Objective To explore the protective effects and mechanism of Shenqi Qiangjing granules containing serum on hydrogen peroxide(H2O2)induced oxidative damage and ferroptosis in mouse spermatogonia(GC-1 spg).Methods Thir-ty SPF grade healthy male SD rats were selected and randomly divided into blank group,levocarnitine group,and Shenqi Qiangjing granules group.After 7 days of intragastric administration,drug-containing serum was collected from each group.Using mouse sper-matogonia as a cell model,they were randomly divided into the normal control group,the model control group,blank serum group,levocarnitine containing serum group,and Shenqi Qiangjing granules containing serum group.Except for the normal control group,the other groups used hydrogen peroxide at a concentration of 600 μmol·L-1 to induce injury to mouse spermatogonia for 4 hours,and then established oxidative stress injury models,after 24 hours of medication intervention in each group.The survival rate of cells was detected using CCK-8 method;The levels of intracellular reactive oxygen species(ROS),malondialdehyde(MDA),catalase(CAT),glutathione(GSH),and superoxide dismutase(SOD)were detected by ELISA;The intracellular iron level was detected by iron ion colorimetry;The activities of Caspase-3 and Caspase-9 were detected by colorimetry;The mRNA levels of glu-tathione peroxidase 4(GPX4)and ACSL4 were determined by qRT-PCR.Results Compared with the normal control group,the cell proliferation activity of the model control group decreased significantly,the levels of ROS,MDA and Fe3+were significantly increased,while the activities of CAT,GSH and SOD were significantly decreased in the model control group,however,Caspase-3 and Caspase-9 activities were significantly increased,the results showed significant difference(P＜0.05).Compare with the model control group,Shenqi Qiangjing granules containing serum could significantly increase the cell proliferation activity,decrease the levels of ROS,MDA and Fe3+,increase the activities of CAT,GSH and SOD,and decrease the activities of Caspase-3 and Caspase-9,the results showed significant difference(P＜0.05).The qRT PCR results showed that compared with the model control group,the expression of GPX4 mRNA was upregulated and ACSL4 mRNA was downregulated in blank serum group containing Shenqi Qiangjing granules,and the differences were statistically significant(P＜0.05).Conclusions Shenqi Qiangjing granules have significant protective effects on hydrogen peroxide-induced oxidative stress injury and ferroptosis of spermatogonia in mice.The mechanism may be related to the decrease of Caspase-3 and Caspase-9 activities and the inhibition of oxidative stress injury and ferroptosis.

ObjectiveTo explore the mechanisms associated with Mahoniae Caulis alkaloids （MA） in ameliorating depression by network pharmacology， molecular docking， and animal experiments. MethodsThe component targets of MA were obtained through Swiss Target Prediction and TCMIP database. The depression targets were collected through TCMIP， Genecards， HPO， DrugBank and OMIM database. The depression targets were collected through TCMIP， Genecards， HPO， DrugBank and OMIM database. Protein-protein interaction （PPI） network was constructed by protein interaction analysis （STRING） database. Gene ontology （GO） and Kyoto Encyclopedia of Genes and Genomes （KEGG） analyses were performed through Bioinformatics （DAVID） database. The docking of components and targets was performed by AGFR. The mouse model of depression was established by intraperitoneal injection of corticosterone （CORT） once a day for 35 consecutive days. Sixty mice were randomly allocated into control （0.9% normal saline）， model （CORT， 20 mg·kg^-1）， positive control （fluoxetine hydrochloride， 3.6 mg·kg^-1）， and MA （10， 5， and 2.5 mg·kg^-1） groups. Each group was administrated with corresponding medicine or normal saline once a day for 28 consecutive days. The depression-like behavior of mice was observed. The pathological changes of prefrontal cortex in mice were observed by hematoxylin-eosin staining. Terminal deoxynucleotidyl dUTP transferase nick end labeling （TUNEL） was employed to observe the apoptosis of neurons in the prefrontal cortex. Enzyme-linked immunosorbent assay was employed to assess the serum levels of brain-derived neurotrophic factor （BDNF）， dopamine （DA）， 5-hydroxytryptamine （5-HT）， and norepinephrine （NE） in mice. The mRNA levels of cyclic adenosine monophosphate （cAMP） pathway-related factors and inflammatory factors were determined by Real-time PCR. Western blot was employed to determine the expression of cAMP pathway-related factors and connexin 43 （Cx43）. ResultsA total of 434 component targets and 545 depression targets were obtained， including 84 common targets， among which 10 core targets were screened out. GO analysis predicted 34 biological processes， 15 cell components， and 11 molecular functions. The KEGG pathways were mainly related to gap junction and cAMP signaling pathway. The core components had good binding affinity with the core targets. The results of animal experiments showed that compared with the control group， CORT prolonged the immobility time of mice in forced swimming and tail suspension tests （P<0.01）， lowered the serum levels of NE， BDNF， and 5-HT （P<0.05）， up-regulated the mRNA levels of nuclear factor-κB （NF-κB） and interleukin-6 （IL-6） in the brain tissue （P<0.05）， and down-regulated the mRNA levels of cyclic adenosine monophosphate effector binding protein （CREB） and BDNF （P<0.05） and the protein levels of protein kinase （PRKACA）， phosphorylation （p）-CREB/CREB， BDNF， and Cx43 （P<0.05） in the brain tissue. Compared with the model group， high-dose MA reduced the immobility time of mice in forced swimming （P<0.05） and tail suspension （P<0.01） tests， raised the serum levels of NE， BDNF， and 5-HT （P<0.01）， down-regulated the mRNA level of NF-κB （P<0.01）， and up-regulated the mRNA level of BDNF （P<0.01） and protein levels of PRKACA， p-CREB/CREB， BDNF， and Cx43 （P<0.05）. ConclusionMA alleviates the CORT-induced depressive behavior of mice. It may play an antidepressant role by regulating cAMP signaling pathway and gap junction pathway， improving synaptic plasticity and gap junction function， and reducing neuroinflammation.

