ObjectiveTo explore the syndromes and mechanisms of depression induced by maternal separation （MS） combined with chronic restraint stress （RS） in mice. MethodOn postnatal day 0 （PD0）， the offspring mice were randomized into a blank group （NC） and a modeling group. The mouse model of depression was established by MS+RS for 21 days. After removal of female mice on PD21， the modeled mice were randomized into model， Wenyang， Jieyu， Wenyang Jieyu， and fluoxetine groups， with 15 mice in each group. The sucrose preference， tail suspension， and open field tests were carried out to evaluate the anxiety and depression-like behavior in mice. Enzyme-linked immunosorbent assay was used to measure the adrenocorticotrophic hormone （ACTH） and corticosterone （CORT） levels in mouse plasma. High performance liquid chromatography-electrochemical detector was used to determine the content of monoamine neurotransmitters in the hippocampus. Real-time fluorescence quantitative polymerase chain reaction was employed to determine the mRNA levels of genes in the 5-hydroxytryptamine （5-HT） system， hypothalamic-pituitary-adrenal （HPA） axis， and brain-derived neurotrophic factor （BDNF） signaling pathway in the hippocampus. Immunohistochemistry was employed to determine the expression levels of proteins in the 5-HT system and HPA axis in the hippocampus. The Simple Western system was used to determine the protein levels of BDNF and tyrosine kinase receptor B （TrkB） in the hippocampus. ResultCompared with the NC group， the model group exhibited depression-like behavior， which was significantly relieved by Wenyang Jieyu prescription and fluoxetine. Compared with the NC group， the model group showed elevated levels of CORT and ACTH in the plasma （P<0.01）， which， however， were lowered by Wenyang Jieyu prescription and fluoxetine （P<0.05， P<0.01）. Compared with the NC group， the model group showed inhibited expression of neurotransmitters in the hippocampus （P<0.05， P<0.01）， while Wenyang Jieyu prescription and fluoxetine restored the expression of neurotransmitters （P<0.05， P<0.01）. Compared with NC group， the model group showed inhibition of the 5-HTergic nerve and abnormal activation of the HPA axis， and Wenyang Jieyu prescription and fluoxetine regulated the abnormal state of the 5-HTergic nerve and HPA axis. Compared with NC group， the modeling down-regulated the mRNA and protein levels of BDNF and TrkB in the hippocampus （P<0.05， P<0.01）， which， however， were recovered in Wenyang， Jieyu， Wenyang Jieyu， and fluoxetine groups （P<0.05， P<0.01）. ConclusionThe mouse model of depression induced by MS+RS may present the syndrome of Yang deficiency and liver depression. Wenyang Jieyu prescription may increase the content of hippocampal neurotransmitters by regulating the 5-HT system and the BDNF signaling pathway mediated by the HPA axis， thereby alleviating depression-like behavior in mice.

ObjectiveTo explore the effects of Wenyang Jieyu prescription （WJP） on neuroinflammation and synaptic plasticity in the mouse model of depression induced by maternal separation combined with restraint stress. MethodThe mice on postnatal day 0 （PD0） were randomized into a control group and a modeling group. Maternal separation combined with restraint stress was employed to establish the mouse model of depression. After the removal of female mice， the modeled mice were randomized into model， Wenyang prescription （5.85 g·kg^-1）， Jieyu prescription （12.03 g·kg^-1）， WJP （16.71 g·kg^-1）， and fluoxetine （2.6 mg·kg^-1） groups on the weaning day （PD21）， with 15 mice in each group. The mice were administrated with corresponding drugs mixed with the diet from PD21 to PD111. The sucrose preference test， open field test， O-maze test， and novel object recognition test were then carried out to evaluate the depression state， memory， and learning ability of the mice. Immunohistochemistry （IHC） was employed to observe the ionized calcium-binding adapter molecule-1 （Iba-1） in hippocampal microglia. High performance liquid chromatography （HPLC） was employed to measure the content of noradrenaline （NE） and epinephrine （E） in the hippocampus. Enzyme-linked immunosorbent assay （ELISA） was employed to determine the content of interleukin （IL）-18 and IL-1β in the hippocampus. Western blot was employed to determine the protein levels of NOD-like receptor protein 3 （NLRP3）， apoptosis-associated speck-like protein containing a CARD （ASC）， cysteine aspartate-specific protease-1 （Caspase-1）， IL-1β， synaptophysin （Syn）， and postsynaptic density 95 （PSD95）. ResultCompared with control group， the model group showed decreased sucrose preference rate， time spent in central zone within 5 min， total movement distance， time spent in the open arm， and cognition index （P<0.05， P<0.01）. The microglia in the model group presented amoeba-like appearance， the Iba1 increased. Moreover， the model group showed decreased content of NE and E （P<0.01）， elevated levels of IL-1β and IL-18 （P<0.01）， down-regulated protein levels of PSD95 and Syn （P<0.05， P<0.01）， and up-regulated protein levels of NLRP3， ASC， Caspase-1， and IL-1β （P<0.05， P<0.01）. Compared with model group， WJP and fluoxetine increased the sucrose preference rate， time spent in central zone within 5 min， total movement distance， time spent in the open arm， and cognition index （P<0.05， P<0.01）. They recovered the microglia and the Iba1 decreased. Moreover， the drugs increased the content of NE and E （P<0.05， P<0.01）， lowered the levels of IL-1β and IL-18 （P<0.01）， up-regulated the protein levels of PSD95 and Syn （P<0.01）， down-regulated the protein levels of NLRP3， ASC， Caspase-1， and IL-1β （P<0.05， P<0.01）. ConclusionWJP can treat the depressive behavior induced by maternal separation combined with restraint stress in mice， with the performance outperforming Wenyang prescription and Jieyu prescription. It may alleviate the neuroinflammation induced by microglia and improve the synaptic plasticity by regulating the NLRP3 pathway and increasing neurotransmitters in the hippocampus.

ObjectiveTo explore the mechanism of Wenyang Jieyu prescription in regulating hippocampal neuron apoptosis and improving synaptic plasticity in the mouse model of depression induced by maternal separation combined with restraint stress. MethodThe mice on postnatal day 0 （PD0） were randomly assigned into a control group （n=10） and a modeling group （n=50）. Maternal separation combined with restraint stress was adopted to establish the mouse model of depression， and the modeled mice were randomized into model， Wenyang prescription， Jieyu prescription， Wenyang Jieyu prescription， and fluoxetine groups （n=10） on the weaning day （PD21）. From PD21 to PD111， the mice were fed with the diets mixed with corresponding medicines. The sucrose preference test， open field test， O-maze test， and novel object recognition test were then conducted to evaluate the depression， memory， and learning abilities of mice. Immunohistochemistry （IHC） was employed to measure the atomic absorbance （AA） of postsynaptic density protein 95 （PSD95） in the hippocampus. Terminal-deoxynucleoitidyl transferase-mediated nick-end labeling （TUNEL） was employed to detect the apoptosis of hippocampal neurons. Western blot was employed to determine the protein levels of brain-derived neurotrophic factor （BDNF）， phosphorylated tyrosine kinase receptor B/tyrosine kinase receptor B （p-TrkB/TrkB）， phosphorylated protein kinase B/protein kinase B （p-Akt/Akt）， phosphorylated mammalian target of rapamycin/mammalian target of rapamycin （p-mTOR/mTOR）， B-cell lymphoma-2 （Bcl-2）， Bcl-2-associated X （Bax）， cysteinyl aspartate-specific proteinase-3 （Caspase-3）， synaptophysin （Syn）， and PSD95. ResultCompared with the control group， the modeling decreased the sucrose preference rate， time spent in central zone within 5 min， total movement distance， time spent in the open arm， and cognition index （P<0.01）. Furthermore， it decreased the expression of PSD95， increased the neuron apoptosis in the hippocampus （P<0.01）， down-regulated the protein levels of BDNF， p-TrkB/TrkB， p-Akt/Akt， p-mTOR/mTOR， Bcl-2， PSD95， and Syn （P<0.01）， and up-regulated the protein levels of Bax and Caspase-3 （P<0.05） in the hippocampus. Compared with the model group， Wenyang Jieyu prescription and fluoxetine increased the sucrose preference rate， time spent in central zone within 5 min， total movement distance， time spent in the open arm， and cognition index （P<0.05， P<0.01）. Moreover， the drugs increased the expression of PSD95， reduced the neuron apoptosis （P<0.01）， up-regulated the protein levels of BDNF， p-TrkB/TrkB， p-Akt/Akt， p-mTOR/mTOR， Bcl-2， PSD95， and Syn （P<0.01）， and down-regulated the protein levels of Bax and Caspase-3 （P<0.01）. ConclusionWenyang Jieyu prescription outperformed Wenyang prescription and Jieyu prescription in the treatment of the depressive behavior induced by maternal separation combined with restraint stress in mice. It exerted the therapeutic effect by reducing the hippocampal neuron apoptosis and improving the synaptic plasticity via the BDNF/Akt/mTOR pathway.

