associated with hepatocellular injury, inflammation, and fibrosis. IL-6 may also serve as a biomarker for acute and chronic liver disease, and changes in its levels can reflect disease activity and severity. Compound 215 may directly inhibit components of the IL-6 gene transcription or translation. Compound 215 may also reduce IL-6 expression by blocking inflammatory signaling pathways, such as JAK-STAT and NF-κB. In terms of immune regulation, Compound 215 may reduce IL-6 production by regulating the activity and function of immune cells. In terms of metabolism, Compound 215 may affect IL-6 expression by altering cellular metabolism, for example, by regulating metabolic pathways associated with inflammation. IL-6 production can be stimulated by endotoxins, and Compound 215 may reduce IL-6 expression by improving intestinal barrier function and reducing endotoxin translocation. A decrease in IL-6 expression helps alleviate the inflammatory state in the body. In liver disease, IL-6 may promote hepatocellular damage and fibrosis, and its reduced expression may help protect the liver. IL-6 is involved in regulating metabolic processes, and reducing its expression may help improve symptoms associated with metabolic syndrome, such as insulin resistance. Moderately reducing IL-6 levels may help balance the inflammatory response and promote the repair of damaged tissues. The decrease in IL-6 relative mRNA expression reflects the potential therapeutic effect of R215 in regulating immune responses, reducing inflammation, and promoting physical health.

[0069] Compared with the control group, the relative mRNA expression of TNF-α in the model group was significantly increased (P < 0.0001). After 6 weeks of treatment with Compound 215, the relative mRNA expression of TNF-α in the Compound 215-treated group was significantly decreased compared with the model group (P < 0.01). TNF-α is a major inflammatory cytokine produced by activated macrophages and plays a key role in regulating immune responses and inflammatory processes. In liver diseases, TNF-α is involved in hepatocyte damage and apoptosis and is associated with the development of hepatitis, cirrhosis, and liver cancer. Anti-TNF-α therapy has potential therapeutic effects in liver damage caused by certain autoimmune liver diseases and inflammatory bowel disease. Compound 215 may directly inhibit components of the TNF-α gene transcription or translation. Compound 215 may also reduce TNF-α expression by blocking inflammatory signaling pathways. Reduced relative mRNA expression of TNF-α may help alleviate inflammatory responses in the liver and other organs. In terms of immunomodulation, reduced TNF-α expression may help regulate the immune system and reduce autoimmune liver damage. TNF-α is involved in the progression of liver fibrosis, and Compound 215 may help inhibit the development of fibrosis by reducing its expression. Decreased relative TNF-α mRNA expression may create a more favorable environment for liver repair and regeneration. Figure 2 Result B fully demonstrates that compound R215 can effectively reduce liver inflammation.

[0070] CD31, also known as PECAM-1 (platelet endothelial cell adhesion molecule), is a cell surface glycoprotein highly expressed on cell types such as vascular endothelial cells, platelets, monocytes / macrophages, and lymphocytes. It is a marker of liver vascular endothelial cells and is involved in intercellular adhesion and signaling. In immunofluorescence studies, CD31 is often used to label vascular endothelial cells, allowing for the study of liver vascular structure and angiogenesis. CD31 expression can be used to assess tumor microvessel density, helping to understand the angiogenic and imaging characteristics of tumors. Figure 2In Figure C, compared to the control group, the fluorescence intensity of CD31 increased in the model group. After six weeks of treatment with Compound 215, the fluorescence intensity of CD31 in the treatment group was significantly reduced, indicating that Compound 215 inhibits abnormal angiogenesis in the liver, particularly angiogenesis associated with tumor growth or inflammation. CD31 is involved in regulating cell adhesion during inflammation. Compound 215 may indirectly reduce CD31 expression and fluorescence intensity by alleviating the inflammatory response. Compound 215 may help repair damaged endothelial cells, restore their normal barrier function, and reduce abnormal CD31 expression. Compound 215 may reduce CD31 expression by improving microcirculation in organs such as the liver and reducing the activation state of endothelial cells. Regarding apoptosis, Compound 215 may affect the apoptotic process of endothelial cells, reducing CD31 release caused by cell damage or death. Compound 215 may also reduce vascular permeability and the gaps between endothelial cells, which may help reduce fluid and protein exudation and improve inflammation and edema. The decrease in CD31 fluorescence intensity may also indicate the inhibitory effect of Compound 215 on tumor angiogenesis, which helps limit tumor nutrient supply and growth. Compound 215 may reduce damage to vascular endothelial cells caused by oxidative stress and inflammatory factors, thereby reducing CD31 expression. Compound 215 may reduce CD31 expression by intervening in signaling pathways in vascular endothelial cells, such as those regulating VEGF and TNF-α. The decrease in CD31 fluorescence intensity after Compound 215 treatment may reflect the combined effects of Compound R215 in anti-angiogenesis, anti-inflammatory, antioxidant, and microcirculatory improvement, which help to improve pathological conditions and promote tissue repair and organ function recovery.

