Study on the Induction of Apoptosis and Mechanism of Human Prostate Cancer LNCap Cells by Huangjing Agglutinin PCL-2
Prostate cancer is a malignant tumor that occurs in the male prostate tissue. In recent years, with the aging of the population and the change of diet structure, the incidence rate and mortality of prostate cancer in China have increased significantly, and will become the main disease endangering the health of men in China. The pathogenesis of prostate cancer is relatively complex. Currently, the main treatment methods for prostate cancer in clinical practice include surgery and postoperative radiation therapy. However, traditional treatment methods cannot significantly improve the five-year survival rate of prostate cancer patients. Drug therapy is mainly used for conservative treatment of patients who cannot be treated surgically or in advanced stages, and its significant adverse reactions limit the clinical treatment effect. There is currently no effective curative drug. Therefore, finding new effective drugs for the treatment of prostate cancer has become a challenge for scientific researchers.
In recent years, the search for highly efficient and low toxicity anti-cancer active ingredients from natural products has gradually become a research hotspot. The search for drugs or effective ingredients against prostate cancer has important guiding significance for clinical prevention and treatment of prostate cancer. Medicinal plants are an advantageous source of anti-tumor agents due to their wide distribution and variety of resources. Plant lectins (lectins) are a class of sugar binding specific non immunogenic proteins or glycoproteins that are widely present in various plants. They can agglutinate various cells such as red blood cells and cancer cells. As plant derived protein substances, lectins have significant inhibitory effects on tumor cell proliferation and killing of cancer cells. Their toxic and side effects on normal cells are much smaller than those of chemotherapy drugs. Developing lectins as a potential class of anti-tumor drugs has extremely broad prospects.
Huangjing is the dried rhizome of Polygonatum cyrtonema Hua, a plant in the Liliaceae family. Huangjing has a long history of medicinal use and precise therapeutic effects. It is an important dual-purpose Chinese medicine for both medicine and food, mainly used clinically to treat diseases such as yin deficiency, dry cough, spleen and stomach weakness, and soreness and weakness of the waist and knees. Huangjing should contain polysaccharides, saponins, flavonoids, anthraquinones, and amino acids. Previous studies have found that the Huangjing lectin PSL extracted from Huangjing has significant agglutination activity, which can inhibit the proliferation and Warburg effect of prostate cancer bone metastasis PC3 cells, and suppress tumor cell glycolysis by reducing the expression of HK2. At present, there are few reports on the study of Huangjing lectin on lymph node metastasis cells LNCap in prostate cancer cells. This study extracted and isolated a type of Polygonatum sibiricum lectin PCL-2 from fresh Polygonatum sibiricum medicinal materials, and investigated the effect of PCL-2 on apoptosis of human prostate cancer LNCap cells and related anti-tumor mechanisms, providing a research basis for the development of anti prostate cancer drug Polygonatum sibiricum lectin PCL-2.

 

The incidence of prostate cancer is increasing year by year in China, and it is the second largest cancer killer that causes male deaths. At present, a lot of research has been done on prostate cancer both domestically and internationally. Studies have shown that the prostate cancer LNCap cell line is widely used for studying the molecular mechanisms, drug screening, and drug evaluation of prostate cancer development and metastasis, and has become the most commonly used cell line for prostate cancer research. Huangjing lectin PCL-2 is a glycoprotein component extracted from fresh Huangjing medicinal materials. The mechanism of PCL-2 on the proliferation and apoptosis of prostate cancer LNCap cells has not been extensively studied. The results of WST-1 and colony formation experiments in this study showed that PCL-2 significantly inhibited LNCap cell proliferation in a concentration dependent manner.

Apoptosis is one of the mechanisms by which cells maintain a stable internal environment. Inducing tumor cell apoptosis has become a hot topic in anti-tumor research and an important indicator for evaluating the effectiveness of anticancer drugs. Apoptosis is regulated by multiple genes and is a complex process involving multiple pathways. ROS is an oxygen free radical and its derivatives that are directly or indirectly converted from oxygen, playing an important role in cellular signal transduction and maintaining body balance. Mitochondria are the main site of ROS production, and the accumulation of ROS causes the breakdown of mitochondrial membrane potential, triggering cell apoptosis. ROS aggregation is considered one of the earliest events in the cascade reaction of cell apoptosis. The Bcl-2 protein family is a special protein family, in which Bad, Bid, and Bax have the effect of promoting cell apoptosis, while Bcl-2 and Bcl-w have the effect of inhibiting cell apoptosis. Bcl-2 is an endometrial protein expressed in various tumor cells, which can exert anti apoptotic effects by inhibiting the release of cytochrome C, enhancing mitochondrial membrane potential, and suppressing calcium ion release; The role of Bax is opposite to that of Bcl-2. It promotes the release of cytochrome C, activates the apoptosis effector Caspase, changes cell membrane permeability, and promotes apoptosis. Studies have shown that changes in the ratio of Bcl-2 to Bax in cells can regulate apoptosis. The Caspase family is closely related to cell apoptosis and is considered a common pathway for cell apoptosis signal transduction. Among them, Caspase-3 is the executing factor of apoptosis, and the activation of this protein is a sign that cell apoptosis enters the irreversible stage. Research has found that the most common way ROS induces cancer cell apoptosis is through programmed cell death induced by the Caspases family. In addition, ROS can induce cell apoptosis by downregulating Bcl-2 and upregulating Bax and Bad. Research has found that rituximab promotes cell apoptosis by inhibiting Bcl-2 protein expression and activating the p38-MAPK signaling pathway, leading to the accumulation of ROS. This study used the 2,7-dichlorofluorescein diacetate (DCFH-DA) fluorescent probe method to measure the level of ROS in cells. The study showed that PCL-2 can significantly promote the production and accumulation of ROS in LNCap cells in a dose-dependent manner. After treatment with PCL-2, the mRNA and protein expression levels of Bax gene were upregulated and the mRNA and protein expression levels of Bcl2 gene were decreased in LNCap cells with increasing drug concentration. The protein expression levels of activated Caspase-3 and activated Caspase-9 increased. From this, it can be seen that the accumulation of ROS in LNCap cells causes changes in the levels of Bax and Bcl-2 proteins, which in turn initiate the Caspase cascade reaction, activate Caspase-9 and Caspase-3, and induce apoptosis in LNCap cells.
Huangjing is abundant in resources in China, mainly distributed in Shaanxi, Yunnan, Guizhou, Sichuan and other places, and is one of the authentic medicinal herbs in Shaanxi Province. The extraction, separation, and purification method of Huangjing lectin PCL-2 is simple and has a high yield, which can be used for large-scale sample preparation. According to the results of this study, it can be concluded that Polygonatum sibiricum lectin PCL-2 has a significant inducing effect on apoptosis of human prostate cancer LNCap cells, and has the potential to be developed as an anti prostate cancer drug. This study provides new clues for the clinical application of Polygonatum sibiricum lectin in the treatment of prostate cancer. This article only conducted preliminary exploration on the anti human prostate cancer activity and mechanism of PCL-2 at the cellular level in vitro. Its specific mechanism of action and anti-tumor activity at the animal level in vivo still need further research.