OBJECTIVE To study the immunomodulatory effect of Guipi pills on D-galactose（D-gal）-induced aging mice. METHODS The immune-related targets and related pathways for Guipi pills to exert immune effects were screened by network pharmacology and verified through pharmacodynamic experiments. Totally 105 male ICR mice were randomly divided into blank control （CON） group， model control （MOD） group， positive control （POS） group， Guipi pills low-dose （GD） group and Guipi pills high-dose （GG） group. Except for the CON group， other groups were subcutaneously injected with 400 mg/kg D-gal to induce the aging model； CON group and MOD group were given distilled water， POS group was given 300 mg/kg pidotimod oral solution intragastrically， GD group and GG group were given Guipi pills 300， 600 mg/kg intragastrically， once a day， for 8 weeks. After medication， the serum and spleen were collected， and the contents of interleukin 2 （IL-2）， IL-4， IL-6 and tumor necrosis factor α （TNF-α）， and the contents of immunoglobulin G （IgG）， IgM and IgA were detected. The spleen index was calculated and the histopathological changes in the spleen were observed. The activities of superoxide dismutase （SOD）， glutathione peroxidase （GSH- Px） and malondialdehyde （MDA）， and the content of 8-hydroxy-2 deoxyguanosine （8-OHdG） in spleen were detected； the expression of TNF/phosphoinositide-3-kinase-threonine protein kinase （PI3k-Akt）-related proteins in spleen was detected except for POS group. RESULTS The results of network pharmacology showed that TNF， IL-6 and Akt1 were core targets. The results of pharmacodynamic study showed that compared with MOD group， the contents of IL-2， IL-4， IgG， IgM and IgA were increased significantly in Guipi pills groups， while the contents of TNF-α and IL-6 were decreased significantly； the spleen index was increased significantly （P＜0.05 or P＜0.01）. The phenomenon of diffuse proliferation of lymphocytes was improved， the spleen cells were closely arranged， and the line between the white pulp and red pulp was clear. The activities of SOD and GSH-Px in spleen were increased significantly， while the activity of MDA， the content of 8-OHdG， and the protein expressions of TNF-α， PI3K and p-Akt were decreased significantly （P＜0.05 or P＜0.01）. CONCLUSIONS Guipi pills can regulate the immune function of D-gal-induced aging mice， which is related to regulating the TNF/PI3k-Akt pathway， thereby reducing oxidative stress damage in spleen tissue of mice， and regulating protein expressions of TNF-α， PI3K and p-Akt.