[0071] CD68 is a transmembrane glycoprotein highly expressed in monocytes and macrophages and is often used as a macrophage marker. In liver disease, CD68 expression can help identify and quantify Kupffer cells (macrophages within the liver), which play a vital role in the liver's immune response and inflammatory processes. Detecting CD68 through immunofluorescence can assess the degree of liver inflammation and the activation state of macrophages, which is important for understanding the progression of liver disease and treatment response. Figure 2In Figure C, the fluorescence intensity of CD68 in the model group was significantly increased compared to the control group. After six weeks of treatment with Compound 215, the fluorescence intensity of CD68 in the treatment group was significantly decreased, indicating that Compound 215 has a significant anti-inflammatory effect (reducing inflammatory responses in the liver and other tissues). Macrophages play a key role in inflammation and immune responses. The decrease in CD68 indicates that Compound 215 inhibits macrophage activation and aggregation. Macrophage activation is associated with tissue damage, and CD68 may alleviate tissue damage by reducing macrophage involvement. The decrease in CD68 may also reflect the regulatory effect of Compound 215 on the immune system, helping to restore immune balance. Compound 215 may reduce the production of inflammatory factors and CD68 expression by improving the tissue microenvironment. Reducing macrophage activity may help reduce inflammation-related tissue damage and create favorable conditions for tissue repair and regeneration. In liver diseases, macrophage activation is associated with the development of liver fibrosis, and the decrease in CD68 may indicate the anti-fibrotic potential of Compound 215. Macrophages also participate in regulating apoptosis, and the decrease in CD68 expression may be related to the reduction of apoptosis by Compound 215. Compound 215 reduces oxidative stress through its antioxidant components, indirectly affecting macrophage activity and CD68 expression. Sustained macrophage activation is associated with poor disease prognosis, and Compound 215's inhibition of CD68 expression may help improve disease prognosis. The decrease in CD68 may also indicate that Compound 215 has the potential to be a targeted therapy for certain inflammatory diseases. In summary, the decrease in CD68 fluorescence intensity after Compound 215 treatment may reflect the potential therapeutic effects of Compound 215 in modulating immune responses, reducing inflammation, protecting tissues, and improving disease prognosis.

[0072] (3) Liver fibrosis level in rats with cirrhosis and portal hypertension, e.g. Figure 3 shown.

[0073] HE staining: The fixed rat liver was dehydrated with a 75% by volume ethanol aqueous solution, transparentized with xylene, and then embedded in paraffin. The embedded wax block was cut into 4-6 μm thin slices using a microtome, flattened in hot water, mounted on a glass slide, and dried in a 45°C constant temperature oven. The sections were HE stained. The HE-stained sections were placed under an optical microscope to observe pathological changes in the liver tissue and photographed.

[0074] Quantitative analysis of liver fibrosis: Dewax paraffin sections to water; chromate treatment or demercurate precipitation (this step can be omitted for formaldehyde-fixed tissues); wash with tap water and then distilled water; stain the nuclei with Regaud's hematoxylin solution or Weigert's hematoxylin solution for 5-10 minutes; wash thoroughly with water; if over-staining, differentiate with hydrochloric acid and alcohol; wash with distilled water; use Masson's ponceau acid fuchsin solution for 5-10 minutes; soak briefly in 2% glacial acetic acid aqueous solution; differentiate for 3-5 minutes with 1% phosphomolybdic acid aqueous solution; stain directly with aniline blue or light green solution for 5 minutes without washing; soak briefly in 0.2% glacial acetic acid aqueous solution; transparentize with 95% alcohol, anhydrous alcohol, or xylene, and then seal with neutral gum; observe and photograph.

