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1.
Journal of Prevention and Treatment for Stomatological Diseases ; (12): 178-187, 2024.
Article in Chinese | WPRIM | ID: wpr-1006519

ABSTRACT

Objective@#To explore the molecular mechanism of resveratrol (RES) in the treatment of oral squamous cell carcinoma (OSCC) through the use of biological information methods such as network pharmacology and molecular docking and to provide a theoretical reference for the clinical application of RES in the treatment of OSCC.@*Methods@#The Swiss Target Prediction(http://www.swisstargetprediction.ch), SEA (http://sea.bkslab.org)database, and Pharm mapper database(http://lilab-ecust.cn) were used to retrieve RES-related targets, and the DISGENET (www.disgenet.org), OMIM (https://omim.org) and GeneCards (https://www.genecards.org) databases were used to screen OSCC disease targets. The intersection of drugs and disease targets was determined, and Cytoscape 3.7.2 software was used to construct a "drug-diseasetarget pathway" network. The Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) database was used to construct a target protein interaction network, and the DAVID database was used for enrichment analysis of key proteins. Finally, molecular docking validation of key proteins was performed using AutoDock and PyMOL. The enrichment analysis and molecular docking results were integrated to predict the possible molecular mechanisms of RES treatment in OSCC; western blot was used to determine the effect of resveratrol at different concentrations (50, 100) μmol/L on the expression of Src tyrosine kinase (SRC), epidermal growth factor receptor (EGFR), estrogen receptor gene 1 (ESR1), and phosphatidylinositol 3 kinase/protein kinase B (PI3K/AKT) signaling pathway proteins in OSCC HSC-3 cells.@*Results@#A total of 243 targets of RES drugs and 6 094 targets of OSCC were identified. A total of 116 potential common targets were obtained by intersecting drugs with disease targets. These potential targets mainly participate in biological processes such as in vivo protein self-phosphorylation, peptide tyrosine phosphorylation, transmembrane receptor protein tyrosine kinase signaling pathway, and positive regulation of RNA polymerase Ⅱ promoter transcription, and they interfere with the PI3K/AKT signaling pathway to exert anti-OSCC effects. The docking results of resveratrol with OSCC molecules indicated that key targets, such as EGFR, ESR1, and SRC, have good binding activity. The results of cell-based experiments showed that resveratrol inhibited the protein expression of SRC, EGFR, ESR1, p-PI3K, and p-AKT in HSC-3 cells in a dose-dependent manner.@*Conclusion@#RES can inhibit the expression of its targets EGFR, ESR1, SRC, p-PI3K, and p-AKT in OSCC cells.

2.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 124-132, 2024.
Article in Chinese | WPRIM | ID: wpr-1003774

ABSTRACT

ObjectiveTo analyze the antidepressant quality markers(Q-Marker) of Bupleuri Radix(BP) before and after vinegar-processing by ultra-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry(UPLC-Q-TOF-MS), multivariate statistical analysis and network pharmacology. MethodUPLC-Q-TOF-MS was used to analyze the chemical basis of raw and vinegar-processed products of BP, and principal component analysis(PCA) orthogonal partial least squares-discriminant analysis(OPLS-DA) were used to identify the differential components in BP that changed significantly before and after vinegar-processing, which were regarded as candidate quality markers(Q-Marker). Then the disease-drug-component-target network related to antidepressant effect of BP was constructed by network pharmacology, and the antidepressant Q-Marker of raw and vinegar-processed products of BP was determined. Rats were randomly divided into blank group, model group, fluoxetine group(2.67 mg·kg-1) and total saponin group(0.72 mg·kg-1), except the blank group, rats in the other groups were subjected to chronic unpredictable mild stress(CUMS). Three weeks after the start of modeling, rats in each administration group were given the corresponding dose of drugs once a day for 4 weeks, and rats in the blank and model groups were given normal saline with dose of 10 mL·kg-1. At 1 day before modeling, 21 days and 28 days after administration, body mass weighing, sucrose preference test and open field test were performed on each group . After 28 days of administration, real-time fluorescence quantitative polymerase chain reaction(Real-time PCR) was used to detect the mRNA expression levels of phosphatidylinositol 3-kinase(PI3K), protein kinase B(Akt), mammalian target of rapamycin(mTOR), glycogen synthase kinase-3β(GSK-3β), forkhead box transcription factor O3a(FoxO3a) and β-catenin in hippocampal tissues of rats in each group, while protein expression levels of PI3K, Akt, mTOR and FoxO3a in hippocampal tissues of rats in each group were detected by Western blot. ResultThere were 19 components in BP showed significant changes before and after vinegar-processing, and 9 components such as saikosaponin A, saikosaponin B1, saikosaponin B2, saikosaponin C and saikosaponin D were identified as potential Q-Marker through S-plot differential marker screening. Combined with the disease-drug-component-target network, saikosaponin A, saikosaponin B1, saikosaponin B2 and saikosaponin D were identified as antidepressant Q-Marker of raw and vinegar-processed products of BP. According to the results of pharmacodynamic tests, after 28 d of administration, compared with the blank group, the body mass, sucrose preference index and open field total score of rats in model group, fluoxetine group and total saponin group decreased significantly(P<0.01). Compared with the model group, the body mass, sucrose preference index and open field total score in total saponin group increased significantly(P<0.01). Compared with the blank group, mRNA expression levels of PI3K, Akt, mTOR and β-catenin in hippocampus of rats in the model group decreased significantly(P<0.05), while mRNA expression levels of GSK-3β and FoxO3a increased significantly(P<0.05). Compared with the model group, mRNA expression levels of PI3K, Akt, mTOR and β-catenin in hippocampus of rats in the total saponin group were increased significantly(P<0.05), while mRNA expression levels of GSK-3β and FoxO3a decreased significantly(P<0.05). Compared with the blank group, the protein expression levels of Akt and mTOR in hippocampus of the model group decreased significantly(P<0.01), while the protein expression levels of PI3K and FoxO3a increased significantly(P<0.01). Compared with the model group, the expression level of Akt in hippocampus of the total saponin group increased significantly(P<0.01), the mTOR expression level was increased but not statistically significant, while the protein expression levels of PI3K and FoxO3a decreased significantly(P<0.01). ConclusionThe chemical constituents of BP changed greatly after vinegar-processing, and the antidepressant Q-Marker of raw and vinegar-processed products of BP was determined by chemical basis, pharmacodynamics, network pharmacology and signaling pathway, which provided a reference for further research on quality control, pharmacodynamic substance basis and processing mechanism of BP.

3.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 10-17, 2024.
Article in Chinese | WPRIM | ID: wpr-1003761