ObjectiveTo explore the mechanism of Xiaoyaosan in alleviating lipopolysaccharide （LPS）-induced depressive-like behavior in mice based on the c-Jun N-terminal kinase （JNK） pathway. MethodAfter adaptive feeding， C57BL/6J mice were randomly divided into normal group， model group， minocycline group （intrabitoneal injection， 50 mg·kg^-1）， fluoxetine group （intragastric administration， 2.6 mg·kg^-1）， and low-， medium-， and high-dose Xiaoyaosan groups （intragastric administration，6.012 5， 12.025， and 24.050 g·kg^-1）. After 14 days of administration， the model group and each administration group were intraperitoneally injected with 2 mg·kg^-1 LPS， and the normal group was intraperitoneally injected with equal volume of normal saline. Depressive-like behavior in mice was assessed using the open field test and the elevated zero maze test. High-performance liquid chromatography （HPLC） was used to measure the levels of norepinephrine （NE） and epinephrine （E） in the mouse hippocampus. Enzyme-linked immunosorbent assay （ELISA） was performed to determine serum interleukin-1β （IL-1β） levels. Immunohistochemistry was used to measure the protein expression levels of ionized calcium-binding adapter molecule-1 （Iba-1）， c-Fos， and c-Jun. Real-time polymerase chain reaction （Real-time PCR） was used to measure mRNA expression levels of IL-1β， c-Jun， c-Fos， and JNK3 in the mouse hippocampus. Protein expression levels of JNK and phosphorylated （p）-JNK in the mouse hippocampus were measured using capillary protein automated protein expression analysis system （Western）. ResultCompared with the normal group， the model group exhibited significantly reduced central area residence time， crossing times， and travel distance in the open field （P<0.01）， significantly increased serum IL-1β levels （P<0.01）， significantly decreased NE and E levels （P<0.05）， upregulated mRNA expression of IL-1β， JNK3， and c-Fos， and increased protein expression of Iba-1， c-Fos， and c-Jun （P<0.05， P<0.01）. Compared with the model group， the Xiaoyaosan groups showed increased central area residence time and open arm residence time （P<0.05）， increased NE and E levels （P<0.01）， decreased mRNA expression of IL-1β， JNK3， c-Jun， and c-Fos， and decreased protein expression of Iba-1， c-Fos， JNK， and p-JNK （P<0.05， P<0.01）. The minocycline group and the fluoxetine group showed decreased mRNA expression of JNK3， c-Jun， and c-Fos （P<0.05， P<0.01）. The minocycline group showed decreased serum IL-1β and p-JNK protein expression （P<0.01）. The fluoxetine group exhibited increased NE and E levels and decreased c-Fos protein expression （P<0.01）. ConclusionXiaoyaosan can improve depressive-like behavior induced by LPS in mice， and its mechanism may be related to the inhibition of neuroinflammatory responses and the JNK pathway.

Objective:To investigate the effect of on-demand glucocorticoid strategy on the occurrence and outcome of porcine anti-lymphocyte globulin (p-ALG) -associated serum sickness in aplastic anemia (AA) .Methods:The data of AA patients who received in the Anemia Diagnosis and Treatment Center of Haematology Hospital, CAMS & PUMC from January 2019 to January 2022 were collected. Among them, 35 patients were enrolled in the on-demand group, with the glucocorticoid strategy adjusted based on the occurrence and severity of serum sickness; 105 patients were recruited in the usual group by matching the age and disease diagnosis according to 1∶3 ratio in patients who received a conventional glucocorticoid strategy in the same period. The incidences, clinical manifestations, treatment outcomes of serum sickness, and glucocorticoid dosage between the two groups were analyzed.Results:The incidences of serum sickness in the on-demand group and the usual group were 65.7% and 54.3% ( P=0.237) , respectively. The median onset of serum sickness was the same [12 (9, 13) d vs the 12 (10, 13) d, P=0.552], and clinical symptoms and signs, primarily joint, and/or muscle pain, fever, and rash were similar. Severity grades were both dominated by Grades 1-2 (62.8% vs 51.4%) , with only a few Grade 3 (2.9% vs 2.9%) , and no Grades 4-5. No significant difference in the serum sickness distribution ( P=0.530) . The median duration of serum sickness was the same [5 (3, 7) d vs 5 (3, 6) d, P=0.529], and all patients were completely cured after glucocorticoid therapy. In patients without serum sickness, the average dosage of prophylactic glucocorticoid per patient in the usual group was (469.48 ±193.57) mg (0 in the on-demand group) . When compared to the usual group, the average therapeutic glucocorticoid dosage per patient in the on-demand group was significantly lower [ (125.91±77.70) mg vs (653.90±285.56) mg, P<0.001]. Conclusions:In comparison to the usual glucocorticoid strategy, the on-demand treatment strategy could significantly reduce glucocorticoid dosage without increasing the incidence of serum sickness; in addition, the duration of serum sickness and the incidence of above Grade 2-serum sickness were similar.