ObjectiveTo explore the mechanisms associated with Mahoniae Caulis alkaloids （MA） in ameliorating depression by network pharmacology， molecular docking， and animal experiments. MethodsThe component targets of MA were obtained through Swiss Target Prediction and TCMIP database. The depression targets were collected through TCMIP， Genecards， HPO， DrugBank and OMIM database. The depression targets were collected through TCMIP， Genecards， HPO， DrugBank and OMIM database. Protein-protein interaction （PPI） network was constructed by protein interaction analysis （STRING） database. Gene ontology （GO） and Kyoto Encyclopedia of Genes and Genomes （KEGG） analyses were performed through Bioinformatics （DAVID） database. The docking of components and targets was performed by AGFR. The mouse model of depression was established by intraperitoneal injection of corticosterone （CORT） once a day for 35 consecutive days. Sixty mice were randomly allocated into control （0.9% normal saline）， model （CORT， 20 mg·kg^-1）， positive control （fluoxetine hydrochloride， 3.6 mg·kg^-1）， and MA （10， 5， and 2.5 mg·kg^-1） groups. Each group was administrated with corresponding medicine or normal saline once a day for 28 consecutive days. The depression-like behavior of mice was observed. The pathological changes of prefrontal cortex in mice were observed by hematoxylin-eosin staining. Terminal deoxynucleotidyl dUTP transferase nick end labeling （TUNEL） was employed to observe the apoptosis of neurons in the prefrontal cortex. Enzyme-linked immunosorbent assay was employed to assess the serum levels of brain-derived neurotrophic factor （BDNF）， dopamine （DA）， 5-hydroxytryptamine （5-HT）， and norepinephrine （NE） in mice. The mRNA levels of cyclic adenosine monophosphate （cAMP） pathway-related factors and inflammatory factors were determined by Real-time PCR. Western blot was employed to determine the expression of cAMP pathway-related factors and connexin 43 （Cx43）. ResultsA total of 434 component targets and 545 depression targets were obtained， including 84 common targets， among which 10 core targets were screened out. GO analysis predicted 34 biological processes， 15 cell components， and 11 molecular functions. The KEGG pathways were mainly related to gap junction and cAMP signaling pathway. The core components had good binding affinity with the core targets. The results of animal experiments showed that compared with the control group， CORT prolonged the immobility time of mice in forced swimming and tail suspension tests （P<0.01）， lowered the serum levels of NE， BDNF， and 5-HT （P<0.05）， up-regulated the mRNA levels of nuclear factor-κB （NF-κB） and interleukin-6 （IL-6） in the brain tissue （P<0.05）， and down-regulated the mRNA levels of cyclic adenosine monophosphate effector binding protein （CREB） and BDNF （P<0.05） and the protein levels of protein kinase （PRKACA）， phosphorylation （p）-CREB/CREB， BDNF， and Cx43 （P<0.05） in the brain tissue. Compared with the model group， high-dose MA reduced the immobility time of mice in forced swimming （P<0.05） and tail suspension （P<0.01） tests， raised the serum levels of NE， BDNF， and 5-HT （P<0.01）， down-regulated the mRNA level of NF-κB （P<0.01）， and up-regulated the mRNA level of BDNF （P<0.01） and protein levels of PRKACA， p-CREB/CREB， BDNF， and Cx43 （P<0.05）. ConclusionMA alleviates the CORT-induced depressive behavior of mice. It may play an antidepressant role by regulating cAMP signaling pathway and gap junction pathway， improving synaptic plasticity and gap junction function， and reducing neuroinflammation.

