References

References

[1] Pei Zhengxue. Casual Talk on the Treatment of Leukemia. Collection of Pei Zhengxue's Medical Talks and Cases, Gansu Science and Technology Press, 2004:24-25

[2] Pei Zhengxue. Supporting the Body and Nourishing the Root—Immunity. Collection of Pei Zhengxue's Medical Experiences, Gansu Science and Technology Press, 2003:234-246

[3] Pei Zhengxue. Acute Monocytic Leukemia. Practical Internal Medicine Combining Traditional Chinese and Western Medicine, People's Health Publishing House, 1996:321-336

[4] Zhang Guiqiong. The Effect of Pei's Hematopoietic Granules on the Proliferative Activity of Splenic Lymphocytes and Cytokines in H₂₂ Tumor-Bearing Mice. Dissertation, 2007

[5] Huang Bangrong. Experimental Study on the Impact of Pei's Hematopoietic Granules on the Immune System of H₂₂ Tumor-Bearing Mice. Dissertation, 2007

[6] Bai Lijun. Experimental Study on the Impact of Pei's Hematopoietic Granules on the Bone Marrow Hematopoietic System of Aplastic Anemia Model Mice. Dissertation, 2006

[7] Wang Xiaoli. Experimental Study on the Impact of Pei's Hematopoietic Granules on the Immune System of Aplastic Anemia Model Mice. Dissertation, 2006

[8] Wang Ning. Experimental Study on the Impact of Pei's Hematopoietic Granules on Apoptosis of H₂₂ Tumor Cells and the Expression of p53 and Caspase-3 Proteins. Dissertation, 2008

[9] Wang Zhuo. Experimental Study on the Impact of Pei's Hematopoietic Granules on Apoptosis of H₂₂ Tumor Cells and the Expression of NF-κB. Dissertation, 2008

[10] Ding Jiexia. The Effect of Pei's Hematopoietic Granules Combined with 5-FU on the Cell Cycle and Apoptosis Rate of Mouse H₂₂ Tumor Cells. Dissertation, 2009

[11] Zhang Chouchou. The Effect of Pei's Hematopoietic Granules Combined with 5-FU on Bone Marrow DNA and Serum TNF-α and IFN-γ Levels in Tumor-Bearing Mice. Dissertation, 2009

[12] Xue Wenhan. Teacher Pei Zhengxue's Experience in Treating Primary Liver Cancer. Journal of Traditional Chinese Medicine Classics, December 1998

[13] Li Min, Xue Wenhan, Li Wei, et al. Observation of the Therapeutic Effects of Pei's "Lanzhou Formula" Combined with Chemotherapy in 10 Cases of Cancer. National Medicine Forum, 1998

[14] Pei Zhengxue, Editor-in-Chief. Radionuclide Therapy and "Supporting the Body and Nourishing the Root." Collection of Pei Zhengxue's Medical Talks and Cases, Gansu Science and Technology Press, 2004:31-33

[15] Pei Zhengxue. Clinical Notes on the Treatment of Esophageal Cancer. Journal of Traditional Chinese Medicine, July 2002

[16] Xu Shuyun, Bian Rulian, Chen Xiu. Pharmacological Experimental Methods. People's Health Publishing House, 2002:1757-1827

[17] Dai Xinyi, Chen Linxiang, Zhou Daihan, et al. Experimental Study on the Anti-Tumor Effects of Shen Tao Soft Liver Pills on Tumor-Bearing Animals. China Oncology, October 2001, Issue 7:426-428

[18] Li Yikui. Pharmacological Experimental Methods for Traditional Chinese Medicine. Shanghai Science and Technology Press, 1991:36

[19] Sun Zhen, Chen Shiliang, Gu Wenying, et al. Experimental Study on the In Vivo Anti-Tumor Effects of Grifola Frondosa Polysaccharides. Pharmaceutical Biotechnology, August 2001, Issue 5:279-283

[20] Lv Houjun. The Relationship Between VEGF Expression in Liver Cancer Tissue and Micrometastasis in Lymph Nodes. Basic and Clinical Oncology, 2008, Issue 21(4):348

[21] Pei Zhengxue. Collection of Pei Zhengxue's Medical Experiences. Gansu Science and Technology Press, 2003:378-381

[22] Tao Yong, Wu Min. The Effect of Fangji Huangqi Decoction on the Functions of MΦ and T Cells in Spleen-Deficient Mice. Journal of Anhui College of Traditional Chinese Medicine, 2000, Issue 19(1):48-49

[23] Liao Zijun. Modern Oncology Pharmacology. World Book Publishing Company, 2002:409