ABSTRACT

ObjectiveTo observe the therapeutic effect of Qiwei Baizhusan(QWBZS) on diabetic encephalopathy(DE) rat model, and to explore the possible mechanism of QWBZS in the treatment of DE based on phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)/glycogen synthase kinase-3β(GSK-3β) signaling pathway. MethodForty-eight SPF male Wistar rats were randomly divided into blank group(8 rats) and high-fat diet group(40 rats). After 12 weeks of feeding, rats in the high-fat diet group were intraperitoneally injected with 35 mg·kg-1 of 1% streptozotocin(STZ) for 2 consecutive days to construct a DE model, and rats in the blank group were injected with the same amount of sodium citrate buffer. After successful modeling, according to blood glucose and body weight, model rats were randomly divided into model group, low, medium and high dose groups of QWBZS(3.15, 6.3, 12.6 g·kg-1), combined western medicine group(metformin+rosiglitazone, 0.21 g·kg-1), with 6 rats in each group. The administration group was given the corresponding dose of drug by gavage, and the blank group and the model group were given an equal volume of 0.9% sodium chloride solution by gavage, 1 time/day for 6 weeks. Morris water maze was used to detect the spatial memory ability of DE rats. Fasting insulin (FINS) level was detected by enzyme-linked immunosorbent assay(ELISA) and insulin resistance index(HOMA-IR) was calculated. Hematoxylin-eosin(HE) staining was used to observe the morphological changes of hippocampus in rats, ELISA was used to detect the indexes of oxidative stress in hippocampal tissues, real-time fluorescence quantitative polymerase chain reaction(Real-time PCR) was used to detect mRNA expression levels of PI3K, Akt, nuclear transcription factor-κB(NF-κB), tumor necrosis factor-α(TNF-α) and interleukin-1β(IL-1β) in hippocampus, and Western blot was used to detect the protein expression of PI3K, Akt, phosphorylated(p)-Akt, GSK-3β and p-GSK-3β in hippocampus of rats. ResultCompared with the blank group, FINS and HOMA-IR values of the model group were significantly increased(P<0.01), the path of finding the original position of the platform was significantly increased, and the escape latency was significantly prolonged(P<0.01), the morphology of neuronal cells in hippocampal tissues was disrupted, the levels of reactive oxygen species(ROS) and malondialdehyde(MDA) in hippocampus of rats were increased, and the activity of superoxide dismutase(SOD) was decreased(P<0.05, P<0.01), mRNA expression levels of PI3K and Akt were decreased(P<0.01), mRNA expression levels of NF-κB, TNF-α and IL-1β were increased(P<0.05, P<0.01), the protein expression levels of PI3K, p-Akt and p-GSK-3β were significantly decreased, and the protein expression of GSK-3β was significantly increased(P<0.01). Compared with the model group, the FINS and HOMA-IR values of the medium dose group of QWBZS and the combined western medicine group were significantly decreased(P<0.01), the path of finding the original position of the platform and the escape latency were significantly shortened(P<0.01), the hippocampal tissue structure of rats was gradually recovered, and the morphological damage of nerve cells was significantly improved, the contents of ROS and MDA in hippocampus of rats decreased and the level of SOD increased(P<0.01), the mRNA expression levels of PI3K and Akt were increased(P<0.01), and the mRNA expression levels of NF-κB, TNF-α and IL-1β were decreased (P<0.05, P<0.01), the protein expression levels of PI3K, p-Akt and p-GSK-3β were significantly increased(P<0.01), and the expression of GSK-3β was significantly decreased(P<0.01). ConclusionQWBZS can alleviate insulin resistance in DE rats, it may repair hippocampal neuronal damage and improve learning and cognitive ability of DE rats by activating PI3K/Akt/GSK-3β signaling pathway.

4.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 118-126, 2024.
Article in Chinese | WPRIM | ID: wpr-1003415

ABSTRACT

ObjectiveTo observe the effect of earthworm protein on the expression of phosphatidylinositol 3-kinase/protein kinase B/nuclear factor E2-related factor 2 (PI3K/Akt/Nrf2) pathway in the aorta of spontaneously hypertensive rats (SHR) and explore mechanism of earthworm protein in treating hypertensive vascular endothelial dysfunction (VED). MethodTen 10-week-old Wistar Kyoto (WKY) rats and fifty SHR rats were selected for a week of adaptive feeding. WKY rats were selected as the normal group, and fifty SHR rats were randomized according to body weight into model, valsartan (8×10-3 g·kg-1·d-1), and high-, medium-, and low-dose (0.2, 0.1, 0.05 g·kg-1·d-1, respectively) earthworm protein groups. The normal and model groups were administrated with equal volume of double distilled water by gavage. During the drug intervention period, the general situations of rats in each group were observed and their blood pressure was monitored at specific time points every other week before and after administration. After 8 weeks of drug intervention, enzyme-linked immunosorbent assay was employed to measure the levels of angiotensin-Ⅱ (Ang-Ⅱ) and endothelin-1 (ET-1) in the serum of rats in each group. The corresponding kits were used to determine the levels of nitric oxide (NO), malondialdehyde (MDA), glutathione peroxidase (GPX), superoxide dismutase (SOD), and ferrous ion (Fe2+). Hematoxylin-eosin (HE) staining was employed to observe the changes in the intima of the aorta. Fluorescence quantitative polymerase chain reaction (Real-time PCR) was employed to measure the mRNA levels of PI3K, Akt, Nrf2, heme oxygenase-1 (HO-1), and glutathione peroxidase 4 (GPX4) in the aortic tissue. Western blotting was used to determine the protein levels of p-PI3K (Tyr467/199), PI3K, p-Akt (Ser473), Akt, Nrf2, HO-1, and GPX4 in the thoracic aorta. ResultCompared with the normal group, the model group had decreased body mass, increased irritability, severe endothelial damage, elevated blood pressure and serum levels of Ang-Ⅱ, ET1, MDA, and Fe2+ (P<0.01), lowered NO level (P<0.01), and down-regulated mRNA and protein levels of p-PI3K (Tyr467/199), PI3K, p-Akt (Ser473), Akt, Nrf2, HO-1, and GPX4 in the aortic tissue (P<0.01). Compared with the model group, drug intervention caused no significant change in the body mass, calmed the rats, alleviated the endothelial damage, lowered blood pressure and serum levels of Ang-Ⅱ, ET1, MDA, and Fe2+ (P<0.01), elevated the NO level (P<0.05), and up-regulated the mRNA and protein levels of p-PI3K (Tyr467/199), PI3K, p-Akt (Ser473), Akt, Nrf2, HO-1, and GPX4 (P<0.05). ConclusionThe earthworm protein can exert antihypertensive effects by ameliorating VED in SHR. Specifically, it may regulate the PI3K/Akt/Nrf2 signaling pathway to inhibit oxidative stress and ferroptosis.

5.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 99-109, 2024.
Article in Chinese | WPRIM | ID: wpr-1003413

ABSTRACT

ObjectiveTo investigate the effect of Tangzhi pills on the improvement of insulin resistance (IR) in the liver with type 2 diabetes (T2DM) by regulating phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway based on differential genes and its possible molecular mechanism. MethodT2DM rat models were prepared by high fat (HFD) diet combined with streptozotocin (STZ) intraperitoneal injection. The experiment was divided into blank group, model group, metformin hydrochloride group (0.18 g·kg-1), Tangzhi pills high (1.08 g·kg-1), medium (0.54 g·kg-1) and low (0.27 g·kg-1) dose groups. Rat serum, liver, and pancreatic tissue were collected, and the pathological tissue of the liver and pancreas was observed using hematoxylin-eosin (HE) staining. The fasting blood glucose level (FBG) was detected, and oral glucose tolerance (OGTT) tests were conducted. Enzyme-linked immunosorbent assay (ELISA) was used to detect fasting serum insulin (FINS) and glycated hemoglobin (GHb) levels in rats. IR homeostasis model index (HOMA-IR), β cellular homeostasis index (HOMA-β), and insulin sensitivity index (ISI) were calculated. Biochemical methods were used to determine the levels of triglyceride (TG), total cholesterol (TC), low-density lipoprotein (LDL-C), and high-density lipoprotein (HDL-C) in rat serum. Transcriptomics obtained differentially expressed mRNA from liver tissue and enriched differentially expressed pathways. Real-time reverse transcriptase polymerase chain reaction (Real-time PCR) was used to detect the mRNA expression of cyclic adenylate responsive element binding protein 3-like protein 2 antibody (CREB3l2), B-lymphocyte tumor 2 (Bcl-2), Toll-like receptor 2 (TLR2), cyclin-dependent kinase inhibitor 1A (CDNK1A), and DNA damage induced transcription factor 4-like protein (DDIT4) in liver tissue. Western blot was used to detect the protein expression of phosphorylated phosphatidylinositol 3-kinase (p-PI3K), phosphorylated protein kinase B (p-Akt), glucose transporter 4 (GLUT4), insulin receptor (INSR), and insulin receptor substrate 2 (IRS2). ResultThe pharmacodynamic experiment results showed that compared with model group, Tangzhi pills groups repaired liver and pancreatic tissue to varying degrees, reduced blood sugar (P<0.01), and promoted a decrease in serum FINS, GHb, and HOMA-IR (P<0.05, P<0.01). In addition, HOMA-β and ISI increased (P<0.05, P<0.01). The levels of TC, TG, and LDL-C decreased (P<0.05, P<0.01), while the levels of HDL-C increased (P<0.05, P<0.01). The transcriptomics experimental results confirmed that the PI3K/Akt signaling pathway was significantly expressed in both the blank group and model group, as well as in the high-dose Tangzhi pills group and model group. CDNK1A, DDIT4, CREB3l2, Bcl-2, and TLR2 were significantly differentially expressed mRNA during TG intervention in T2DM. Compared with the model group, the protein expression of p-PI3K, p-Akt, GLUT4, INSR, and IRS2 increased in all Tangzhi pills groups (P<0.01). The mRNA expression of CREB3l2, Bcl-2, and TLR2 increased (P<0.01), while that of CDNK1A and DDIT4 decreased (P<0.01). ConclusionTangzhi pills may regulate the PI3K/Akt signaling pathway based on the differential mRNA expression of CREB3l2, Bcl-2, TLR2, CDNK1A, and DDIT4, thereby improving IR in the liver with T2DM.