[0075] Figure 3 The results of a pathological study on the improvement of liver fibrosis in rats with portal hypertension and cirrhosis were presented. Figure 3 The staining results of Masson and Sirius Red are specifically shown.

[0076] according to Figure 3 Liver pathology results showed that compared with the control group, the CCL4-induced model group showed severe liver tissue structural abnormalities, with some hepatocyte necrosis and hemorrhage, a small number of hepatocytes edematous, mild cytoplasmic staining, lack of clear cell outlines, and more disorganized hepatic cords (structure disorder of the rat hepatic lobules, irregular hepatocyte morphology, and obvious inflammatory cell infiltration in the portal areas and hepatic lobules). Treatment with Compound 215 did not significantly improve the liver tissue sections of the rats. Figure 3 The results showed that Compound 215 did not significantly improve the level of liver fibrosis in rats with cirrhosis and portal hypertension. Specifically, Masson staining showed no significant difference in the stained area between the treatment group and the model group. Sirius red staining also showed no significant difference in the stained area between the treatment group and the model group.

[0077] Unexpectedly, the results of this example show significant differences in the mechanism of action of the traditional Chinese medicine compositions for treating liver cirrhosis. Existing traditional Chinese medicine compositions such as Fuzheng Huayu Capsules, Compound 861, and Kangxian Ruangan Granules all treat portal hypertension in liver cirrhosis by mechanisms such as anti-liver fibrosis and inhibition of inflammatory responses. However, the results of this example show that Compound 215 does not treat portal hypertension in liver cirrhosis by improving the level of liver fibrosis, but involves a completely new mechanism, as shown in the following table. Figure 7 .

[0078] (4) Hepatic sinusoidal remodeling and pathological angiogenesis in rats with cirrhosis and portal hypertension. Figure 4 Show, among them, Figure 4 A is the gene expression level and protein expression level of VEGF; Figure 4 B is the fenestration rate of the hepatic sinusoids; Figure 4 C is the immunohistochemical result of liver vWF; Figure 4 D is the immunofluorescence result of vWF in the liver.

[0079] VEGF (vascular endothelial growth factor) is a marker of angiogenesis. Figure 4 In Figure A, the relative mRNA expression of VEGF in the treatment group was significantly decreased compared with the model group (P < 0.05); the protein expression of VEGF in the treatment group was also significantly decreased compared with the model group (P < 0.01). VEGF expression is associated with endothelial cell fenestrae (endothelial cell permeability). After six weeks of treatment with Compound 215, both the relative mRNA expression and protein expression of VEGF in the intervention group were significantly decreased. VEGF is a key factor in promoting angiogenesis, playing an important role in regulating angiogenesis, endothelial cell proliferation, and increasing vascular permeability. The significant decrease in the relative mRNA and protein expression of VEGF in the treatment group suggests that Compound 215 may inhibit abnormal angiogenesis in the liver. VEGF is involved in regulating the tissue microenvironment. Compound 215 may improve the tissue microenvironment by reducing VEGF expression, thereby reducing tissue hypoxia and acidity and promoting tissue repair. The decrease in relative mRNA and protein expression of VEGF may promote the restoration of normal tissue structure. In addition, the reduction in the relative mRNA and protein expression of VEGF may also help stabilize vascular function and reduce vascular leakage and abnormal permeability.

[0080] Figure 4 In B, the results of liver scanning electron microscopy observations showed that compared with the model group, the fenestration rate of the hepatic sinusoids in the treatment group increased and the hepatic sinusoidal remodeling was improved. When using Compound 215 to treat portal hypertension in cirrhosis, this example found that Compound 215 can significantly improve liver structure and function. The fenestration rate of the hepatic sinusoids is a key indicator for measuring the status of hepatic sinusoidal endothelial cells. Its increase means that the integrity and function of the hepatic sinusoidal endothelial cells have been restored, and the capillarization of the hepatic sinusoids has been reversed, which helps to improve the blood microcirculation of the liver. Specifically, Compound 215 may promote the repair and functional recovery of hepatic sinusoidal endothelial cells and reduce the activation and dysfunction of endothelial cells.

