Buyang Huanwu Decoction(BYHWD), as one of the classic formulas in traditional Chinese medicine(TCM) for the treatment of cerebral ischemic stroke(CIS), has demonstrated definite effects in clinical practice. However, the material basis and mechanism of treatment have not been systematically elucidated. This study employed network pharmacology and molecular docking to analyze the potential targets and mechanisms of blood-and brain-penetrating active components of BYHWD in reducing cell apoptosis in CIS. Cell experiments were then carried out to validate the prediction results. In the experiments, five active components including hydroxysafflor yellow A( HSYA), tetramethylpyrazine( TMP), astragaloside Ⅳ( AS-Ⅳ), amygdalin( AMY), and paeoniflorin(PF) were selected to explore the pharmacological effects of BYHWD. HT22 cells were treated with BYHWD, and the cell counting kit-8(CCK-8) method was employed to examine the toxic and side effects of BYHWD. A cell model of oxygen-glucose deprivation/reoxygenation( OGD/R) was constructed, with apoptosis and pyroptosis as the main screening indicators. The levels of lactate dehydrogenase(LDH) and glutathione(GSH) were measured to assess the cell membrane integrity. Flow cytometry was employed to detect apoptosis, and the activities of caspase-3 and caspase-1 were measured to clarify the status of apoptosis and pyroptosis. ELISA was employed to determine the levels of interleukin(IL)-1β and IL-18 to confirm pyroptosis. HSYA and AMY were identified in this study as the active components regulating apoptosis and pyroptosis. TUNEL was employed to detect the apoptosis rate, and Western blot was employed to determine the expression levels of apoptosis-related proteins B-cell lymphoma-2(Bcl-2), Bcl-2-associated X protein(Bax), and caspase-3, which confirmed that the anti-apoptotic effect of the combined component group was superior to that of the single component groups. The molecular docking results revealed strong binding affinity of HSYA and AMY with SDF-1α and CXCR4.AMD3100, a selective antagonist of CXCR4, was then used for intervention. The results of Western blot showed alterations in the expression levels of apoptosis-associated proteins, SDF-1α, and CXCR4. In conclusion, HSYA and AMY influence cellular apoptosis by modulating the SDF-1α/CXCR4 signaling cascade.
Drugs, Chinese Herbal/chemistry* ; Apoptosis/drug effects* ; Animals ; Neurons/cytology* ; Mice ; Molecular Docking Simulation ; Cell Line ; Glucose/metabolism* ; Humans ; Neuroprotective Agents/pharmacology*

OBJECTIVE: Comorbid pain and depression are common but remain difficult to treat. Electroacupuncture (EA) can effectively improve symptoms of depression and relieve pain, but its neural mechanism remains unclear. Therefore, we used resting-state functional magnetic resonance imaging (rs-fMRI) to detect cerebral changes after initiating a mouse pain model via constriction of the infraorbital nerve (CION) and then treating these animals with EA. METHODS: Forty male C57BL/6J mice were divided into 4 groups: control, CION model, EA, and sham acupuncture (without needle insertion). EA was performed on the acupoints Baihui (GV20) and Zusanli (ST36) for 20 min, once a day for 10 consecutive days. The mechanical withdrawal threshold was tested 3 days after the surgery and every 3 days after the intervention. The depressive behavior was evaluated with the tail suspension test, open-field test, elevated plus maze (EPM), sucrose preference test, and marble burying test. The rs-fMRI was used to detect the cerebral changes of the functional connectivity (FC) in the mice following EA treatment. RESULTS: Compared with the CION group, the mechanical withdrawal threshold increased in the EA group at the end of the intervention (P < 0.05); the immobility time in tail suspension test decreased (P < 0.05); and the times of the open arm entry and the open arm time in the EPM increased (both P < 0.001). There was no difference in the sucrose preference or marble burying tests (both P > 0.05). The fMRI results showed that EA treatment downregulated the amplitude of low-frequency fluctuations and regional homogeneity values, while these indicators were elevated in brain regions including the amygdala, hippocampus and cerebral cortex in the CION model for comorbid pain and depression. Selecting the amygdala as the seed region, we found that the FC was higher in the CION group than in the control group. Meanwhile, EA treatment was able to decrease the FC between the amygdala and other brain regions including the caudate putamen, thalamus, and parts of the cerebral cortex. CONCLUSION EA can downregulate the abnormal activation of neurons in the amygdala and improve its FC with other brain regions, thus exerting analgesic and antidepressant effects. Please cite this article as: Yin X, Zeng XL, Lin JJ, Xu WQ, Cui KY, Guo XT, Li W, Xu SF. Brain functional changes following electroacupuncture in a mouse model of comorbid pain and depression: a resting-state functional magnetic resonance imaging study. J Integr Med. 2025; 23(2): 159-168.
Animals ; Electroacupuncture ; Male ; Magnetic Resonance Imaging ; Depression/diagnostic imaging* ; Mice, Inbred C57BL ; Brain/diagnostic imaging* ; Disease Models, Animal ; Mice ; Pain/diagnostic imaging* ; Acupuncture Points