[24] Fan Weike. Fundamentals of Modern Oncology. People's Health Publishing House, 2005:220

[25] Gong Feili. Medical Immunology. Beijing: Science Press, 2004:6

[26] Chen Weifeng. Medical Immunology. People's Health Publishing House, 2005:100-115

[27] Xu Dujuan, Chen Minzhu. The Anti-Tumor Effects and Mechanisms of Astragalus Polysaccharides. Chinese Hospital Pharmacy Journal, 2005, Issue 25(10):923-925

[28] Folkman. Antiangiogenesis in Cancer Therapy: Endostatin and Its Mechanisms of Action. Exp Cell Res, 2006, Issue 312(5):594

[29] Zou Qifei, Zhang Feng. The Expression of Angiogenin and Vascular Endothelial Growth Factor in Liver Cancer and Their Clinical Significance. Jiangsu Medicine, 2006, Issue 32(5):436

[30] Pan Yunlong. Research on Nanogold or IL-12 Inhibiting the Proliferation of Vascular Endothelial Cells and Angiogenesis in H₂₂ Liver Cancer. Doctoral Dissertation, 2007

Research on Pei Zhengxue's Series of Formulas and Medicines

Abstract

Objective: To explore the regulatory effect of Pei's Hematopoietic Granules (PG) on the immune system of H₂₂ tumor-bearing mice and its inhibitory effect on tumor tissue.

Methods: A H₂₂ tumor-bearing mouse model was established by subcutaneous injection at the right anterior axillary region. The inhibitory effect of PG on mouse tumors was observed, and the concentrations of EGF were measured using ELISA, while the concentrations of IL-1 were detected using RIA.

Results:

1. The tumor inhibition rates in all PG groups were higher than those in the model group, with the medium-dose group showing particularly significant results (P<0.05);
2. The thymus index (TI) and spleen index (SI) in all PG groups increased, with significant increases in the medium- and high-dose groups (P<0.05), and the medium-dose group showed a significant increase in SI (P<0.05);
3. The EGF concentrations in all PG groups were lower than those in the model group, with the medium-dose group showing a significant difference (P<0.05);
4. The IL-1 concentrations in all PG groups were lower than those in the model group, with the medium- and high-dose groups showing significant differences (P<0.05).

Conclusion: PG exerts an anti-tumor effect by increasing the weight of the thymus and spleen in H₂₂ tumor-bearing mice, inhibiting EGF formation and IL-1 secretion.

Keywords: Pei's Hematopoietic Granules (PG); H₂₂ (transplanted hepatocellular carcinoma); IL-1; EGF; immune function

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Foreword

Primary liver cancer (PLC) is a common malignant tumor of the digestive tract and one of the most prevalent malignancies worldwide. It has a rapid onset, severe progression, and poor prognosis, making it a major threat to human health. Approximately 6.35 million new cases of malignant tumors are diagnosed globally each year, and according to a report by the World Health Organization (WHO), the annual number of new cases is expected to reach 15 million by 2020. Among these, liver cancer accounts for 260,000 cases (4% of all malignant tumors). China accounts for about 55% of global liver cancer cases, ranking second among cancer-related deaths, and has been listed as a key disease for cancer prevention and control in China's Cancer Prevention and Control Plan Outline.

Currently, the treatment of liver cancer mainly involves surgical resection in the early stages, supplemented by radiotherapy and chemotherapy. However, existing data show that the rate of surgical resection is low, and the recurrence rate after resection is high. Moreover, liver cancer is a tumor that is insensitive to radiotherapy and chemotherapy and prone to metastasis. As most patients are diagnosed at an advanced stage, they often lose the opportunity for surgery. While the medical community seeks and uses anti-cancer drugs, it has found that many chemical anti-cancer drugs often affect normal cells when acting on target cells, and most chemotherapeutic agents used clinically have varying degrees of mutagenic and genotoxic effects. Consequently, treating tumors simultaneously increases the risk of patients developing a second type of cancer. In contrast, plant-based medicines seem to have less obvious genotoxicity.

Traditional Chinese medicine believes that the fundamental cause of human disease lies in "internal deficiency of vital energy." The "Suwen·Yipian Cifa Lun" states: "When vital energy is preserved within, pathogenic factors cannot invade"; the "Suwen·Pingre Bing Lun" says: "Wherever pathogenic factors gather, qi must be deficient." In terms of treatment, it proposes the therapeutic principle of "supporting the body and strengthening the root." The "vital energy" of the human body is also called "true qi," as stated in the "Lingshu·Ci Jie Zhen Xie Pian": "True qi is what we receive from heaven, combined with the qi of food, to nourish the body." True qi encompasses both innate and acquired factors. Throughout history, physicians have attributed the innate aspect to the two kidneys, calling it "yuan qi" (primordial qi), and the acquired aspect to the spleen and stomach, calling it "zhong qi" (grain qi). "The spleen is the foundation of postnatal life and the source of qi and blood production," thus governing zhong qi; "the kidneys are the foundation of prenatal life and store marrow to generate essence," thus governing yuan qi. The transformation and transportation of postnatal spleen soil rely on the warming of innate kidney yang; the ascent of innate true yang...