6.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 130-139, 2024.
Article in Chinese | WPRIM | ID: wpr-999169

ABSTRACT

ObjectiveTo observe the effects of Hedysari Radix polysaccharide on the apoptosis of gastric sinus smooth muscle cells and explore the underlying mechanism via the insulin-like growth factor-1 (IGF-1)/phosphatidylinositol 3-kinase (PI3K)/serine-threonine kinase (Akt) pathway in the rat model of diabetic gastroparesis (DGP). MethodSixty-two Wistar male rats were randomized into a blank group (n=12) and a modelling group (n=50). The rat model of DGP was established by small-dose multiple intraperitoneal injections of streptozotocin combined with an irregular high-fat and high-sugar diet for 4 weeks. The modeled rats were randomized into model group, mosapride citrate (1.35 mg·kg-1), and high-, medium-, and low-dose (200, 100, and 50 mg·kg-1, respectively) Hedysari Radix polysaccharide groups. The rats were administrated with corresponding drugs by gavage, and those in the blank and model groups with equal volumes of pure water by gavage once a day for 8 consecutive weeks. The random blood glucose and body mass were measured every 2 weeks, and gastric emptying rate was calculated. Hematoxylin-eosin (HE) staining was used to observe the pathological changes of smooth muscle in gastric antrum, and terminal deoxynucleoitidyl transferase-mediated nick-end labeling (TUNEL) was employed to detect the apoptosis of smooth muscle cells in the gastric antrum. The expression of IGF-1, phosphorylated (p)-PI3K, and p-Akt in the smooth muscle of gastric sinus tissue was detected by immunohistochemistry. Western blot was employed to determine the protein levels of IGF-1, p-PI3K/PI3K, p-Akt/Akt, B-cell lymphoma-2 (Bcl-2), and Bcl-2-associated X protein (Bax) in the smooth muscle of the gastric antrum. ResultCompared with the blank group, the model group showed elevated random blood glucose at all time points (P<0.01), decreased body mass and gastric emptying rate (P<0.01), increased apoptotic index of smooth muscle cells in the gastric antrum (P<0.01), down-regulated protein levels of IGF-1, p-PI3K/PI3K, p-Akt/Akt, and Bcl-2, and up-regulated protein level of Bax (P<0.01). Compared with the model group, the 8 weeks of drug administration lowered the random blood glucose, increased the body mass and gastric emptying rate (P<0.05, P<0.01), decreased the apoptotic index of smooth muscle cells in the gastric antrum (P<0.05, P<0.01), up-regulated the protein levels of IGF-1, p-PI3K/PI3K, p-Akt/Akt, and Bcl-2, and down-regulated the protein level of Bax (P<0.05, P<0.01). Compared with the mosapride citrate group,the administration of low-dose Hedysari Radix polysaccharide for 6 and 8 weeks lowered the random blood glucose and decreased the body mass (P<0.05, P<0.01),low and medium-dose Hedysari Radix polysaccharide decreased the gastric emptying rate and the apoptotic index of smooth muscle cells in the astragaloside low-dose group decreased (P<0.05). The protein levels of IGF-1,p-PI3K/PI3K,p-Akt/Akt and Bcl-2(low dose)were down-regulated and the protein level of Bax was up-regulated by low doses of Hedysari Radix polysaccharide (P<0.05, P<0.01). Compared with high-dose Hedysari Radix polysaccharide, low-dose Hedysari Radix polysaccharide elevated random blood glucose and reduced body mass after 6 and 8 weeks of administration (P<0.05, P<0.01), and the low and medium doses decreased the gastric emptying rate, increased the apoptotic index of smooth muscle cells in the gastric antrum (P<0.05, P<0.01), down-regulated the protein levels of IGF-1, p-PI3K/PI3K, p-Akt/Akt, and Bcl-2, and up-regulated the protein level of Bax (P<0.05, P<0.01). Compared with the medium-dose group,the low-dose group of Hedysari Radix polysaccharide had lower body mass,lower gastric emptying rate in rats,higher apoptotic index of smooth muscle cells in gastric sinus tissue after 6 and 8 weeks of administration (P<0.05, P<0.01), and lower protein expression of IGF-1,p-PI3K/PI3K,p-Akt/Akt. ConclusionHedysari Radix polysaccharide protects the smooth muscle cells in gastric antrum against apoptotic injury and promotes gastric motility by activating the IGF-1/PI3K/Akt signaling pathway, as manifested by the up-regulated expression of IGF-1, p-PI3K, p-Akt, and Bcl-2 and down-regulated expression of Bax.

7.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 87-94, 2024.
Article in Chinese | WPRIM | ID: wpr-999164

ABSTRACT

ObjectiveTo establish a mouse model of basilar artery dolichoectasia (BAD) and explore the mechanism of modified Tongqiao Huoxuetang (JTQHX) in regulating BAD via phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) pathway. MethodSixty C57/BL6 female mice were randomized into sham operation (injected with 10 U·mL-1 inactivate elastase), model, atorvastatin calcium tablets (2.6 mg·kg·d-1), and low- and high-dose (crude drug 3.4, 17 g·kg-1·d-1, respectively) JTQHX groups. The mouse model of BAD was established by injection with 10 U·mL-1 elastase. After 14 days of modeling, the sham operation group and model group were administrated with equal volumes of pure water by gavage, and other groups with corresponding drugs for 2 months. The levels of interleukin-6 (IL-6) and calpain (LpA) in the serum were measured by enzyme-linked immunosorbent assay (ELISA). Verhoeff 's Van Gieson (EVG) staining was employed to observe the pathological changes of blood vessels. Terminal-deoxynucleotidyl transferase mediated nick end labeling (TUNEL) was employed to examine the apoptosis rate of vascular smooth muscle cells (VSMCs). Image Pro Plus was used to observe and calculate the curvature index, elongation length, percentage increase in vessel diameter, and curvature angle of the basilar artery vessels in mice. Western blot was employed to determine the expression levels of PI3K and Akt in the vascular tissue. ResultCompared with the sham operation group, the model group showed lowered IL-6 level (P<0.01), no significant change in LpA level, increased apoptosis of VSMCs (P<0.01), and increased curvature index, elongation length, percentage increase in vessel diameter, and curvature angle (P<0.01). Furthermore, the modeling up-regulated the protein levels of PI3K and Akt in blood vessels (P<0.01) and aggravated the destruction of the inner elastic layer, atrophy of the muscular layer, and hyaline changes in the connective tissue of the medial membrane of the basilar artery wall. Compared with the model group, 2 months of treatment with JTQHX elevated the IL-6 level (P<0.01), reduced the apoptosis of VSMCs (P<0.01), decreased the curvature index, elongation length, percentage increase in vessel diameter, and curvature angle (P<0.05, P<0.01), and down-regulated the protein levels of PI3K and Akt in blood vessels (P<0.01). In addition, the treatment alleviated the destruction of the inner elastic layer, atrophy of the muscular layer, and hyaline changes in the connective tissue of the medial membrane of the basilar artery wall. ConclusionJTQHX inhibits the elongation, expansion, and curvature of basilar artery vessels and alleviates the pathological changes by reducing the apoptosis of VSMCs and down-regulating the expression of PI3K/Akt pathway.