Objective:To analyze the diagnostic value of cell-free plasma metagenomic next-generation sequencing (mNGS) pathogen identification for severe aplastic anemia (SAA) bloodstream infection.Methods:From February 2021 to February 2022, mNGS and conventional detection methods (blood culture, etc.) were used to detect 33 samples from 29 consecutive AA patients admitted to the Anemia Diagnosis and Treatment Center of the Hematology Hospital of the Chinese Academy of Medical Sciences to assess the diagnostic consistency of mNGS and conventional detection, as well as the impact on clinical treatment benefits and clinical accuracy.Results:①Among the 33 samples evaluated by mNGS and conventional detection methods, 25 cases (75.76%) carried potential pathogenic microorganisms. A total of 72 pathogenic microorganisms were identified from all cases, of which 65 (90.28%) were detected only by mNGS. ②All 33 cases were evaluated for diagnostic consistency, of which 2 cases (6.06%) were Composite, 18 cases (54.55%) were mNGS only, 2 cases (6.06%) were Conventional method only, 1 case (3.03%) was both common compliances (mNGS/Conventional testing) , and 10 cases (30.3%) were completely non-conforming (None) . ③All 33 cases were evaluated for clinical treatment benefit. Among them, 8 cases (24.24%) received Initiation of targeted treatment, 1 case (3.03%) received Treatment de-escalation, 13 cases (39.39%) received Confirmation, and the remaining 11 cases (33.33%) received No clinical benefit. ④ The sensitivity of 80.77%, specificity of 70.00%, positive predictive value of 63.64%, negative predictive value of 84.85%, positive likelihood ratio of 2.692, and negative likelihood ratio of 0.275 distinguished mNGS from conventional detection methods (21/12 vs 5/28, P<0.001) . Conclusion:mNGS can not only contribute to accurately diagnosing bloodstream infection in patients with aplastic anemia, but can also help to guide accurate anti-infection treatment, and the clinical accuracy is high.

Objective:Short-term efficacy and safety of afatrombopag conversion therapy in patients with aplastic anemia (AA) who were previously ineffectively treated with intense immunosuppressive therapy (IST) combined with TPO receptor Agonist (TPO-RA) or who were unable to tolerate the side effects of TPO-RA.Methods:Analysis of patients with severe aplastic anemia (SAA) treated in Institute of Hematology & Blood Diseases Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College from January 2021 to December 2021 who received IST combined with TPO-RA (eltrombopag/hatrombopag) for at least 6 months, but was ineffective for at least 3 months or patients who cannot continue to use TPO-RA due to side effects, and switched from TPO-RA to avatrombopag (APAG) , and treated for at least 6 months. This study analyzed its short-term efficacy and evaluated its safety.Results:The median age was 54 (14-68) years old among the 16 patients with AA (8 male and eight female) . A total of ten patients (refractory group) who did not respond to IST combined with TPO-RA were converted to APAG median therapy for 6 (6-10) months. Only seven patients (70% ) obtained trilineage HR [four cases of complete treatment response (CTR) , one case of good treatment response (GPR) , and two cases of partial treatment response (PR) ], all of which began to take effect at 3 months of APAG treatment. There were six patients with TPO-RA intolerance, and APAG was treated for 6 (2 to 8) months. About four patients (67% ) received HR (three GPR cases and one PR case) , of which two patients received PR at 3 months and four patients received HR at 6 months of APAG treatment. No APAG-related grade 2 or more adverse events occurred during the APAG therapy, no thrombotic events occurred, no fibrologic tissue hyperplasia was found in the bone marrow pathology reexamination at 6 months of treatment, and none of the patients discontinued the drug due to adverse events.Conclusion:APAG may be a better switching treatment option for patients with AA who are refractory or are intolerant to TPO-RA.