Research on Pei Zhengxue's Series of Formulas and Medicines

The transformation and transportation of postnatal spleen soil rely on the nourishment of postnatal spleen soil; the spleen and kidneys complement each other and jointly maintain the growth of bodily functions. Therefore, in the principle of "supporting the body and strengthening the root," "strengthening the spleen" and "tonifying the kidneys" are the two most important aspects. Experimental studies have proven that the "spleen" and "kidneys" in traditional Chinese medicine have significance in various systems, including the immune system, endocrine system, metabolic system, autonomic nervous system, and gastrointestinal-endocrine system. Strengthening the spleen and tonifying the kidneys can improve the functions of these systems, which is the essence of the "supporting the body and strengthening the root" principle. There is a great deal of commonality between the concept of "supporting the body and strengthening the root" in traditional Chinese medicine and modern immunology [6-14]. The "vital energy" mentioned in traditional Chinese medicine generally represents the body's intrinsic ability to resist disease, which is equivalent to the physiological function of the human immune system. The immune system consists of non-specific immunity and specific immunity. The "zhong qi" and "wei qi" in traditional Chinese medicine are similar to non-specific immunity in modern immunology, while "yuan qi" is similar to specific immunity. Specific and non-specific immunity promote each other to jointly achieve the body's immune response, and "zhong qi" and "yuan qi" also promote each other to jointly form the body's vital energy effect. "Supporting the body and strengthening the root" can enhance the body's immune function. With the development of immunology and molecular biology, research has confirmed that the occurrence and development of malignant tumors are closely related to the decline of the body's immune function. In the treatment of tumors, patients often cannot achieve complete cure due to local tumor eradication (surgery), radiotherapy, chemotherapy, etc.; sometimes, due to severe damage to the immune function, distant dissemination and metastasis occur, affecting quality of life and survival time.

Professor Pei Zhengxue, drawing on his more than forty years of clinical experience, believes that "deficiency of vital energy" is the fundamental cause of the occurrence and development of malignant tumors, and that "supporting the body and strengthening the root" is the basic principle for treating malignant tumors. When applying this principle, he takes "strengthening the spleen and tonifying the kidneys" as its essence, and thus formulated the "Lanzhou Formula." Through continuous practice, enrichment, and reorganization, it was finally developed into "Pei's Hematopoietic Granules" (composed of raw Rehmannia, Cornus fruit, Chinese yam, Prince Ginseng, Northern Sand Ginseng, Lu Dangshen, Cinnamon twig, Schisandra berry, White Peony, Floating Wheat, etc.). Years of clinical application have proven that this granule has excellent effects in treating various malignant tumors, preventing tumor recurrence and metastasis, improving patients' immune function weakened by radiotherapy and chemotherapy, enhancing patients' quality of life, and prolonging survival time. This experiment aims to scientifically and objectively study and demonstrate the anti-tumor effects and mechanisms of Pei's Hematopoietic Granules, providing sufficient scientific theoretical basis for their widespread clinical application.

I. Basis for the Research Topic

Pei's Hematopoietic Granules are a preparation developed by Professor Pei Zhengxue, a renowned expert in integrated traditional Chinese and Western medicine in China, based on his specially formulated prescription for treating leukemia—the "Lanzhou Formula"—through continuous practice, enrichment, and reorganization. It is mainly composed of raw Rehmannia, Cornus fruit, Chinese yam, Prince Ginseng, Northern Sand Ginseng, Lu Dangshen, Cinnamon twig, Schisandra berry, White Peony, Floating Wheat, etc., with heavy use of Prince Ginseng, Lu Dangshen, and Northern Sand Ginseng to strengthen the spleen; and the core ingredients of Liuwei Dihuang Tang to tonify the kidneys [16,17]. Thirty years ago, this formula was named the "Lanzhou Formula" at the Blood Disease Conference held in Suzhou in 1974 because it completely cured a patient with acute monocytic leukemia (M), Ma Changsheng. In 2003, Pei's Hematopoietic Granules successfully cured another leukemia patient (L₂), Liu Ligang, and the news was reported in the Nanchang Daily and the Gansu Daily. In 1997, this formula was designated as an internal preparation of the Gansu Provincial Academy of Medical Sciences and named "Pei's Hematopoietic Granules." Starting in 2002, this drug was used as a clinical research project of the Gansu Provincial Academy of Medical Sciences and underwent evaluation of its achievements. It won the First Prize of the Huangfu Mi Scientific and Technological Achievement Award of Gansu Province in 2003.