8.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 42-51, 2023.
Article in Chinese | WPRIM | ID: wpr-984582

ABSTRACT

ObjectiveTo explore the mechanisms of internal treatment (Renshen Baidusan), external treatment (Yurui Enema), and combination of the two methods in treating intestinal mucosal injury in the rat model of ulcerative colitis (UC) from the changes of phosphatidylinositol-3 kinase (PI3K)/protein kinase B (Akt)/nuclear factor-κB (NF-κB) pathway. MethodFifty SPF-grade SD rats were randomized into blank, model, Renshen Baidusan (15.6 g·kg-1), Yurui Enema (25 g·kg-1), and combined treatment (15.6 g·kg-1 Renshen Baidusan + 25 g·kg-1 Yurui Enema) groups (n=10). The rat model of UC was established in other groups except the blank group by 2,4, 6-trinitrosulfonic acid (TNBS)/ethanol. The rats were administered with corresponding drugs once a day for 14 consecutive days since the 8th day after modeling. The histopathological changes of colon were observed by hematoxylin-eosin (HE) staining. Enzyme-linked immunosorbent assay (ELISA) was employed to measure the levels of tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-4, and IL-10 in the colon tissue. The apoptosis of colon epithelial cells was detected by terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL). The location and expression of B-cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein (Bax), TNF-α, and IL-6 in the colon tissue were examined by immunohistochemistry. Real-time quantitative fluorescence polymerase chain reaction (Real-time PCR) and Western blot were employed to determine the mRNA and protein levels, respectively, of the proteins in the PI3K/Akt/NF-κB pathway in the colon tissue. ResultIn the model group, HE staining showed a large number of inflammatory cell infiltration in the mucosa and submucosa. Compared with the blank group, the model group showed elevated levels of TNF-α and IFN-γ and lowered levels of IL-4 and IL-10 in the colon tissue, increased apoptosis rate of colon epithelial cells, increased positive expression of Bax, TNF-α, and IL-6, and decreased positive expression of Bcl-2 (P<0.05). Moreover, the model group showed up-regulated mRNA levels of PI3K, Akt, and NF-κB and protein levels of PI3K, p-PI3K, Akt, p-Akt, p65, p-p65, Bax, and cleaved Caspase-3, increased Bax/Bcl-2 and cleaved Caspase-3/Caspase-3 ratios, and down-regulated protein levels of NF-κB suppressor protein α(IκBα), Bcl-2, and Caspase-3 in the colon tissue (P<0.05). Compared with the model group, the internal treatment, the external treatment, and the combination (referred to as the three groups) alleviated the colonic mucosal injury, lowered the levels of TNF-α and IFN-γ and elevated the levels of IL-4 and IL-10 in the colon tissue, decreased the apoptosis rate of colon cells, inhibited the positive expression of Bax, TNF-α, and IL-6, and promoted the positive expression of Bcl-2 (P<0.05). Furthermore, the combination group down-regulated the mRNA level of PI3K (P<0.05). The three groups down-regulated the mRNA levels of Akt and NF-κB and the protein levels of p-PI3K, Akt, p-Akt, p65, p-p65, Bax, and cleaved Caspase-3 in the colon tissue, decreased the Bax/Bcl-2 and cleaved Caspase-3/Caspase-3 ratios, and up-regulated the protein levels of IκBα, Bcl-2, and Caspase-3 (P<0.05). ConclusionRenshen Baidusan, Yurui Enema, and their combination may inhibit the activation of PI3K/Akt/NF-κB signaling pathway and regulate the expression of genes and proteins related to this pathway to achieve anti-inflammatory and anti-apoptotic effects, thus restoring the intestinal mucosal barrier function of UC rats.

9.
Chinese Critical Care Medicine ; (12): 256-262, 2023.
Article in Chinese | WPRIM | ID: wpr-992013

ABSTRACT

Objective:To explore the protective effect of sivelestat (SV) against sepsis-induced acute kidney injury (AKI) and its molecular mechanism.Methods:According to the random number table method, 64 male Wistar rats were divided into sham operation group (Sham group), sepsis due to cecal ligation and puncture group (CLP group), low dose of SV treatment group (SL group, 50 mg/kg SV was injected into the tail vein at 12 hours and 24 hours after CLP), and high dose of SV treatment group (SH group, 100 mg/kg SV was injected into the tail vein at 12 hours and 24 hours after CLP), with 16 rats in each group. 48 hours after CLP, the 48-hour survival of rats were recorded, all rats were sacrificed and samples were harvested. Enzyme-linked immunosorbent assay (ELISA) was used to detect the serum levels of kidney injury molecule-1 (KIM-1), interleukins (IL-1β, IL-6), tumor necrosis factor-α (TNF-α) and neutrophil elastase (NE). Hematoxylin-eosin (HE) staining was used to observe histopathological changes and assess renal tubule injury score. Masson staining was used to detect the collagen volume fraction (CVF) of kidney tissue. Western blotting was used to detect the protein expressions of phosphatidylinositol 3-kinase (PI3K), phosphorylation PI3K (p-PI3K), protein kinase B (AKT), phosphorylation AKT (p-AKT), nuclear factor-κB p65 (NF-κB p65) and NE. The protein expressions of p-PI3K, p-AKT, NF-κB p65 were detected by immunohistochemistry.Results:Compared with Sham group, the 48-hour survival rate of CLP group was significantly reduced. Histopathological results showed that large tubular epithelial cells and brush margins were shed, tubular casts were formed, some tubular atrophy, glomerular hyperemia, renal interstitial inflammatory cell infiltration and increased renal tubular injury score. Renal interstitial fibrosis was obvious and CVF increased. The levels of KIM-1, IL-1β, IL-6, TNF-α and NE in serum were significantly elevated in the CLP group. The proteins expression of inflammatory pathway-related p-PI3K/PI3K, p-AKT/AKT, NF-κB p65 and NE were significantly increased in kidney tissue. It suggested that septic rats had renal injury and the PI3K/AKT inflammatory pathway was activated. Compared with CLP group, there was no significant difference in 48-hour survival in SL group and SH group (68.75%, 75.00% vs. 56.25%, both P > 0.05), but kidney injury was significantly relieved. Specifically: renal tubular injury score and CVF significantly decreased [tubular injury score: 2 (1, 2), 1 (1, 1) vs. 2 (2, 3); CVF: (22.36±0.86)%, (18.74±1.05)% vs. (58.38±0.79)%, all P < 0.05]; the serum levels of KIM-1, IL-1β, IL-6, TNF-α and NE also decreased significantly [KIM-1 (ng/L): 145.03±8.88, 117.58±7.02 vs. 158.22±12.00; IL-1β (ng/L): 108.32±9.00, 92.98±8.06 vs. 133.78±8.48; IL-6 (ng/L): 124.33±10.11, 115.42±8.17 vs. 165.19±5.70; TNF-α (ng/L): 321.56±19.29, 289.68±21.57 vs. 424.88±22.76, NE (mol/L): 93.84±9.14, 75.01±10.56 vs. 113.45±6.39, all P < 0.05]; the proteins expression of inflammatory pathway-related p-PI3K/PI3K, p-AKT/AKT, NF-κB p65 and NE were significantly decreased (p-PI3K/PI3K: 0.93±0.06, 0.67±0.04 vs. 1.27±0.08; p-AKT/AKT: 0.78±0.09, 0.47±0.05 vs. 0.96±0.12; NF-κB p65/GAPDH: 1.43±0.13, 0.85±0.08 vs. 1.88±0.17; NE/GAPDH: 1.45±0.06, 0.91±0.04 vs. 1.71±0.08, all P < 0.05), the positive expressions of p-PI3K, p-AKT and NF-κB p65 in kidney tissue were decreased [p-PI3K positive expression area: (13.36±1.84)%, (8.03±1.12)% vs. (21.56±1.20)%; p-AKT positive expression area: (21.57±0.91)%, (15.21±2.76)% vs. (30.81±2.12)%; NF-κB p65 positive expression area: (25.17±1.38)%, (17.07±2.11)% vs. (37.85±2.50)%, all P < 0.05]. Serum inflammatory factor level, and PI3K/AKT pathway related protein, NF-κB p65, NE protein expression level and p-PI3K, p-AKT, NF-κB p65 positive area and other indicators in renal tissue in SH group were further lower than those in SL group (all P < 0.05). Conclusions:SV can ameliorate sepsis-induced AKI. The mechanism may be related to the inhibition of PI3K/AKT pathway, and high dose of SV has better efficacy.

