| Background and PurposeSyndrome differentiation and treatment is the basic principle of TCM(Traditional Chinese Medicine)on understanding and treatment of diseases.Among them,the coldness,heat,warmth,and coldness of TCM is one of the important basis for the treatment of diseases.We conducted statistical analysis of the commonly used anti-tumor TCM,and found that most of the anti-tumor medicines own obviously cold/hot properties.However,there are many controversies about whether cold/hot compounds promote or inhibit tumor metastasis.Is it because the cold/hot compounds act on the corresponding sensory receptors on the tumor cells,thereby causing different effects on the biological behavior of the tumor cells?Thermo-TRPs(Thermo-Transient Receptor Potential Channels)play important role in sensing temperature changes.Thermo-TRPs include a total of 11 members including hot-sensitive receptors TRPV1-4,TRPM2-5,cold-sensitive receptors TRPM8,TRPC5 and TRPA1.Thermo-TRPs can sense and respond to temperature changes,and cold/hot compounds can affect the body's perception of temperature.In addition,many cold/hot compounds have been reported to have a certain regulatory effect on Thermo-TRPs,suggesting that TRPs may be a common target of cold/hot compounds?We analyzed the expression levels of Thermo-TRPs in different cancers and found that Thermo-TRPs clustered among different tumors.In breast and skin cancer,the expression of hot-sensitive receptors increased significantly;while in colon,liver and gastric cancer,cold-sensitive receptors showed high expression.Based on the analysis results,we tested Thermo-TRPs protein expression levels of various cancers and found that the cold-sensitive receptor TRPC5 was significantly overexpressed in gastric cancer.Malignant gastric cancer proliferates quickly and is highly invasive,and metastasis is an important cause of death in patients with gastric cancer.Preliminary analysis and preliminary exploration of experiments have shown that TRPC5 is significantly expressed in gastric cancer.Whether TRPC5 can be used as a therapeutic target for gastric cancer,and effective compounds can be selected from TCM to inhibit the progress of gastric cancer metastasis?Therefore,through bioinformatics analysis combined with a series of in vivo and in vitro experiments,we verified the regulatory effect of kaempferol on TRPC5,as well as the molecular mechanism and biological basis of gastric cancer metastasis inhibition.Research Contents and ResultsIn the first part of the experimental study,bioinformatics analysis and analysis of clinical gastric cancer samples revealed that TRPC5 was highly expressed in patients with gastric cancer,and its expression level was negatively correlated with the survival time of patients.Further research atthe cellular level has shown that the expression level of TRPC5 in a variety of gastric cancer cells is higher than that of normal gastric epithelial cells,and its expression level is positively correlated with the calcium influx mediated by it.The second part of the experimental study of this subject comprehensively analyzed transwell,inverted invasion,scratches and other in vitro experiments and in vivo metastasis experiments to verify that the high expression of TRPC5 can promote the metastasis of gastric cancer,and inhibit its function and expression can inhibit gastric cancer metastasis.The third part of the experimental study of this subject first verified that kaempferol can inhibit the opening of TRPC5 and its calcium influx through binding with TRPC5 through calcium imaging and drug protein binding experiments.Further in vitro and in vivo metastasis experiments,including transwell,inverted invasion and scratches,have verified that kaempferol can significantly inhibit the metastasis of gastric cancer both in vivo and in vitro.The fourth part of the experimental research of this subject first combines database analysis and live cell imaging experiments to find that the function of TRPC5 is closely related to cell motility.Based on this,further database analysis and literature research have found that the regulation of gastric cancer cell motility by TRPC5 may be closely related to the formation of filopodia,and the experimental results also show that TRPC5 regulates the formation of filopodia of gastric cancer cells.Regulates its mobility and affects the metastasis of gastric cancer.The last part of the experimental research in this topic is to deeply explore and explore the mechanism of TRPC5 through various molecular biological methods and bioinformatics analysis.The experimental results show that the activation of TRPC5 channel can promote the formation of intracellular Ca2+-CaM complex,and then promote the activation of MLC.The activation of MLC can promote the activation of cortactin,positioning it at the tip of the filopodia,thereby enhancing the stability of the filopodia.In general,TRPC5 regulates the formation of filopodia through the p-MLC/ATP/p-cortactin signal axis,which affects the mobility of gastric cancer cells and their metastatic process.Conclusion and SignificanceThe study combined bioinformatics analysis,clinical samples,and multiple experiments in vivo and in vitro confirmed that the expression of TRPC5 was negatively correlated with the occurrence and development of gastric cancer from multiple perspectives,suggesting that TRPC5 may be a potential target for the treatment of gastric cancer.Further research on the mechanism revealed that TRPC5 can regulate the formation of filopodia in gastric cancer through p-MLC/ATP/p-cortactin signal axis,reveal the specific mechanism of filopodia in promoting gastric cancer metastasis,and provide a new mechanism for gastric cancer metastasis.Treatment strategy.Discover the active ingredient kaempferol in a variety of traditional anti-tumor Chinese medicines,and verify its anti-gastric cancer metastasis effect and mechanism,providing a basis for the development of clinical anti-gastric cancer metastatic drugs. |
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