Study On The Induction Of Apoptosis In Kidney Cancer Cells By Total Flavonoids Of Herb Of Spicate Clerodendranthus Wort And Its Related Signaling Pathways

Research Background and PurposeIn human health,diseases are a serious problem,with cancer being a major issue,and kidney cancer is one type of cancer that affects people.In the treatment of cancer,surgery and drug combination therapy are commonly used methods,and this combination therapy has become a common clinical treatment method,with better efficacy and relatively reduced side effects compared to single treatment.To find a more effective drug,Cat’s Claw has been found to have good therapeutic effects on some kidney diseases and has been applied clinically.Cat’s Claw contains multiple active ingredients,with its main component being total flavonoids,which have antioxidant,anti-inflammatory,and anti-cancer effects.Recent studies have shown that total flavonoids of Cat’s Claw have a potential therapeutic effect on renal cell carcinoma,but their mechanism is still unclear.This study selected the most common human renal clear cell carcinoma 786-O cell line as a model to explore the mechanism of total flavonoids of Cat’s Claw on the apoptosis of human renal clear cell carcinoma 786-O cells,and used the following research methods for experiments to provide multiple experimental data support for clinical treatment of kidney cancer,and also provide an idea for the development of new drugs.Research methods and results（1）Effects of total flavones of Feline sphaeria on the activity of two cell linesCancer cells and normal cells were treated with different concentrations of total flavonoids from S.catina.The activity data of experimental cells were detected by CCK-8method,so as to study the changes in cell proliferation or inhibition.Meanwhile,the semi-inhibitory concentration(IC₅₀)of total flavonoids from S.catina on 786-O cells in24 h was calculated.The experimental results showed that:for normal cells,the total flavonoid of C.catoria had a promoting effect when the concentration of total flavonoid was low and the time was short,and the effect of total flavonoid of C.catoria had a significant inhibitory effect on both kinds of cells when the concentration and the time were increased.However,through the comparison of the activity of the two kinds of cells,it was found that the total flavonoid of C.catoria had a stronger inhibitory effect on kidney cancer cells.The proliferation of 786-O cells was inhibited by total flavonoids in a time and concentration dependent manner.The IC50 of total flavonoids in total flavonoids was 139.9μg·ml^–1for 24 h.（2）The effect of total flavonoids of Feline sphaerocephala on morphology of 786-O cellsThe 786-O cells were treated with different concentrations of total flavonoids in different time periods.The cell morphology was observed by inverted microscope,and the cell area was calculated by software.The results showed that the cells grew well when treated with low concentration.When drug concentration increased,cell growth was inhibited and the growth environment was affected,resulting in cell membrane rupture and cell death.At the same time,the cell area calculated by the software was also reduced to a large extent.By observing the cell death morphology under the inverted microscope,we inferred that the cell death was induced by apoptosis.（3）The effect of total flavonoids of Felicorrhiza solanum on 786-O cell deathAO/EB dye method and flow cytometry were used to detect the apoptosis of 786-O cells treated with total flavonoids at different concentrations at different time periods.It was found that nearly half of the kidney cancer cells died at a concentration of 139.9μg·ml^–1,similar to the previous IC50 results.（4）The effect of total flavonoids of Ceratophora catina on gene expression in 786-O cellsAfter the total flavonoids of S.catolaria were treated on renal carcinoma cells,transcriptome experiment was conducted,and the obtained data were analyzed at the same time,then core genes were selected for q RT-PCR verification.The results showed that the differentially expressed genes were mainly in cell cycle,apoptosis,signal transduction and other related pathways.After q RT-PCR verification,it was found that the differential gene expression trend of RNA-Seq sequencing results and verification was roughly the same,indicating that The apoptosis of renal carcinoma cells may be caused by the regulation of apoptosis-related genes by the total flavonoids of S.catoria.conclusionTotal flavonoids from Cat Whisker Grass inhibited the activity of 786-O cells and promoted their apoptosis in a concentration-and time-dependent manner.Through transcriptome sequencing,we found that total flavonoids from Cat Whisker Grass might induce apoptosis of renal cancer cells by regulating the expression of genes such as EGR1,FOS,DUSP1,RIPK1,and NOTCH3.