10.
Chinese Critical Care Medicine ; (12): 140-145, 2023.
Article in Chinese | WPRIM | ID: wpr-991992

ABSTRACT

Objective:To investigate whether microRNA-21-5p (miR-21-5p) alleviates hyperoxia-induced acute lung injury (HALI) through activating the phosphatidylinositol 3 kinase/serine-threonine protein kinase (PI3K/Akt) signaling pathway by regulating apoptosis of type Ⅱ alveolar epithelial cell (AECⅡ).Methods:Seventy-two male Sprague-Dawley (SD) rats were divided into normozone-controlled group, HALI group, PI3K/Akt signaling pathway inhibitor LY294002+HALI group (LY+HALI group), miR-21-5p overexpression+LY294002+HALI group (miR-21-5p+LY+HALI group), miR-21-5p overexpression+HALI group (miR-21-5p+HALI group), and dimethyl sulfoxide (DMSO)+HALI group by random number table method with 12 rats in each group. Animal models of HALI were prepared using 95% concentrations of oxygen. The animals in the normozone-controlled group were fed normally under normoxia. Transfection of lung tissue by miR-21-5p adeno-associated viral vector AAV6-miR-21-5p was performed by instillation of 200 μL titer (1×10 12 TU/mL) through a tracheal catheter 3 weeks prior to modeling. DMSO and LY294002 were administered via the tail vein at 0.3 mg/kg 1 hour before modeling. After 48 hours of modeling, carotid artery blood was collected to detect oxygenation index (OI) and respiratory index (RI), and real-time fluorescence quantitative reverse transcription-polymerase chain reaction (RT-PCR) was used to detect miR-21-5p expression. Lung tissue was collected, and the levels of inflammatory factors including tumor necrosis factor-α (TNF-α) and interleukins (IL-6, IL-1β) were measured by enzyme-linked immunosorbent assay (ELISA), and the ratio of pulmonary wet/dry weight (W/D) was determined, and the pathological changes of lung histopathology were observed under the light microscopy after hematoxylin-eosin (HE) staining. Each group was purified AECⅡ cells from 6 rats, the apoptosis rate was detected by flow cytometry, and the expression levels of phosphatase and tensin homologous gene (PTEN), and proteins from the PI3K/Akt signaling pathway were detected by Western blotting. Results:Compared with the normozone-controlled group, alveolar septal thickening and massive inflammatory cell infiltration were found after hyperoxia exposure, RI, inflammatory factors, lung W/D ratio, pathological score, AECⅡ cells early apoptosis rate, PTEN protein expression and phosphorylation level of Akt were increased, while OI and miR-21-5p expression were decreased, indicating the successful preparation of the model. After pretreatment, LY294002 could aggravate the pathological injury of lung tissue in HALI rats, RI, inflammatory factors and lung W/D ratio were further increased, and OI was further reduced compared with HALI group. At the same time, it could promote the AECⅡ cell apoptosis, further up-regulate the expression of PTEN, and reduce the phosphorylation of Akt. However, miR-21-5p pretreatment could negatively regulate PTEN, activate PI3K/Akt signal pathway, inhibit AECⅡ cell apoptosis, and reduce HALI, which was shown by the decreased level of inflammatory factors in miR-21-5p+LY+HALI group compared with LY+HALI group [TNF-α (μg/L): 100.33±3.48 vs. 116.55±2.53, IL-6 (ng/L): 141.06±3.70 vs. 161.31±3.59, IL-1β (μg/L): 90.82±3.69 vs. 112.23±2.87, all P < 0.05], RI, lung injury pathology score, lung W/D ratio, and AECⅡ cell early apoptosis rate were significantly decreased [RI: 0.81±0.02 vs. 1.05±0.07, pathology score: 0.304±0.008 vs. 0.359±0.007, lung W/D ratio: 5.29±0.03 vs. 5.52±0.08, apoptosis rate: (27.20±2.34)% vs. (34.17±1.49)%, all P < 0.05], OI and expressions of miR-21-5p were significantly increased [OI (mmHg, 1 mmHg≈0.133 kPa): 266.71±2.75 vs. 230.12±4.04, miR-21-5p (2 -ΔΔCt): 2.21±0.13 vs. 0.33±0.03, both P < 0.05], and PTEN protein expression in AECⅡ cell was significantly reduced (PTEN/GAPDH: 0.50±0.06 vs. 0.93±0.06, P < 0.05), and phosphorylation level of Akt was significantly increased [phosphorylated Akt (p-Akt) protein (p-Akt/GAPDH): 0.86±0.05 vs. 0.56±0.06, P < 0.05]. Conclusion:miR-21-5p attenuates HALI by inhibiting AECⅡ cell apoptosis, possibly through negative regulation of PTEN to activate PI3K/Akt signaling pathway.

11.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 253-263, 2023.
Article in Chinese | WPRIM | ID: wpr-997679

ABSTRACT

Osteoporosis (OP), a common systemic skeletal disease in the elderly, is characterised by bone loss and bone microstructural degeneration. Its clinical manifestations include increased bone fragility and bone pain. Furthermore, OP increases the risk of fracture due to the high bone fragility, which leads to lifelong disability or death, imposing a heavy economic and psychological burden on the patients and their families. The pathogenesis of OP is extremely complex and associated with a variety of factors such as proliferation and differentiation of osteoblasts, impairment of osteoclast activity and function, and abnormalities in autophagy activation. Recent studies have found that mammalian target of rapamycin (mTOR) signaing pathway is involved in the regulation of bone homeostasis, which can promote bone formation and improve bone metabolism and bone microstructure by regulating osteoblast proliferation and differentiation and osteoclast function and activating cellular autophagy, thus playing a crucial role in the prevention and treatment of OP. The prevention and treatment of OP with Chinese medicine has a long history, clear efficacy, multiple targets of action, low adverse effects, and wide medicine sources. Therefore, this paper briefly describes the role of mTOR signaling pathway in the development of OP by reviewing the latest research reports and summarizes in detail the latest research results on the treatment of OP with Chinese medicine extracts and prescriptions via the mTOR signaling pathway. This review aims to provide a basis for the in-depth research on the relationship between mTOR signaling pathway and OP and the clinical application of traditional Chinese medicine in the prevention and treatment of OP.