[0081] CD31- and vWF-positive staining areas reflect sinusoidal capillarization and sinusoidal endothelial cell dysfunction, respectively. Sinusoidal capillarization refers to the loss of the unique fenestrae of sinusoidal endothelial cells, forming a basement membrane, which impairs the exchange between hepatocytes and the blood.

[0082] Figure 4 C and Figure 4In Figure D, compared with the control group, the immunofluorescence staining intensity of vWF in the model group was significantly increased, and the vWF-positive staining area reflected more severe sinusoidal endothelial cell dysfunction. After six weeks of treatment with Compound 215, the immunofluorescence staining intensity of vWF in the treatment group was significantly reduced, indicating significant improvement in sinusoidal endothelial cell dysfunction. The reduction in vWF-positive areas after Compound 215 treatment reflects improved sinusoidal microvascular function. Alternatively, Compound 215 may reduce hepatic inflammation, thereby reducing endothelial cell activation and vWF expression. Oxidative stress is a factor in sinusoidal endothelial cell dysfunction, and the antioxidant components of Compound 215 may help reduce oxidative damage and protect endothelial cells. The reduction in vWF-positive areas may be related to reduced vascular permeability, suggesting that Compound 215 may have a role in reducing vascular leakage. Compound 215 may also reduce hepatic vascular resistance, thereby promoting normal blood-hepatocyte exchange. The reduction in vWF-positive areas can serve as an important indicator for monitoring the therapeutic efficacy of Compound 215 and disease progression. In conclusion, the reduction of vWF-positive staining area after treatment with Compound 215 reflects from multiple perspectives its significant technical effects in improving the function of hepatic sinusoidal endothelial cells, inhibiting inflammation and fibrosis, regulating immune response, and improving hemodynamics.

[0083] CD31 is a marker of hepatic sinusoidal capillarization. Figure 4In Figure E, compared with the control group, the immunofluorescence staining intensity and extent of CD31 in the model group were significantly increased. After six weeks of treatment with Compound 215, the intensity and extent of CD31 immunofluorescence staining in the treatment group decreased. Compound 215 may promote the restoration of sinusoidal capillarization and reduce basement membrane formation, thereby bringing the CD31 expression pattern closer to normal. The decreased intensity and extent of CD31 staining may indicate an improved sinusoidal microenvironment, for example, by reducing the impact of inflammation on sinusoidal structure. The decreased intensity and extent of CD31 staining may also be related to improved hepatic hemodynamics, with Compound 215 helping to enhance hepatic blood perfusion. Compound 215 may reduce abnormal angiogenesis by inhibiting the expression of angiogenic factors such as VEGF, thereby reducing CD31 staining intensity. With the reduction in CD31-positive areas, the exchange between hepatocytes and the blood may be improved, contributing to enhanced liver metabolism and detoxification capabilities. The reduced extent of CD31 staining reflects a reduction in sinusoidal capillarization, suggesting that Compound 215 helps restore normal sinusoidal function. Compound 215 may reduce inflammatory cell infiltration in the sinusoidal region and improve the environment of sinusoidal endothelial cells through its anti-inflammatory and immunomodulatory effects. Compound 215 acts directly on sinusoidal endothelial cells, promoting their functional recovery, including the formation and maintenance of fenestrae and improving sinusoidal permeability. These results demonstrate the significant efficacy of Compound 215 in improving sinusoidal endothelial cell structure, inhibiting abnormal angiogenesis, alleviating inflammation, and promoting the recovery of hepatocyte function.

[0084] After treatment with Compound 215, the CD31 and vWF positive staining areas in the treatment group decreased, indicating that Compound 215 can effectively reduce the expression of vWF and CD31, suggesting that hepatic sinusoidal capillarization is reduced and endothelial dysfunction is improved, proving that Compound 215 can help maintain the normal structure and function of the liver.

