| ObjectiveThis paper intends to elucidate the efficacy and mechanism of Hexuetongbi formula for the clinical prevention and treatment of oxaliplatin-induced peripheral neuropathy,as w ell as to give an effective and safe treatment strategy for oxaliplatin-induced peripheral neuropathy.This paper presents high-quality evidence-based medicine evidence for the prevention and treatment of peripheral neuropathy with traditional Chinese medicine,based on clinical and mechanistic study of Hexuetongbi formula in the prevention and treatment of peripheral neuropathy.This paper not only broadens the application of the concept of"Preventive treatment of disease" in traditional Chinese medicine in the clinical treatment of malignant tumors,but also deepens the theoretical connotation of "Preventive treatment of disease" in traditional Chinese medicine and serves as a reference for methodological research in traditional Chinese medicine.Methods1.Clinical efficacy of Hexuetongbi formula in the prevention and treatment of oxaliplatin-induced peripheral neuropathyFrom September 2019 to December 2020,patients with malignant tumors who underwent chemotherapy with oxaliplatin at the First Affiliated Hospital of Guangzhou University of Traditional Chinese Medicine and Shenzhen Bao’an Hospital of Traditional Chinese Medicine were studied.In this paper,two groups were established for the goal of a randomized and open controlled study:the control group without pharmacological intervention and the Hexuetongbi formula external hand-foot washing treatment group.During the four rounds of chemotherapy,the reactions of the two groups of patients were monitored.The NCI-CTCAE scale,TNS scale,and QLQ-CIPN20 scale were used in this study to assess the incidence and severity of peripheral neuropathy in patients,as well as to assess the clinical efficacy of Hexuetongbi formula in the prevention and treatment of oxaliplatin-induced peripheral neuropathy.The information is shown as x±s frequency.and percentage.The t-test.chi-square test,and repeated measures analysis of variance were used in the study.Statistical significance was defined as P<0.05.2.Pharmacodynamic study of Hexuetongbi formula on oxaliplatin-induced peripheral neuropathy rat model18 SPF-grade SD healthy male rats were separated into six groups prior to the experiment:a blank control group,an oxaliplatin model group.and four groups of Hexuetongbi formula medicinal bath groups with varying dosages of oxaliplatin following modeling.In the Hexuetongbi formula group,four groups of rats were given varied concentrations of Hexuetongbi formula medicinal bath.The medicinal liquid concentrations were 4.25g/L,8.5g/L 17g/L.and 34g/L,and the therapy was sustained for more than three courses.The trial was halted if the medicated bath group’s behavioral test results was similar to the blank control group after any course of treatment.Every day,the rats in the normal control and model control groups were washed in the same volume of fresh water,and the temperature and timing circumstances were identical to those in the Hexuetongbi formula group.The rats’ overall health and toxic reactions were monitored throughout the trial,and behavioral tests,such as the Mechanical Withdrawal Threshold and the Cold-stimulated Paw Withdrawal Reflex test,were conducted every week.Preliminary trials identified the correct dose of Hexuetongbi formula in later experiments.Frequency and x±s are used to represent data.The analysis included one-way ANOVA and repeated measures ANOVA.Statistical significance was defined as P<0.05.Following the pre-experiment to determine the optimal concentration of the medicinal liquid,21 SD rats were used in the formal experiment and separated into three groups:normal control,oxaliplatin model,and Hexuetongbi formula medicated bath.Hexuetongbi formula medicinal bath was provided to rats in the Hexuetongbi formula group,and the treatment course was referred to the pre-experiment.Every day,the rats in the normal control and model control groups were washed in the same volume of fresh water,and the temperature and timing circumstances were identical to those in the Hexuetongbi formula group.The rats’ overall health and toxic reactions were monitored throughout the trial,and behavioral tests,such as the Mechanical Withdrawal Threshold and the Cold-stimulated Paw Withdrawal Reflex test,were conducted every week.The sensory and motor conditions of rats’ quadrupedal limbs were analyzed using various behavioral test techniques,and the therapeutic impact of Hexuetongbi formula on peripheral neuropathy was assessed.x±s and frequency are used to represent data.The analysis included one-way ANOVA and repeated measures ANOVA.Statistical significance was defined as P<0.05.Pathological sections were taken from the L4-6 segment dorsal root ganglia of the rat model used in the formal experiment.The three groups of rats’ diseased sections of the dorsal root ganglion were examined.and the morphological alterations of the rats’ dorsal root ganglion neurons were detected.The impact of the Hexuetongbi formula on the tissue and cells of the rat L4-6 dorsal root ganglion was assessed in this w ay.3.Network pharmacology study on the pharmacological effects of Hexuetongbi formulaThrough the TCMSP database,TCMID database,TCM Database@Taiwan database,and PharmGKB database,all of the chemical components of the five traditional Chinese medicines in Hexuetongbi formula were retrieved using the network pharmacology research approach.Candidate chemical components are evaluated for bioavailability(OB),drug-like characteristics(DL),and half-life once the search results are acquired(HL).TCMSP was used to find its target genes.The Genecard database and NCBI GENE can be used to find out about known target genes for the therapy of CIPN.The retrieved genes were compared to the chemical components’ screened target genes,and the intersection was used to create the "traditional Chinese medicine-target-disease" relationship network.Cytoscape v3.7.0 software was used for visualization.On the String website,the selected targets were subjected to PPI analysis of target genes,and a protein interaction network was created.For GO enrichment analysis.KEGG pathway analysis,and histogram and bubble chart making,this research employed R 3.6.3 software,the String data package was referenced,and the ClusterProfiler.Enrichplot,and Ggplot2 software packages were used.4.Molecular mechanism of Hexuetongbi formula in the treatment of oxaliplatininduced peripheral neuropathyThe tissue RNA and protein were collected from the L4-6 segment dorsal root ganglia of the rat experimental model used in the formal experiment.Quantitative Real-time PCR and Western Blot detection were used to determine gene expression in cells.The data were reported as x±s.and one-way ANOVA and repeated measures ANOVA were used to analyze them.Statistical significance was defined as P<0.05.Results1.Clinical efficacy of Hexuetongbi formula in the prevention and treatment of oxaliplatin-induced peripheral neuropathyA total of 83 individuals were included in the trial,5 of whom were later eliminated,leaving 78 patients in the final analysis.There were 39 patients in each of the control and Hexuetongbi formula treatment groups.At baseline,all 78 patients had P-values greater than 0.05,indicating no significant difference.In treatment periods 1-3 at different phases of the trial,there was no significant difference in the incidence of peripheral neuropathy between the two groups(P>0.05).There was a significant difference in the incidence between the two groups in stage 4(P<0.05).There was a significant difference in CTCAE scores between the two groups(P<0.05)according to the between-group effect analysis of NCI-CTCAE scale scores.The findings of the within-group effect analysis revealed that CTCAE ratings differed significantly at different phases(P<0.05).The interaction effect of evaluation stage and group revealed a significant difference between the treatment and control groups in the trajectory of CTCAE scores over time(P<0.05).The CTCAE score of the control group rapidly grew over time,but the treatment group’s CTCAE score climbed slowly at first and eventually stayed steady.There was no significant change in CTCAE ratings between the treatment and control groups throughout stages 1-3(P>0.05).CTCAE ratings were significantly different between the treatment and control groups at stage 4(P<0.05).The TNS scale scores between-group effect analysis revealed a significant difference in TNS ratings between the two groups(P<0.05).The findings of the within-group effect analysis revealed that TNS ratings differed significantly at different phases(P<0.05).The interaction effect of evaluation stage and group revealed a significant difference between the treatment and control groups in the trend of TNS scores over time(P<0.05).The TNS score of the control group steadily climbed over time,whereas the TNS score of the treatment group increased slowly at first and eventually declined.At stages 1 and 2,there was no significant change in TNS scores between the treatment and control groups(P>0.05).However,there was a significant difference in TNS ratings between the treatment and control groups at stages 3 and 4(P<0.05).The CIPN scores of the two groups were substantially different(P<0.05)according to the between-group effect analysis of the QLQ-CIPN20 scale scores.The findings of the within-group effect analysis revealed that CIPN scores differed significantly at different phases(P<0.05).The interaction effect of evaluation stage and group revealed that the treatment and control groups’ CIPN scores trended substantially differently over time(P<0.05).The control group’s CIPN score rapidly climbed over time,but the treatment group’s CIPN score increased slowly at first and subsequently stayed steady.At stages 1 and 2,there was no significant difference in CIPN scores between the treatment and control groups(P>0.05).The CIPN scores of the treatment group and the control group were substantially different(P<0.05)at stages 3 and 4.2.Pharmacodynamic study of Hexuetongbi formula on oxaliplatin-induced peripheral neuropathy rat modelExcept for abnormal activity of rats caused by oxaliplatin-induced peripheral neuropathy.no erythema.rash,or edema were observed on the skin of the four feet of rats exposed to Hexuetongbi formula and other parts of the skin that may have been exposed to the medicinal solution during the entire animal experimental treatment period.,Damage phenomenon.Normal skin tone is also present.The weight gain of the normal control group was the most noticeable in both studies.After the 21st day of taking oxaliplatin,the body weight of the model control group and all medicated bath groups began to decline dramatically,and the difference between them and the normal control group was statistically significant(P<0.05).The body weight of the rats in the model control group and the medicated bath group gradually increased when the use of oxaliplatin was discontinued(day 28).However,there was no statistical significance between the two trends.Blood was taken from the rats after the experiment to assess liver function,kidney function,and the myocardial enzyme spectrum.There was no harm to the liver,kidneys,or heart.The MWT of the H1 and H2 groups was still substantially different from the normal model group(P<0.05)but not statistically different from the model control group(P>0.05)in the pre-experimental Mechanical Withdrawal Threshold.The MWT of the H3 and H4 groups did not differ substantially from the normal control group(P>0.05),but it did differ significantly from the model control group(P<0.05).At the same time,no significant difference was seen