Objective:To investigate effect of cedrol on radiosensitivity of prostate cancer(PCa)cells and its mechanism.Methods:mRNA and protein expressions of cGAS and STING in PCa tissues and cells were detected by qRT-PCR and Western blot,respectively.Human PCa cells PC-3 were divided into control group,radiation group,cedrol group,combination group and inhibitor group.Radiosensitivity of cells in each group was detected by plate cloning test;apoptosis,migration and invasion were detected by flow cytometry,scratch test and Transwell chamber,respectively;γ-H2AX immunofluorescence staining was used to analyze effect of cedrol on DNA damage repair;after PC-3 cells and CD8+T cells were co-cultured,cells were divided into T cell group,co-culture group,cedrol group and inhibitor group,MTT and flow cytometry were used to detect proliferation and apoptosis of CD8+T cells,IFN-γ,IL-2 and TNF-α levels in supernatant of CD8+T cells were detected by ELISA;Western blot was used to detect prolife-rating cell nuclear antigen(PCNA),Bcl-2 associated protein(Bax),programmed death recepter ligand 1(PD-L1)and cGAS-STING signal pathway protein.Results:cGAS and STING protein and mRNA expressions in PCa tissues and cells were decreased(P<0.05),which were lowest in PC-3 cells.Compared with control group,colony formation rate,cell survival rate,scratch healing rate and cell inva-sion rate of PC-3 cells in radiation group and cedrol group were decreased obviously,apoptosis rate and number of γ-H2AX focus were increased obviously(P<0.05);compared with radiation group and cedrol group,combined group inhibited proliferation,migration and invasion of PC-3 cells,and induced DNA damage and apoptosis(P<0.05);inhibitor group attenuated inhibitory effect of com-bined group on proliferation,migration and invasion of PC-3 cells,and promoted DNA damage and apoptosis(P<0.05);compared with T cell group,A490,levels of IFN-γ,IL-2 and TNF-α in CD8+T cells in co-culture group were decreased,apoptosis rate was in-creased(P<0.05);compared with co-culture group,A490,levels of IFN-γ,IL-2 and TNF-α in CD8+T cells in cedrol group were in-creased,apoptosis rate was decreased(P<0.05);compared with cedrol group,A490,levels of IFN-γ,IL-2 and TNF-α in CD8+T cells in inhibitor group were decreased,apoptosis rate was increased(P<0.05).Compared with control group,expressions of PCNA and PD-L1 proteins in PC-3 cells in cedrol group were reduced obviously,expressions of Bax,cGAS and STING proteins were increased obviously(P<0.05);inhibitors of cGAS-STING pathway could reverse effect of cedrol on above proteins(P<0.05).Conclusion:Cedrol may enhance radiosensitivity of PCa cells by activating cGAS-STING signal pathway and inhibiting immune escape.

Aim To explore the effect of hydroxysafflor yellow A(HSYA)on lipocalin 2(LCN2)-induced fer-roptosis in HT22 cells and the related mechanism.Methods Thirty male Sprague-Dawley(SD)rats were used to establish the middle cerebral artery occlu-sion/reperfusion(MCAO/R)model by the suture method.The rats were randomly divided into the Sham group,the MCAO/R group,and the MCAO/R+HSYA group.The infarct area was measured by TTC staining,and the degree of neurological deficit was evaluated by the Z-Longa scoring method.The expressions of LCN2 and 24P3R in brain tissues were detected by Western blot.LCN2 protein was added to HT-22 cells,and the cells were divided into the normal group,the LCN2 group,and the LCN2+HSYA group.The optimal con-centration of LCN2-induced neuronal ferroptosis was screened by LDH assay and Western blot,and the ex-pression levels of ferritin,FPN1,GPX4,SLC7A11,COX2,and 24P3R were detected.LCN2 was knocked down by siRNA transfection,and the expressions of GPX4 and ferritin were detected.The contents of glu-tathione(GSH),malondialdehyde(MDA),GPX4,and Fe2+were determined by colorimetry,and the expres-sion of GPX4 was detected by immunofluorescence.The binding force between HSYA and LCN2 was ana-lyzed by molecular docking technology.Results Ani-mal experiments showed that HSYA could reduce the cerebral infarction area and decrease the neurological function score of MCAO/R rats.Compared with the sham group,the levels of LCN2 and 24P3R increased in the MCAO/R group,while HSYA inhibited their ex-pressions.Cell experiments showed that the optimal concentration of LCN2 to induce ferroptosis in HT22 cells was 2 μmol·L-1.After knocking down LCN2 by siRNA transfection,compared with the LCN2 group,the expression levels of GPX4 and ferritin in the siLCN2 group increased significantly.Compared with the nor-mal group,the expressions of SLC7A11,GPX4,FPN1,ferritin,and GSH in the LCN2 group decreased signifi-cantly,while the concentration of Fe2+,and the expres-sions of MDA,COX2,and 24P3R increased.HSYA could increase the expressions of SLC7A11,GPX4,FPN1,ferritin,and GSH,reduce the contents of Fe2+and MDA,and inhibit the expressions of COX2 and 24P3R.Molecular docking showed that the binding en-ergy between HSYA and LCN2 was-8.0 kJ·mol-1.Conclusion HSYA can inhibit LCN2-induced ferrop-tosis in HT22 cells through the SLC7A11/GPX4 signa-ling pathway.