[0085] (5) Effects of Compound 215 on vasoactive molecules in rats with cirrhosis and portal hypertension, including changes in vasoconstrictor (ET-1) and vasodilator (eNOS, NO) molecules, such as Figure 5 As shown, Figure 5 A shows the protein expression results of phosphorylated eNOS and eNOS in the liver; Figure 5 B is the concentration of nitric oxide in the liver; Figure 5 C is the gene expression level of ET-1, which constricts blood vessels; Figure 5 D is the immunofluorescence result of ET-1.

[0086] eNOS (endothelial nitric oxide synthase) is an enzyme expressed in liver endothelial cells that plays a key role in regulating vascular tone and blood pressure. eNOS catalyzes the conversion of L-arginine to nitric oxide (NO). Nitric oxide gas, a signaling molecule, promotes vasodilation, thereby lowering blood pressure. Regulating eNOS activity in the liver is crucial for the prevention and treatment of conditions such as portal hypertension. The activity of p-eNOS (phosphorylated endothelial nitric oxide synthase) can be regulated by multiple mechanisms, including phosphorylation. When eNOS is phosphorylated by specific kinases (such as protein kinase A or protein kinase G), its ability to produce nitric oxide (NO) is enhanced. When phosphorylated (i.e., p-eNOS), eNOS activity is enhanced, producing more NO, which contributes to vasodilation and lowers blood pressure. Measuring p-eNOS levels can assess endothelial cell function and vascular health.

[0087] ET-1 (endothelin-1) is a potent vasoconstrictor peptide that can bind to endothelin receptors to cause contraction of vascular smooth muscle cells and increase vascular resistance.

[0088] Compared with the model group, the phosphorylation level of eNOS in the treatment group was significantly increased (P<0.05). Figure 5 As shown in A. Compared with the model group, the concentration of NO in the treatment group was significantly increased (P<0.05). Figure 5 As shown in Figure B. Therefore, Compound 215 has a positive effect on the key vasodilator eNOS. The phosphorylation level of eNOS was significantly increased, indicating that Compound 215 can effectively activate the activity of the eNOS enzyme. As a result, the concentration of NO also increased significantly. As a key vasodilator molecule, NO can promote the relaxation of smooth muscle cells in the blood vessel wall by increasing the amount released by endothelial cells, thereby reducing vascular tension. This result shows that Compound 215 can not only reduce the burden on the liver but also help alleviate complications such as ascites and esophageal variceal bleeding caused by portal hypertension. In addition, the increase in NO has the effects of inhibiting platelet aggregation and inhibiting leukocyte adhesion, which helps reduce the risk of thrombosis, improve the liver's microcirculatory environment, and promote liver cell repair and regeneration, which has a positive impact on improving patients' quality of life and long-term prognosis.

[0089] Figure 5 C and Figure 5In D, compared with the model group, the gene expression level of ET-1 in the treatment group was significantly decreased (P<0.01), and the fluorescence intensity of ET-1 was weakened, indicating that Compound 215 can reduce the level of ET-1. The reduction in the expression level of ET-1 gene means that the contraction of blood vessels is weakened and the vascular tension is reduced, thus having a positive therapeutic effect on portal hypertension. As the level of ET-1 decreases, the vascular resistance of the liver decreases, blood flow is improved, and portal vein pressure decreases. This result may also be related to the positive effects of Compound 215 in anti-inflammatory, antioxidant, and hepatic sinusoidal remodeling. In summary, Compound 215 may help improve hepatic sinusoidal hemodynamics, increase the fenestration rate of hepatic sinusoids, and improve hepatic sinusoidal remodeling.

[0090] (6) Function of liver sinusoidal endothelial cells, the results are as follows Figure 6 As shown, Figure 6 A shows the contractility changes of liver sinusoidal endothelial cells (LSEC) under different stimuli; Figure 6 B is the ET-1 immunofluorescence result of liver sinusoidal endothelial cells; Figure 6 C is a scanning electron micrograph of hepatic sinusoidal endothelial cells.