More than thirty years of clinical application have proven that Pei's Hematopoietic Granules (i.e., the "Lanzhou Formula") not only have excellent efficacy against leukemia but also show good effects on malignant tumors and immune dysfunction caused by radiotherapy and chemotherapy. Clinical practice has demonstrated that this formula is highly effective against esophageal cancer, liver cancer, malignant lymphoma, aplastic anemia, and other diseases. According to incomplete statistics during my time assisting the teacher in clinical practice, it has completely cured two cases of esophageal cancer, three cases of malignant lymphoma, five cases of aplastic anemia, and two cases of small liver cancer [18]. Xue [19] used the "Lanzhou Formula" to treat primary liver cancer, resulting in a significant reduction in tumor size, complete recovery of liver function, improved quality of life, and prolonged survival time; Li et al. [20] conducted an observation of the therapeutic effects of combining the Lanzhou Formula with chemotherapy in ten cancer cases, showing that the short-term efficacy of the Lanzhou Formula group was superior to that of the chemotherapy-only group (p<0.05); moreover, the toxicity and side effects of the Lanzhou Formula group were significantly milder than those of the chemotherapy-only group (p<0.01). Pei's Hematopoietic Granules were used in combination with radionuclides to treat fifteen cases of liver cancer, with the treatment group receiving radionuclides plus PG and the control group receiving radionuclides alone, revealing that the treatment group experienced significantly fewer side effects than the control group, and there was also a significant difference in therapeutic efficacy between the two groups [21].

In recent years, numerous animal experiments have proven [22-25]: Pei's Hematopoietic Granules can significantly increase the levels of hemoglobin, white blood cells, and platelets in the peripheral blood of aplastic anemia model mice (established through whole-body irradiation with a 3.0 Gy linear accelerator), effectively restoring the hematopoietic function of the bone marrow in these mice; significantly inhibit Mtp53 and increase the expression of Caspase-3 protein, inducing apoptosis of tumor cells; inhibit the expression of NF-κB, affecting gene transcription function, blocking the transition of tumor cells from G₁ phase to S phase, thereby reducing their proliferative capacity; promote the proliferative ability of splenic lymphocytes stimulated by ConA, enhance the secretion of IL-2 and the expression of IFN-γ mRNA, thus strengthening the cellular immune function of tumor-bearing mice. Pei's Hematopoietic Granules take "supporting the body and strengthening the root" as the overarching principle, focusing on overall regulation of the body, thereby improving the state of immune deficiency to varying degrees and achieving clinical efficacy. This "supporting the body and strengthening the root" approach in traditional Chinese medicine shares similarities with modern medicine's immunology, molecular biology, genomics, and other fields [10]. Although previous graduate students have already done some work in this area, there are still many aspects that require further exploration.

EGF is a powerful cell division-promoting factor closely related to tumor development [26,27]. Hepatocytes are one of the main target cells for EGF action, and in vitro studies have shown that EGF can promote the growth of liver cancer cells. The epidermal growth factor receptor (EGFR) can promote tumor cell proliferation, invasion, and metastasis, while inhibiting tumor cell apoptosis [28]. After EGF binds to EGFR, it can promote DNA synthesis in liver cancer cells through ion channels, signal transduction, and gene expression mechanisms, thereby promoting the occurrence and development of liver cancer [29]. IL-1 is one of the most potent inflammatory mediators in the body, and IL-1β (interleukin-1β, IL-1β) is an important member of the IL-1 family, with extremely low expression levels under normal physiological conditions. In recent years, it has been found that many tumor cells exhibit abnormally elevated levels of IL-1β expression, playing a crucial role in tumor growth, local invasion, metastasis, and angiogenesis [30, 31].

This experiment establishes an H₂₂ tumor-bearing mouse model to observe the effects of Pei's Shengxue Granules on serum EGF and IL-1 levels in mice, exploring the anti-tumor efficacy of Pei's Shengxue Granules and the anti-tumor mechanisms of traditional Chinese medicine from the perspective of cytokines. This aims to provide sufficient scientific theoretical basis for its widespread clinical application and lay the foundation for developing this formulation into a new type of highly effective traditional Chinese medicine preparation for the prevention and treatment of tumors.

II. Experimental Study