Objective:To evaluate the safety and effectiveness of a new Chinese-made surgical biopatch for atrial septum under the establishment of atrial septal defect animal model in miniature pigs.Methods:From June 2018 to April 2019, 26 pigs were divided into experimental group (15 pigs) and the control group (11 pigs). Animal models of atrial septal defect were established by traditional surgical methods. The to-be-evaluated and listed surgical biological patches (with a diameter of 10 mm) were implanted in the experimental group and the control group to repair the atrial septal defect. Cardiac ultrasound and blood examination of all animals were performed before and at 7, 30, 90, 180 days after operation, the results were analyzed with repetitive measurement and analysis of variance. At 90 days and 180 days after the operation, tissue samples were taken from animals after euthanasia. Pathological examination of heart and major organs were conducted. The independent sample t test and rank sum test were used to compare the data between the two groups, and the nonparametric was used to compare the patch calcification score between the two groups. Results:In total of 26 animals, 14 animals in the experimental group(6 at 90 days, 8 at 180 days) and 9 animals in the control group(4 at 90 days, 5 at 180 days) reached the end of the experiment. The other 3 animals (1 in the experimental group and 2 in the control group) died of arrhythmia, whole heart failure and right heart failure, the results of pathological examination showed that the causes of death were unrelated to the experimental materials. Cardiac ultrasound showed no patch leakage in all animals. There was no statistically significant difference in cardiac ultrasound and blood examination between the two groups at different time points after operation (all P>0.05). The pathological results showed that all the implants were intact and had good biocompatibility. There was no significant difference in the mean endothelialization rate between the experimental group and the control group at 90 and 180 days after operation ((80.8±29.1)% vs. (82.5±23.6)%, t=0.095, P=0.927; (78.8±36.4)% vs. (82.0±19.2)%, t=0.182, P=0.859) on 90 and 180 days, there was no significant difference in the patch calcification score between the two groups (1.00(1.25) vs. 2.00(0.75), Z=6.500, P=0.214; 0(0.75) vs. 1.00(2.00), Z=12.000, P=0.139). Conclusion:The new Chinese-made surgical biopatch for atrial septum has comparable safety and efficacy to that of the marketable patch in miniature pig atrial septal defect animal model.

Objective:To evaluate the safety and effectiveness of a new Chinese-made surgical biopatch for atrial septum under the establishment of atrial septal defect animal model in miniature pigs.Methods:From June 2018 to April 2019, 26 pigs were divided into experimental group (15 pigs) and the control group (11 pigs). Animal models of atrial septal defect were established by traditional surgical methods. The to-be-evaluated and listed surgical biological patches (with a diameter of 10 mm) were implanted in the experimental group and the control group to repair the atrial septal defect. Cardiac ultrasound and blood examination of all animals were performed before and at 7, 30, 90, 180 days after operation, the results were analyzed with repetitive measurement and analysis of variance. At 90 days and 180 days after the operation, tissue samples were taken from animals after euthanasia. Pathological examination of heart and major organs were conducted. The independent sample t test and rank sum test were used to compare the data between the two groups, and the nonparametric was used to compare the patch calcification score between the two groups. Results:In total of 26 animals, 14 animals in the experimental group(6 at 90 days, 8 at 180 days) and 9 animals in the control group(4 at 90 days, 5 at 180 days) reached the end of the experiment. The other 3 animals (1 in the experimental group and 2 in the control group) died of arrhythmia, whole heart failure and right heart failure, the results of pathological examination showed that the causes of death were unrelated to the experimental materials. Cardiac ultrasound showed no patch leakage in all animals. There was no statistically significant difference in cardiac ultrasound and blood examination between the two groups at different time points after operation (all P>0.05). The pathological results showed that all the implants were intact and had good biocompatibility. There was no significant difference in the mean endothelialization rate between the experimental group and the control group at 90 and 180 days after operation ((80.8±29.1)% vs. (82.5±23.6)%, t=0.095, P=0.927; (78.8±36.4)% vs. (82.0±19.2)%, t=0.182, P=0.859) on 90 and 180 days, there was no significant difference in the patch calcification score between the two groups (1.00(1.25) vs. 2.00(0.75), Z=6.500, P=0.214; 0(0.75) vs. 1.00(2.00), Z=12.000, P=0.139). Conclusion:The new Chinese-made surgical biopatch for atrial septum has comparable safety and efficacy to that of the marketable patch in miniature pig atrial septal defect animal model.