[0091] Figure 6 In Figure 1, LSECs refer to liver sinusoidal endothelial cells (LSECs), which are endothelial cells in the liver's microvasculature. They form the inner wall of the liver sinusoids and possess a unique fenestration structure, playing a key role in material exchange and immune regulation. LSECs play a crucial role in maintaining liver function and capillarization of the liver sinusoids. Figure 6 A is the result of the three-dimensional collagen cell contraction test. This test observes the changes in LSEC by comparing the ratio of the collagen contraction area to the original uncontracted area of the culture medium (the blue circle is the contraction area). Specifically, Figure 6In panel A, "LSEC" corresponds to untreated sinusoidal endothelial cells; "LSEC+R215" corresponds to sinusoidal endothelial cells treated with compound 215; "LSEC+ET-1" corresponds to sinusoidal endothelial cells treated with ET-1; and "LSEC+ET-1+R215" corresponds to sinusoidal endothelial cells treated with both ET-1 and compound 215. LSECs exhibit a certain degree of contractility within the collagen lattice, but significantly relax upon stimulation with compound 215 (P < 0.05). See the corresponding results for the "LSEC+R215" treatment group. LSECs significantly contracted upon ET-1 stimulation (P < 0.001). See the corresponding results for the "LSEC+R215" treatment group. Simultaneous stimulation with ET-1 and compound 215 resulted in significant relaxation of LSECs compared to treatment with ET-1 alone (P < 0.05). These results demonstrate that compound 215 has a positive effect on the morphology of sinusoidal endothelial cells. LSECs play a crucial role in liver microcirculation. They not only provide nutrients and oxygen to liver cells but also regulate immune responses and clear toxins from the blood. In the pathological state of portal hypertension in cirrhosis, the function and structure of LSECs may be impaired, leading to obstructed blood flow in the hepatic sinusoids. Treatment with Compound 215 significantly dilated LSECs and improved endothelial cell function.

[0092] Figure 6 In Figure B, serum containing Compound 215 at different concentrations (10%, 20%, and 30%) had varying effects on ET-1 expression. Compared to the model group, treatment with Compound 215 at different concentrations significantly reduced the fluorescence intensity of ET-1. Compound 215 significantly inhibited ET-1 expression, and the inhibitory effect of Compound 215 on ET-1 was dose-dependent. In this example, 20% and 30% Compound 215 were more effective in suppressing ET-1 expression.

[0093] Openings in the sinusoidal endothelial cells enable the exchange of substances between hepatocytes and blood, including insulin, lipoproteins, etc., to maintain metabolic balance. Figure 6 C is the result of compound 215 increasing the fenestration rate of liver sinusoidal endothelial cells (indicated by the red arrows in the figure). Figure 6 C includes the window opening rates of different groups when the scale bar = 20.0 μm and the scale bar = 10.0 μm. Figure 6C Results showed that compared with the control group, the fenestrae of the liver sinusoidal endothelial cells in the model group were significantly reduced. After treatment with Compound 215, the fenestrae in the treatment group increased significantly (red arrows indicate LSEC fenestrae), indicating that Compound 215 can increase the fenestrae of liver sinusoidal endothelial cells. Portal hypertension can impair the function of liver sinusoidal endothelial cells and reduce the fenestrae. In terms of restoring endothelial cell function, Compound 215 may improve endothelial cell activity and promote the recovery of their normal physiological function, thereby increasing the fenestrae. This recovery helps improve the liver's microenvironment and enhance the liver's ability to exchange blood and nutrients. In terms of alleviating the inflammatory response, Compound 215 can reduce the inflammatory burden of liver sinusoidal endothelial cells by inhibiting the release of proinflammatory cytokines (such as TNF-α and IL-6). The reduction of the inflammatory response helps restore the structure and function of liver sinusoidal endothelial cells, thereby promoting an increase in the fenestrae. In terms of improving liver hemodynamics, Compound 215 can dilate liver vessels and improve blood flow, reducing portal vein pressure, thereby improving blood oxygen supply to the liver, promoting functional recovery of sinusoidal endothelial cells, and increasing fenestration rate. Healthy hepatocytes can promote the normal function of sinusoidal endothelial cells. This interaction helps enhance fenestration rate, thereby improving the liver's metabolic and detoxification capabilities. Compound 215 may also improve the interaction between hepatocytes and sinusoidal endothelial cells by regulating the liver's microenvironment, enhancing intercellular signaling. This regulation helps stabilize liver function and improve overall liver health. Furthermore, Compound 215 may promote normal sinusoidal structure and function by affecting extracellular matrix and matrix remodeling, further promoting an increase in fenestration rate. The health of the extracellular matrix is crucial for normal cellular function. Compound 215 may also modulate signaling pathways associated with sinusoidal remodeling, such as reducing transforming growth factor-beta (TGF-β) activity, inhibiting hepatic stellate cell activation, and reducing extracellular matrix deposition, thereby promoting liver structural recovery.