Aim To explore the effect of hydroxysafflor yellow A(HSYA)on lipocalin 2(LCN2)-induced fer-roptosis in HT22 cells and the related mechanism.Methods Thirty male Sprague-Dawley(SD)rats were used to establish the middle cerebral artery occlu-sion/reperfusion(MCAO/R)model by the suture method.The rats were randomly divided into the Sham group,the MCAO/R group,and the MCAO/R+HSYA group.The infarct area was measured by TTC staining,and the degree of neurological deficit was evaluated by the Z-Longa scoring method.The expressions of LCN2 and 24P3R in brain tissues were detected by Western blot.LCN2 protein was added to HT-22 cells,and the cells were divided into the normal group,the LCN2 group,and the LCN2+HSYA group.The optimal con-centration of LCN2-induced neuronal ferroptosis was screened by LDH assay and Western blot,and the ex-pression levels of ferritin,FPN1,GPX4,SLC7A11,COX2,and 24P3R were detected.LCN2 was knocked down by siRNA transfection,and the expressions of GPX4 and ferritin were detected.The contents of glu-tathione(GSH),malondialdehyde(MDA),GPX4,and Fe2+were determined by colorimetry,and the expres-sion of GPX4 was detected by immunofluorescence.The binding force between HSYA and LCN2 was ana-lyzed by molecular docking technology.Results Ani-mal experiments showed that HSYA could reduce the cerebral infarction area and decrease the neurological function score of MCAO/R rats.Compared with the sham group,the levels of LCN2 and 24P3R increased in the MCAO/R group,while HSYA inhibited their ex-pressions.Cell experiments showed that the optimal concentration of LCN2 to induce ferroptosis in HT22 cells was 2 μmol·L-1.After knocking down LCN2 by siRNA transfection,compared with the LCN2 group,the expression levels of GPX4 and ferritin in the siLCN2 group increased significantly.Compared with the nor-mal group,the expressions of SLC7A11,GPX4,FPN1,ferritin,and GSH in the LCN2 group decreased signifi-cantly,while the concentration of Fe2+,and the expres-sions of MDA,COX2,and 24P3R increased.HSYA could increase the expressions of SLC7A11,GPX4,FPN1,ferritin,and GSH,reduce the contents of Fe2+and MDA,and inhibit the expressions of COX2 and 24P3R.Molecular docking showed that the binding en-ergy between HSYA and LCN2 was-8.0 kJ·mol-1.Conclusion HSYA can inhibit LCN2-induced ferrop-tosis in HT22 cells through the SLC7A11/GPX4 signa-ling pathway.