12.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 218-226, 2023.
Article in Chinese | WPRIM | ID: wpr-997675

ABSTRACT

Non-small cell lung cancer (NSCLC) is a malignant tumor of the respiratory system with a high incidence. The early symptoms are not typical, and most patients are diagnosed at an advanced stage, which seriously threatens the lives and health of people. Surgery, chemotherapy, and targeted therapy are the main means of treatment at present, but the consequent drug resistance and adverse reactions restrict these treatment methods with certain limitations. In recent years, with the development of traditional Chinese medicine (TCM) in tumor resistance, TCM has attracted extensive attention for its obvious therapeutic effect and fewer adverse reactions. Numerous signaling pathways are involved in the formation and development of NSCLC, where phosphatidylinositol 3 kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathway is one of the key regulatory pathways. The PI3K/Akt/mTOR signaling pathway affects the proliferation, invasion, and metastasis of NSCLC cells by affecting the cycle, inhibiting the apoptosis, inhibiting the autophagy of tumor cells, and promoting tumor neovascularization. As revealed by the recent literature, Chinese medicine plays an indispensable role in NSCLC cell autophagy, cell cycle, apoptosis, invasion and metastasis, neovascularization, and reversal of drug resistance by regulating the PI3K/Akt/mTOR signaling pathway. Although some Chinese medicinal extracts or compounds have made great breakthroughs in some mechanisms of action in the treatment of NSCLC, these studies only remain at the level of in vitro cell experiments and animal studies in vivo. Researchers are faced with the great challenge of "transforming the research results of Chinese medicines into clinical applications". Based on the current research status in China and abroad, this paper reviewed Chinese medicine in the intervention in NSCLC through the regulation of PI3K/Akt/mTOR signaling pathway in recent years, in order to open up new ideas for NSCLC drug therapy research and also provide a useful reference for further mechanism research.

13.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 29-36, 2023.
Article in Chinese | WPRIM | ID: wpr-996807

ABSTRACT

ObjectiveTo explore the inhibitory effect of water extract of Broussonetiae Fructus on hepatocellular carcinoma (HCC) induced by diethyl nitrosamine (DEN) in mice based on homologous phosphatase and tensin homolog/phosphatidylinositol 3-kinase/protein kinase B (PTEN/PI3K/Akt) signaling pathway. MethodThe primary HCC mouse model was constructed by intraperitoneal injection of DEN solution, and the HCC mice were randomly divided into model group, sorafenib group (0.01 g·kg-1·d-1), low-dose Broussonetiae Fructus water extract group (0.9 g·kg-1·d-1), medium-dose Broussonetiae Fructus water extract group (1.8 g·kg-1·d-1), and high-dose Broussonetiae Fructus water extract group (3.6 g·kg-1·d-1), with 10 mice in each group. Another 10 C57BL/6 mice were selected as a control group and intraperitoneally injected with an equal volume of normal saline. Mice were treated with different concentrations of Broussonetiae Fructus water extract when liver cancer-like white nodules appeared. sorafenib group was treated with sorafenib. The control group and model group were intraperitoneally injected with normal saline. The activities of alkaline phosphatase (ALP), aspartate aminotransferase (AST), alanine aminotransferase (ALT), and γ-glutamyl transferase (γ-GT) in the serum of mice were detected by the biochemical analyzer. The expression levels of alpha-fetoprotein (AFP) and carcinoembryonic antigen (CEA) were detected by enzyme-linked immunosorbent assay (ELISA). The degree of hepatocyte canceration and hepatocyte injury were observed by Hematoxylin-eosin (HE) and Masson staining. The proliferation of HCC cells was observed by immunohistochemical staining. The apoptosis of HCC cells in mice was observed by erminal-deoxynucleotidyl transferase mediated nick end labelling (TUNEL) staining. The expression levels of PTEN, PI3K, Akt, and p-Akt proteins related to the PTEN/PI3K/Akt signaling pathway were detected by Western blot. ResultCompared with the control group, the activities of ALP, AST, ALT, and γ-GT, as well as the expression levels of AFP and CEA in the model group were significantly increased (P<0.01). Carcinogenesis and inflammatory cell infiltration were obvious in liver tissue of mice, and a large number of blue collagen fiber hyperplasia was found. The number of Ki67 positive cells was significantly increased (P<0.01), and the expression level of PTEN protein was significantly decreased, while PI3K and p-Akt protein expression was increased (P<0.01). Compared with the model group, the activities of ALP, AST, ALT, and γ-GT, as well as the expression levels of AFP and CEA in the medium-dose and high-dose Broussonetiae Fructus water extract groups were significantly decreased (P<0.05, P<0.01). The degree of carcinogenesis and inflammatory cell infiltration in liver tissue were reduced, and the collagen fiber hyperplasia was significantly reduced. The number of Ki67 positive cells was significantly decreased, and the number of TUNEL positive apoptotic cells was significantly increased (P<0.05, P<0.01). PTEN protein expression was increased, while p-Akt protein expression was significantly decreased (P<0.05, P<0.01). ConclusionThe water extract of Broussonetiae Fructus has a significant inhibitory effect on DEN-induced primary HCC in mice, and its mechanism may be related to the regulation of key protein expressions in the PTEN/PI3K/Akt signaling pathway.

14.
Chinese Journal of Anesthesiology ; (12): 170-175, 2023.
Article in Chinese | WPRIM | ID: wpr-994169

ABSTRACT

Objective:To evaluate the relationship between hippocampal miR-3065-5p and insulin-like growth factor-1/phosphatidylinositol 3-kinase/protein kinase B(IGF-1/PI3K/Akt)signaling pathway in a mouse model of perioperative neurocognitive disorder (PND).Methods:Eighty clean-grade healthy male C75BL/6 mice, aged 12-14 weeks, weighing 20-30 g, were divided them into 4 groups ( n=20 each) using the random number table method: control group (C group), PND group, miR-3065-5p agonist group (Ag group) and miR-3065-5p agonist negative control group (Ag-NC group). PND model was prepared by internal fixation of tibial fracture under anesthesia with 1.5% isoflurane. Two days before developing the model, miR-3065-5p agomir 2 μl was injected into the lateral ventricle in Ag group, miR-3065-5p agomir negative control 2 μl was injected into the lateral ventricle in Ag-NC group. Morris water maze test and open field test were performed at 7 days after surgery. The mice were sacrificed after the end of test, and hippocampal tissues were obtained for determination of the expression of miR-3065-5p, IGF-1 mRNA and Bcl-2 mRNA (by quantitative real-time polymerase chain reaction) and expression of IGF-1, phosphorylated Akt (p-Akt), phosphorylated glycogen synthase kinase-3β (p-GSK3β) and Bcl-2 (by Western blot). Results:There was no significant difference in each parameter in the open field test among the four groups ( P>0.05). Compared with group C, the postoperative escape latency was significantly prolonged, the percentage of time of stay at the target quadrant was decreased, the number of crossing the original platform was reduced, the expression of miR-3065-5p was up-regulated, and the expression of IGF-1 mRNA, Bcl-2 mRNA, IGF-1, p-Akt, p-GSK3β and Bcl-2 was down-regulated in the other three groups ( P<0.05). Compared with PND group and Ag-NC group, the postoperative escape latency was significantly prolonged, the percentage of time of stay at the target quadrant was decreased, the number of crossing the original platform was reduced, the expression of miR-3065-5p was up-regulated, and the expression of IGF-1 mRNA, Bcl-2 mRNA, IGF-1, p-Akt, p-GSK3β and Bcl-2 was down-regulated in Ag group ( P<0.05). Conclusions:Up-regulation of miR-3065-5p can inhibit the activation of IGF-1/PI3K/Akt signaling pathway, which might be one of the mechanisms of PND developed in mice.