[0094] (7) Effects on extrahepatic vessels in cirrhotic rats. Figure 7 As shown, Figure 7 A is the HE staining result of mesentery; Figure 7 B is the immunofluorescence result of CD31 in the mesentery; Figure 7 C is the immunohistochemical result of vasopressin la (V1a) receptor in mesenteric smooth muscle cells; Figure 7 D is the expression result of VlaR (V1a receptor) protein; Figure 7 E is the immunofluorescence result of V1aR.

[0095] The effects of Compound 215 on the extrahepatic vasculature of rats with cirrhosis are primarily manifested in the mesenteric vessels. Vasodilation of the mesentery, a key vascular source of portal blood flow, significantly increases portal venous blood flow. This increased blood flow leads to elevated portal vein pressure, exacerbating symptoms of portal hypertension. This mechanism is particularly pronounced in cirrhosis and related diseases, further impacting the patient's hemodynamic status. Figure 7 In Figure A, the model group showed thinning of the intestinal wall and enlargement of the lumen, suggesting reduced mesenteric vasoconstriction and increased blood volume. After treatment with Compound 215, the treated group showed thickening of the intestinal wall and a reduction in the lumen, suggesting increased mesenteric vasoconstriction, decreased blood flow, and decreased portal venous return. Specifically, intestinal wall thickening may reflect improvements in intestinal microvasculature and remodeling of intestinal wall tissue. Compound 215 improves intestinal blood supply and enhances microcirculation, thereby promoting intestinal tissue repair and regeneration. Intestinal wall thickening generally indicates enhanced intestinal barrier function. Compound 215 has anti-inflammatory and reparative properties, helping to strengthen tight junctions between intestinal epithelial cells and reduce intestinal permeability, thereby reducing the absorption of harmful substances and improving overall health. Compound 215 reduces the lumen, which not only reduces portal venous return but also potentially facilitates more efficient nutrient transport. With a smaller lumen, efficient blood flow ensures adequate oxygen and nutrient supply to the intestinal tissue, promoting liver repair. By improving liver function, Compound 215 alleviates portal hypertension at its source, indirectly promoting normalized intestinal blood supply. Improvement in liver structure and function helps regulate the physiological state of the intestines and makes the intestinal wall healthier.

[0096] Figure 7 In Figure B, compared to the control group, CD31 expression in the mesentery of the model group increased. After treatment with Compound 215, CD31 expression in the mesentery of the treatment group decreased. This result suggests that Compound 215 can reduce mesenteric neovascularization. Compound 215 can effectively reduce abnormal angiogenesis caused by cirrhosis and portal hypertension, helping to restore normal blood circulation. Reduced neovascularization may indicate improved microcirculation. Compound 215 can promote blood circulation and improve intestinal microcirculation, stabilizing mesenteric blood flow and reducing the risk of intestinal ischemia. Excessive neovascularization may lead to mesenteric congestion and edema, impairing intestinal function. Treatment with Compound 215 can reduce this unnecessary vascular growth, reduce the burden on the intestine, and thus improve its function. Compound 215 can lower portal vein pressure, thereby reducing the abnormal stimulation of portal hypertension on mesenteric vessels. Compound 215 may also inhibit the formation of abnormal neovascularization by enhancing endogenous repair mechanisms. Compound 215 promotes the recovery of liver and intestinal function. These benefits indicate that Compound 215 has outstanding potential value in the treatment of portal hypertension in cirrhosis.

[0097] V1aR is a G protein-coupled receptor that is widely present in multiple tissues in the body, especially in vascular smooth muscle, liver, kidney and brain tissue. Figure 7 C and Figure 7 D The results showed that compared with the control group, the expression of V1aR in the model group was reduced (P<0.001), and after treatment with Compound 215, the expression of V1aR in the treatment group was significantly increased (P<0.01).