between the H3 and H4 groups(P>0.05).After quitting oxaliplatin treatment,the model control group recovered slowly.Its MWT was substantially different from the usual control group towards the end of the trial(P<0.05).There was no significant difference in the number of foot withdrawals between the medicated bath groups and the normal model group in the Cold-stimulated Paw Withdrawal Reflex test(P>0.05).The H1 and H2 groups did not vary substantially from the model control group(P>0.05),however the H3 and H4 groups did differ significantly from the model control group(P<0.05).At the same time,no significant difference was seen between the H3 and H4 groups(P>0.05).After quitting oxaliplatin treatment,the model control group recovered slowly.The number of foot withdrawals was substantially different from the normal control group towards the conclusion of the trial(P<0.05).In the following formal studies,the concentration of 17g/L liquid medication was eventually found.The medicated bath group’s MWT was not substantially different from the normal control group(P>0.05),but it was significantly different from the model control group(P<0.05)in a formal experiment of Mechanical Withdrawal Threshold.Following the discontinuation of oxaliplatin,the model control group recovered slowly.Its MWT differed considerably from the other two groups towards the conclusion of the trial(P<0.05).The medicated bath group did not vary signficantly from the normal control group(P>0.05),but did differ significantly from the model control group(P<0.05).After quitting oxaliplatin,it was still difficult to recover in the model control group,and the number of foots did not drop much.The results were substantially different(P<0.05)from the other two groups towards the conclusion of the trial.The neurons in the normal control group and the Hexuetongbi formula medicinal bath group were plentiful and consistently organized in the pathological trials,with distinct nuclear and cytoplasmic borders,evident nucleoli,and no visible inflammation.More neuronal cell bodies swelled in the model control group,the area around the cells grew larger under a 20x microscope,and many neuronal cell bodies and nuclei vanished after cytoplasmic lysis.Furthermore,the number of satellite cells surrounding the neuron cell body reduced,and more nerve fibers became demyelinated.3.Network pharmacology study on the pharmacological effects of Hexuetongbi formulaThe TCMSP database produced a total of 665 chemical components.Finally,27 active components were evaluated under the restricted circumstances.A total of 399 targets matching to each of the tested 27 active components were collected from the TCMSP database again.There were 141 targets after common targets were removed.The Genecard database yielded a total of 375 target genes associated to CIPN.The intersection was found by comparing the retrieved genes to the 141 target genes of active chemical compounds that had been tested.Hexuetongbi formula yielded a total of 35 targets for the therapy of CIPN.Following a PPI analysis of 35 protein targets via the String website,the top 20 protein targets with the strongest correlation in the network were selected out for GO enrichment analysis and KEGG pathway analysis,based on the degree of each target from high to low.The genes enriched by BP analysis included response to lipopolysaccharide,response to molecule of bacterial origin,response to steroid hormone,and so on,according to the results of GO enrichment analysis.Membrane raft,membrane microdomain,and membrane area are the most enriched genes in CC analysis.A vast number of enriched genes were discovered by MF analysis,including cytokine receptor binding,steroid binding,tumor necrosis factor receptor superfamily binding,and so on.The highest enrichment and association were found in the AGE-RAGE signaling route in diabetic complications,TNF signaling pathway,C-type lectin receptor signaling pathway,and Apoptosis,according to KEGG pathway enrichment analysis.The three pathways of PI3K-Akt、JNK/MAPK、NF-κB/COX-2(PTGS2)may be most closely associated to the therapy of CIPN with Hexuetongbi formula,according to the most relevant targets in the protein interaction network.4.Molecular mechanism of Hexuetongbi formula in the treatment of peripheral neuropathy caused by oxaliplatinThe mRNA expressions of PI3K,Aktl,Akt2,Akt3,MAPK8,PTGS2,and Bcl-2 were all up-regulated,with significant differences(P<0.05),according to QPCR data.Despite the upregulation of MAPK8,PTGS2,and Akt2,no significant difference was found between the Hexuetongbi formula medicinal bath group and the model control group(P>0.05).PI3K,Akt1,Akt3,and Bcl-2 mRNA expressions in the Hexuetongbi formula group,however,were substantially different from those in the model control group(P<0.05).The expression levels of PI3K,Akt1(phospho S473),and Bcl-2 in the Hexuetongbi formula medicated bath group were all up-regulated,with significant differences(P<0.05),according to Western blot detection findings.The change was statistically significant(P<0.05).ConclusionMultiple sessions of oxaliplatin chemotherapy can produce peripheral neuropathy,which can be reduced by using the Hexuetongbi formula.The symptoms of peripheral neuropathy that has already developed might be suppressed or eased.Furthermore,it has strong preventative and therapeutic benefits on oxaliplatin-induced peripheral neuropathy.It has the therapeutic effect of reducing pain and tactile hypersensitivity in peripheral nerves.The mechanism of action may be to ameliorate oxaliplatin-induced peripheral neuropathy by inhibiting death of neurons in the dorsal root ganglia by modulating the expression of the PI3K/Akt pathway and its downstream Bcl-2 protein.It can stimulate nerve cell healing and postpone the progression of lesions even in the situation of damaged neuron cells. |
PDF Full Text Request