15.
Chinese Journal of Hepatobiliary Surgery ; (12): 357-363, 2023.
Article in Chinese | WPRIM | ID: wpr-993338

ABSTRACT

Objective:To investigate the effects and molecular mechanism of neuropilin and tolloid-like 2 (NETO2) on proliferation, migration, cell cycle, and apoptosis in gallbladder cancer (GBC).Methods:The NETO2 mRNA and protein expression in GBC-SD, ZJU-0430, NOZ GBC cells were detected by quantitative real-time polymerase chain reaction and Western blot. NETO2 overexpression and knockdown stable cell lines were constructed by plasmid transfection. Cell counting kit-8 assay, colony formation assay, transwell assay, flow cytometry and WB assay were performed to evaluate proliferation, migration, cell cycle, apoptosis, epithelial-mesenchymal transition (EMT) and changes of phosphatidylinositol-3 kinase/protein kinase B (PI3K/Akt) signaling pathway.Results:GBC-SD and ZJU-0430 cells with NETO2 gene overexpression and NOZ cells with NETO2 gene knockdown were effectively constructed. NETO2 overexpression in gallbladder cancer cell lines significantly improved cell proliferation and migration, advanced cell cycle progression from G0/G1 to S phase, and inhibited cell apoptosis. In the ZJU-0430 and GBC-SD cells, the clone number increased from (78.5±9.2), (217.0±6.4) to (213.5±10.3), (296.3±9.3)( t=10.98, 6.51; P=0.008, 0.023); The number of migrating cells increased from (198.6±8.4), (233.3±11.0) to (382.7±12.4), (379.0±7.3) ( t=16.98, 16.85, both P<0.001); The total apoptosis rate reduced from (29.7±0.9)%, (35.6±1.1)% to (19.2±0.5)%, (29.1±0.4)% ( t=9.74, 9.05; both P<0.001); The expression of EMT related proteins such as N-cadherin, Vimentin, Snail, and Slug were upregulated, while E-cadherin expression was downregulated. Phosphorylated PI3K (p-PI3K) and Akt (p-Akt) protein expression were significantly increased (all P<0.05). In contrast, NETO2 knockdown had the opposite effect on all these parameters. Conclusion:NETO2 influences the EMT process by regulating the PI3K/Akt signaling pathway, thus promotes GBC cell proliferation, migration and cell cycle progression, and inhibits cancer cell apoptosis.

16.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 10-18, 2023.
Article in Chinese | WPRIM | ID: wpr-964940

ABSTRACT

ObjectiveTo explore the mechanism of Dihuang Yinzi in improving astrocyte injury and glycolysis in Alzheimer's disease (AD) mice via regulating the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway, thereby improving the cognitive function of AD mice. MethodForty male APP/PS1 transgenic mice aged four months were randomly divided into a model group and a model + Dihuang Yinzi (0.25 g·kg-1) group, with 20 mice in each group. Forty C57BL/6J mice with the same background and same age were randomly divided into a control group and a control + Dihuang Yinzi (0.25 g·kg-1) group, with 20 mice in each group. The mice in the control + Dihuang Yinzi group and the model + Dihuang Yinzi group were administered with Dihuang Yinzi by gavage, and those in the control group and the model group received an equal volume of sterilized normal saline, once a day for 150 days. Morris water maze test was performed to test the ability of navigation and space exploration of mice. The protein expression of p-PI3K, PI3K, p-Akt, Akt, phosphofructokinase-1 (PFK-1), and aldehyde dehydrogenase 3 family member B2 (ALDH3B2) in mouse brain tissues was measured by Western blot. An immunofluorescence assay was performed to detect astrocyte morphology and the expression level of ALDH3B2. ResultAs compared with the control group, the model group showed prolonged escape latency during the 2nd to 5th days of the location-based navigation (P<0.05, P<0.01), reduced number of times crossing the target area of the platform, shortened residence time in the target quadrant (P<0.05, P<0.01), prolonged residence time in the opposite quadrant (P<0.05), increased surface area of the cell body and total length of cell protrusions of astrocytes (P<0.05, P<0.01), and down-regulated protein expression of p-PI3K, p-Akt, ALDH3B2, and PFK-1 (P<0.01), while the above experimental indexes were not significantly different in the control + Dihuang Yinzi group. Compared with the model group, the model + Dihuang Yinzi group showed shortened escape latency of APP/PS1 mice during the 2nd to 5th days of the location-based navigation (P<0.05, P<0.01), increased number of times crossing the platform, prolonged target quadrant residence time (P<0.05, P<0.01), shortened residence time in the opposite quadrant (P<0.05), reduced surface area of the cell body and total length of cell protrusions of astrocytes (P<0.05), and up-regulated protein expression of p-PI3K, p-Akt, ALDH3B2, and PFK-1 (P<0.01). ConclusionDihuang Yinzi can improve the learning and memory ability of AD mice by activating the PI3K/Akt signaling pathway and up-regulating the protein expression of PFK-1 and ALDH3B2 to protect against astrocyte injury in brain tissues and improve glycolysis.

17.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 1-11, 2023.
Article in Chinese | WPRIM | ID: wpr-978445

ABSTRACT

ObjectiveTo explore the effect and mechanism of Zuojinwan (ZJW) in the treatment of ulcerative colitis (UC) through network pharmacology and experimental validation. MethodUsing network pharmacology and molecular docking, the active components and potential mechanism of ZJW in treating UC were preliminarily identified. Forty-eight male C57BL/6J mice were randomly divided into a normal group, a model group, a sulfasalazine group (300 mg·kg-1), and low-, medium-, and high-dose ZJW groups (1.82, 3.64, 7.28 g·kg-1). The UC model was induced by dextran sulfate sodium (DSS), and oral administration of drugs began on the third day of modeling, lasting for 7 days. The general condition of mice was observed daily, and the disease activity index (DAI) was evaluated. Hematoxylin-eosin (HE) staining was performed to observe histopathological changes in colon tissue. Enzyme-linked immunosorbent assay (ELISA) was used to measure the levels of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-10 in mouse serum. The molecular mechanism was validated using Western blot. ResultNetwork pharmacology predicted that the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway might be a key pathway in the regulation of UC by ZJW. Molecular docking results showed good binding ability between the key components of ZJW and core targets. Animal experiment results showed that compared with the normal group, the model group had shortened colon length (P<0.01), increased DAI scores, spleen index, colon tissue pathology scores, and levels of TNF-α and IL-6 in serum (P<0.05, P<0.01), increased PI3K, phosphorylated Akt (p-Akt), and B-cell lymphoma-2 (Bcl-2)-associated X protein (Bax) expression in colon tissue (P<0.05, P<0.01), and decreased serum IL-10 levels and colon tissue Bcl-2 protein expression (P<0.01). Compared with the model group, the ZJW groups showed significant improvement in UC symptoms, relieved colon tissue pathological damage, downregulated levels of inflammatory cytokines TNF-α and IL-6 in serum (P<0.01), inhibited expression of PI3K, p-Akt, and Bax proteins in colon tissue (P<0.05, P<0.01), and increased serum IL-10 levels and colon tissue Bcl-2 protein expression (P<0.01), with the high-dose group showing the best effect. ConclusionZJW effectively alleviates DSS-induced UC, and its mechanism may be related to the inhibition of the PI3K/Akt signaling pathway and regulation of apoptosis-related protein expression.