[0098] Figure 7 In Figure E, SMA labels smooth muscle cells. Compared with the control group, V1aR expression was reduced in the model group. However, after treatment with Compound 215, the immunofluorescence intensity of V1aR in the treatment group was significantly enhanced, indicating a significant increase in V1aR expression. V1aR acts on smooth muscle cells, and upon activation, it can cause vasoconstriction. Increased V1a binding to the receptor activates the G protein signaling pathway, triggering contraction of arterial smooth muscle cells. This contraction leads to a decrease in mesenteric blood flow. This reduction in mesenteric blood flow further impairs venous return. Due to the connection between the mesenteric vessels and the portal venous system, reduced intestinal blood flow also reduces blood flow returning to the portal vein, thereby lowering portal vein pressure. Specifically, increased V1a causes arterial smooth muscle cell contraction, which in turn causes arterial vasoconstriction. This mesenteric vasoconstriction reduces blood flow, venous return, and, in turn, portal pressure. With mesenteric vasoconstriction and reduced venous return, portal vein pressure decreases.

[0099] It should be noted that the above-mentioned specific embodiments are exemplary, and those skilled in the art can come up with various solutions inspired by the disclosure of the present invention, and these solutions also belong to the disclosure scope of the present invention and fall within the protection scope of the present invention. Those skilled in the art should understand that the present invention specification and its drawings are illustrative and do not constitute a limitation on the claims. The scope of protection of the present invention is defined by the claims and their equivalents. The present invention specification contains multiple inventive concepts, such as "preferably" and "according to a preferred embodiment", which means that the corresponding paragraph discloses an independent concept, and the applicant reserves the right to file a divisional application based on each inventive concept. Throughout the text, the features guided by "preferably" are only an optional method and should not be understood as having to be set, so the applicant reserves the right to abandon or delete the relevant preferred features at any time.

Claims

1. A Chinese medicine composition for treating portal hypertension in cirrhosis of the liver, characterized in that: The Chinese medicine composition comprises: Astragalus, Codonopsis, Atractylodes and Poria. Among them, the ratio of Astragalus: Codonopsis: Atractylodes: Poria is 1-2:1:1:1 by weight.

2. A pharmaceutical preparation, characterized in that The pharmaceutical preparation comprises the traditional Chinese medicine composition according to claim 1 as an active ingredient and any one or more pharmaceutically acceptable carriers.

3. Use of the Chinese medicine composition according to claim 1 or the pharmaceutical preparation according to claim 2 in the preparation of a medicament for reducing portal hypertension in cirrhosis.

4. A method for preparing a Chinese medicine preparation based on the Chinese medicine composition according to claim 1, characterized in that: The method comprises: mixing the ingredients of astragalus, codonopsis, atractylodes and poria in proportion to obtain a mixture; soaking, decocting or extracting the mixture to obtain the Chinese medicine composition; and filtering, concentrating and other steps to obtain a final preparation suitable for clinical application.

5. The Chinese medicine preparation prepared according to the method of claim 4.

6. Use of the Chinese medicine composition according to claim 1, the pharmaceutical preparation according to claim 2, or the Chinese medicine preparation according to claim 5 in the preparation of a medicament for reducing portal hypertension in cirrhosis by upregulating the expression of vasopressin 1a receptor.

7. Use of the Chinese medicine composition according to claim 1, the pharmaceutical preparation according to claim 2 or the Chinese medicine preparation according to claim 5 in the preparation of a medicament for reducing portal hypertension in cirrhosis by increasing mesenteric vasoconstriction and reducing portal vein reflux.

8. Use of the Chinese medicine composition according to claim 1, the pharmaceutical preparation according to claim 2, or the Chinese medicine preparation according to claim 5 in the preparation of a drug for maintaining systemic hemodynamic stability.

9. Use of the Chinese medicine composition according to claim 1, the pharmaceutical preparation according to claim 2 or the Chinese medicine preparation according to claim 5 in the preparation of a medicament for reducing inflammatory response.

10. Use of the traditional Chinese medicine composition according to claim 1, the pharmaceutical preparation according to claim 2, or the traditional Chinese medicine preparation according to claim 5 in the preparation of a medicament for improving hepatic sinusoidal remodeling and inhibiting pathological angiogenesis to reduce portal hypertension in cirrhosis.

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