Objective:To investigate effect of cedrol on radiosensitivity of prostate cancer(PCa)cells and its mechanism.Methods:mRNA and protein expressions of cGAS and STING in PCa tissues and cells were detected by qRT-PCR and Western blot,respectively.Human PCa cells PC-3 were divided into control group,radiation group,cedrol group,combination group and inhibitor group.Radiosensitivity of cells in each group was detected by plate cloning test;apoptosis,migration and invasion were detected by flow cytometry,scratch test and Transwell chamber,respectively;γ-H2AX immunofluorescence staining was used to analyze effect of cedrol on DNA damage repair;after PC-3 cells and CD8+T cells were co-cultured,cells were divided into T cell group,co-culture group,cedrol group and inhibitor group,MTT and flow cytometry were used to detect proliferation and apoptosis of CD8+T cells,IFN-γ,IL-2 and TNF-α levels in supernatant of CD8+T cells were detected by ELISA;Western blot was used to detect prolife-rating cell nuclear antigen(PCNA),Bcl-2 associated protein(Bax),programmed death recepter ligand 1(PD-L1)and cGAS-STING signal pathway protein.Results:cGAS and STING protein and mRNA expressions in PCa tissues and cells were decreased(P<0.05),which were lowest in PC-3 cells.Compared with control group,colony formation rate,cell survival rate,scratch healing rate and cell inva-sion rate of PC-3 cells in radiation group and cedrol group were decreased obviously,apoptosis rate and number of γ-H2AX focus were increased obviously(P<0.05);compared with radiation group and cedrol group,combined group inhibited proliferation,migration and invasion of PC-3 cells,and induced DNA damage and apoptosis(P<0.05);inhibitor group attenuated inhibitory effect of com-bined group on proliferation,migration and invasion of PC-3 cells,and promoted DNA damage and apoptosis(P<0.05);compared with T cell group,A490,levels of IFN-γ,IL-2 and TNF-α in CD8+T cells in co-culture group were decreased,apoptosis rate was in-creased(P<0.05);compared with co-culture group,A490,levels of IFN-γ,IL-2 and TNF-α in CD8+T cells in cedrol group were in-creased,apoptosis rate was decreased(P<0.05);compared with cedrol group,A490,levels of IFN-γ,IL-2 and TNF-α in CD8+T cells in inhibitor group were decreased,apoptosis rate was increased(P<0.05).Compared with control group,expressions of PCNA and PD-L1 proteins in PC-3 cells in cedrol group were reduced obviously,expressions of Bax,cGAS and STING proteins were increased obviously(P<0.05);inhibitors of cGAS-STING pathway could reverse effect of cedrol on above proteins(P<0.05).Conclusion:Cedrol may enhance radiosensitivity of PCa cells by activating cGAS-STING signal pathway and inhibiting immune escape.

Objective To observe the efficacy and safety of Morinda officinalis oligosaccharides in the continuation treatment of mild and moderate depression.Methods An open,single-arm,multi-center design was adopted in our study.Adult patients with mild and moderate depression who had received acute treatment of Morinda officinalis oligosaccharides were enrolled and continue to receive Morinda officinalis oligosaccharides capsules for 24 weeks,the dose remained unchanged during continuation treatment.The remission rate,recurrence rate,recurrence time,and the change from baseline to endpoint of Hamilton Depression Scale(HAMD),Hamilton Anxiety Scale(HAMA),Clinical Global Impression-Severity(CGI-S)and Arizona Sexual Experience Scale(ASEX)were evaluated.The incidence of treatment-related adverse events was reported.Results The scores of HAMD-17 at baseline and after treatment were 6.60±1.87 and 5.85±4.18,scores of HAMA were 6.36±3.02 and 4.93±3.09,scores of CGI-S were 1.49±0.56 and 1.29±0.81,scores of ASEX were 15.92±4.72 and 15.57±5.26,with significant difference(P＜0.05).After continuation treatment,the remission rate was 54.59％(202 cases/370 cases),and the recurrence rate was 6.49％(24 cases/370 cases),the recurrence time was(64.67±42.47)days.The incidence of treatment-related adverse events was 15.35％(64 cases/417 cases).Conclusion Morinda officinalis oligosaccharides capsules can be effectively used for the continuation treatment of mild and moderate depression,and are well tolerated and safe.

To investigate the effect of cantharidin ( CTD) on platelet function and the mechanism of anti-platelet aggregation. Methods Washed platelets were collected from the venous blood of healthy volunteers. The effect of CTD on platelet aggregation and release was determined by aggregometer. The CTD concentration was 2.5 ,5 ,10 μmol • L

Aim To explore the effect of Buyang Huanwu Decoction on cerebral ischemia-reperfusion injury in rats by regulating autophagy through PI3K/AKT pathway. Methods The rats were randomly divided into five groups(n=10): sham operation group(Sham), model group(Model), Buyang Huanwu Decoction group(BYHWD), PI3K inhibitor group(LY294002)and Vehicle group(Vehicle). Except Sham group, the other groups were treated with 2h ischemia and 72 h reperfusion for modeling. The Zea Longa score was used to assess the neurological defects, HE was used to observe brain injury in the ischemic penumbra(IP), immunofluorescence was employed to detect LC3, and Western blot was used to detect pathway and autophagy marker proteins. Results Compared BYHWD group with model group, the neurological score of rats decreased, cerebral infarction volume decreased, the pathological lesions of brain IP were relieved, PI3K and p-AKT/AKT expression increased, and LC3Ⅱ/ decreased and p62 increased(P<0.05). The regulatory effect of BYHWD was weakened by LY294002(P<0.05). Conclusion Buyang Huanwu Decoction alleviates cerebral ischemia-reperfusion injury in rats by activating PI3K/AKT pathway to inhibit autophagy.