18.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 1-10, 2023.
Article in Chinese | WPRIM | ID: wpr-975150

ABSTRACT

ObjectiveTo observe the effects of Wendantang on the expression of inflammatory cytokines, autophagy markers, and key molecules of phosphatidylinositol-3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway in the adipocytes of the rat model of obesity (syndrome of phlegm-dampness) and to explore the material basis of inflammation in obesity (syndrome of phlegm-dampness) and the underlying mechanism of Wendantang intervention. MethodA total of 126 SD rats were randomized into 2 groups: 16 rats in the blank group and 110 rats in the modeling group. The blank group was fed with a basic diet while the modeling group with a high-fat diet to establish the animal model of obesity (syndrome of phlegm-dampness) for 8 weeks. After successful modeling, 48 obese rats were selected according to their body mass and randomized into a model control group, an orlistat (ORLI, 32.40 mg·kg-1) group, a rapamycin (RAPA, 2 mg·kg-1) group, and low-, medium-, and high-dose (4.45, 8.90, 17.80 g·kg-1, respectively) Wendantang groups, with 8 rats in each group. In addition, 8 rats were randomly selected from the blank group to be set as the normal control group. The corresponding agents in each group were administrated by gavage and the model and control groups were administrated with equal amounts of distilled water once daily for 6 weeks. The body mass, Lee's index, body fat ratio, and obesity rate were measured or calculated. The expression of UNC51-like kinase-1 (ULK1), Beclin1, human autophagy-related protein 5 (Atg5), p62, and microtubule-associated protein 1 light chain 3 (LC3) Ⅰ/Ⅱ (markers of autophagy in adipocytes) was detected by the immunohistochemical two-step method. Enzyme-linked immunosorbent assay (ELISA) was employed to determine the expression of tumor necrosis factor (TNF)-α, interleukin-6 (IL-6), IL-1β, monocyte chemotactic protein-1 (MCP-1), IL-4, IL-10, IL-13, and transforming growth factor (TGF)-β in adipocytes. Western blot was employed to measure the protein levels of classⅠ-PI3K, phosphatidylinositol triphosphate (PIP3), Akt, mTORC1, ULK1, TSC1, and TSC2 in adipocytes. ResultCompared with the blank group, the modeling group showed increased body mass and Lee's index (P<0.01), the obesity rate >20%, and phlegm-dampness syndrome manifestations such as physical obesity, decreased mobility, decreased appetite, lusterless and tight fur, loose stools, decreased responsiveness to the outside world, and decreased water intake. Compared with the normal control group, the model control group showed increased body mass, Lee's index, body fat ratio, adipocyte autophagy marker expression, pro- and anti-inflammatory cytokine levels (P<0.05, P<0.01), down-regulated protein levels of classⅠ-PI3K, PIP3, Akt, mTORC1, TSC1, and TSC2 (P<0.01), and up-regulated protein level of ULK1 (P<0.01). The intervention groups showed lower body mass, body fat ratio, adipocyte autophagy marker protein expression, and protein levels of TNF-α, IL-6, IL-1β, MCP-1, IL-4, and IL-13 than the model control group (P<0.05, P<0.01). Moreover, the RAPA and Wendantang (medium and high dose) groups showed lowered levels of IL-10 and TGF-β (P<0.01), and the ORLI group showed down-regulated expression of TGF-β (P<0.01). The expression of key molecules of the signaling pathway was up-regulated (P<0.05, P<0.01) while that of ULK1 was down-regulated (P<0.01) in all the intervention groups. Compared with the RAPA group, the Wendantang groups showed up-regulated expression of all autophagy marker proteins in adipocytes (P<0.01). In addition, the low-dose Wendantang group showed elevated levels of inflammatory cytokines (except TNF-α) (P<0.05, P<0.01) and down-regulated expression of all key molecules of the signaling pathway (P<0.05, P<0.01). The levels of inflammatory cytokines (except IL-16, MCP-1, and IL-10) were elevated in the medium-dose Wendantang group (P<0.05, P<0.01). The expression of key molecules except PI3K of the signaling pathway was down-regulated in the medium- and high-dose Wendantang groups (P<0.05, P<0.01). Compared with the ORLI group, low- and medium-dose Wendantang groups showed up-regulated expression of autophagy markers in adipocytes (P<0.01), and the low-dose group showed elevated levels of inflammatory cytokines (IL-6, IL-4, and TGF-β) (P<0.01) and down-regulated expression of all key molecules of the signaling pathway (P<0.01). The medium-dose Wendantang group showed up-regulated expression of IL-4 (P<0.01) and down-regulated expression of key molecules except PI3K of the signaling pathway (P<0.05, P<0.01). The high-dose Wendantang group showed increased body mass, up-regulated expression levels of autophagy markers (ULK1, LC3 Ⅰ/Ⅱ) (P<0.05, P<0.01), down-regulated expression of PIP3, mTORC1, and TSC1 (P<0.05, P<0.01), and lowered levels of Beclin1, Atg5, TNF-α, and IL-13 (P<0.05, P<0.01). ConclusionThe inflammation in obesity (syndrome of phlegm-dampness) is closely associated with the PI3K/Akt/mTOR pathway-mediated adipocyte autophagy. Wendantang can treat the chronic inflammation in obese rats with the syndrome of phlegm-dampness by regulating this signaling pathway and thus improve adipocyte autophagy.

19.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 254-263, 2023.
Article in Chinese | WPRIM | ID: wpr-973768

ABSTRACT

Colorectal cancer (CRC), as the third most common cancer in the world, develops from the colonic and rectal epithelial cells. With the rapid development of the global economy, the incidence of CRC in developing countries has been increasing year by year. In the past few years, although preventive colonoscopy screening has improved the survival rate of CRC patients, the majority of cases are still detected after symptoms appear. Currently, the clinical treatment of CRC carries high surgical risks and is prone to recurrence, while radiotherapy and chemotherapy have significant side effects and cause a heavy psychological burden on patients. Therefore, there is currently no ideal treatment protocol for CRC. The phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) pathway, as a classical oncogenic pathway, provides potential for the diagnosis and treatment of various malignant diseases, and offers a new direction for the treatment of CRC. In recent years, traditional Chinese medicine (TCM) has become a major focus in cancer treatment due to its advantages in "preventing and treating diseases" and its multiple components, targets, and pathways. Its advantages in having fewer side effect complement western medicine in treatment. Multiple studies have shown that Chinese medicinal monomers and compound formulas can inhibit the proliferation, invasion, migration, and angiogenesis of CRC cells, promote apoptosis and autophagy of cancer cells, and slow down the development of CRC by intervening in the PI3K/Akt signaling pathway, thereby achieving the therapeutic effect on CRC. In recent years, the research progress related to this has been updating rapidly. Previous literature failed to timely incorporate the latest research results, which brought many inconveniences to the literature search of many scholars. Therefore, this article summarized the relevant information from PI3K/Akt pathway, the association between the PI3K/Akt pathway and CRC, and the progress of TCM intervention in the treatment of CRC to provide references for the development of CRC in molecular biology and the clinical development of new drugs in the future.

20.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 230-240, 2023.
Article in Chinese | WPRIM | ID: wpr-973766

ABSTRACT

Hepatic fibrosis is a pathological reparative response of the liver to chronic injury and a crucial step in the progression of chronic liver disease, characterized mainly by the activation of hepatic stellate cells and diffuse deposition of extracellular matrix. Currently, there is no ideal specific drug for the treatment of liver fibrosis in clinical practice. In recent years, with the development and progress of traditional Chinese medicine (TCM) in the treatment of liver fibrosis, TCM has been widely recognized for its significant therapeutic effect and fewer adverse reactions. The phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway is an important pathway that affects the formation and development of liver fibrosis. It mainly plays a role in liver fibrosis by inhibiting the activation and proliferation of hepatic stellate cells, promoting their apoptosis, reducing oxidative stress in liver cells, decreasing the deposition of extracellular matrix, and enhancing liver cell autophagy. This article summarized the mechanisms by which Chinese medicinal monomers regulated the PI3K/Akt pathway to exert their effects on liver fibrosis and their synergistic effects with other signaling pathways, providing a theoretical basis and references for the development of new drugs for the treatment of liver fibrosis with TCM.

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