Objective To elucidate the molecular mechanism of reserpine-induced depression using network toxicology,molecular docking techniques,behavioral assessments of animal models,and histopathological analyses.Methods Core targets were screened using multi-database network toxicology,followed by the construction of a protein-protein interaction network and validation of core targets through molecular docking.Sprague Dawley rats were divided randomly into control and reserpine(0.5 mg/kg)groups,and administered the corresponding treatments once daily for 4 consecutive days.Behavioral changes were assessed using the forced-swim and open-field tests.Serum neurotransmitters were quantified by enzyme-linked immunosorbent assay and neuropathological damage was observed via tissue staining.Target gene expression regulation was verified by Western blot.Results Network toxicology screening and molecular docking simulation demonstrated that reserpine exhibited significant binding affinity with the dopamine D2 receptor,cyclic-AMP response element binding protein,and serine/threonine-protein kinase 1(AKT1).Animal experiments demonstrated that reserpine-treated rats displayed depression-like behaviors,including motor inhibition(P＜0.01),with decreased serum levels of norepinephrine and 5-hydroxytryptamine(P＜0.01),respectively.Pathological observations revealed microglial proliferation in the cerebral cortex,increased apoptosis,and reduced Nissl bodies in the hippocampal CA1 region.Down-regulation of brain-derived neurotrophic factor(BDNF)in brain tissue and decreased expression of hippocampal AKT1 and phosphorylated AKT1 were also observed.Conclusions Reserpine influences monoamine transmitter metabolism and neuronal structural integrity via the inhibition of BDNF and AKT1 protein expression,resulting in depressive-like behavior and cerebral nerve damage in rats.

Objective To elucidate the molecular mechanism of reserpine-induced depression using network toxicology,molecular docking techniques,behavioral assessments of animal models,and histopathological analyses.Methods Core targets were screened using multi-database network toxicology,followed by the construction of a protein-protein interaction network and validation of core targets through molecular docking.Sprague Dawley rats were divided randomly into control and reserpine(0.5 mg/kg)groups,and administered the corresponding treatments once daily for 4 consecutive days.Behavioral changes were assessed using the forced-swim and open-field tests.Serum neurotransmitters were quantified by enzyme-linked immunosorbent assay and neuropathological damage was observed via tissue staining.Target gene expression regulation was verified by Western blot.Results Network toxicology screening and molecular docking simulation demonstrated that reserpine exhibited significant binding affinity with the dopamine D2 receptor,cyclic-AMP response element binding protein,and serine/threonine-protein kinase 1(AKT1).Animal experiments demonstrated that reserpine-treated rats displayed depression-like behaviors,including motor inhibition(P＜0.01),with decreased serum levels of norepinephrine and 5-hydroxytryptamine(P＜0.01),respectively.Pathological observations revealed microglial proliferation in the cerebral cortex,increased apoptosis,and reduced Nissl bodies in the hippocampal CA1 region.Down-regulation of brain-derived neurotrophic factor(BDNF)in brain tissue and decreased expression of hippocampal AKT1 and phosphorylated AKT1 were also observed.Conclusions Reserpine influences monoamine transmitter metabolism and neuronal structural integrity via the inhibition of BDNF and AKT1 protein expression,resulting in depressive-like behavior and cerebral nerve damage in rats.

Objective To explore the protective effects and mechanism of Shenqi Qiangjing granules containing serum on hydrogen peroxide(H2O2)induced oxidative damage and ferroptosis in mouse spermatogonia(GC-1 spg).Methods Thir-ty SPF grade healthy male SD rats were selected and randomly divided into blank group,levocarnitine group,and Shenqi Qiangjing granules group.After 7 days of intragastric administration,drug-containing serum was collected from each group.Using mouse sper-matogonia as a cell model,they were randomly divided into the normal control group,the model control group,blank serum group,levocarnitine containing serum group,and Shenqi Qiangjing granules containing serum group.Except for the normal control group,the other groups used hydrogen peroxide at a concentration of 600 μmol·L-1 to induce injury to mouse spermatogonia for 4 hours,and then established oxidative stress injury models,after 24 hours of medication intervention in each group.The survival rate of cells was detected using CCK-8 method;The levels of intracellular reactive oxygen species(ROS),malondialdehyde(MDA),catalase(CAT),glutathione(GSH),and superoxide dismutase(SOD)were detected by ELISA;The intracellular iron level was detected by iron ion colorimetry;The activities of Caspase-3 and Caspase-9 were detected by colorimetry;The mRNA levels of glu-tathione peroxidase 4(GPX4)and ACSL4 were determined by qRT-PCR.Results Compared with the normal control group,the cell proliferation activity of the model control group decreased significantly,the levels of ROS,MDA and Fe3+were significantly increased,while the activities of CAT,GSH and SOD were significantly decreased in the model control group,however,Caspase-3 and Caspase-9 activities were significantly increased,the results showed significant difference(P＜0.05).Compare with the model control group,Shenqi Qiangjing granules containing serum could significantly increase the cell proliferation activity,decrease the levels of ROS,MDA and Fe3+,increase the activities of CAT,GSH and SOD,and decrease the activities of Caspase-3 and Caspase-9,the results showed significant difference(P＜0.05).The qRT PCR results showed that compared with the model control group,the expression of GPX4 mRNA was upregulated and ACSL4 mRNA was downregulated in blank serum group containing Shenqi Qiangjing granules,and the differences were statistically significant(P＜0.05).Conclusions Shenqi Qiangjing granules have significant protective effects on hydrogen peroxide-induced oxidative stress injury and ferroptosis of spermatogonia in mice.The mechanism may be related to the decrease of Caspase-3 and Caspase-9 activities and the inhibition of oxidative stress injury and ferroptosis.