Intervertebral disc degeneration (IDD) is largely attributed to impaired endogenous repair. Nucleus pulposus-derived stem cells (NPSCs) senescence leads to endogenous repair failure. Small extracellular vesicles/exosomes derived from mesenchymal stem cells (mExo) have shown great therapeutic potential in IDD, while whether mExo could alleviate NPSCs senescence and its mechanisms remained unknown. We established a compression-induced NPSCs senescence model and rat IDD models to evaluate the therapeutic efficiency of mExo and investigate the mechanisms. We found that mExo significantly alleviated NPSCs senescence and promoted disc regeneration while knocking down thioredoxin (TXN) impaired the protective effects of mExo. TXN was bound to various endosomal sorting complex required for transport (ESCRT) proteins. Autocrine motility factor receptor (AMFR) mediated TXN K63 ubiquitination to promote the binding of TXN on ESCRT proteins and sorting of TXN into mExo. Knocking down exosomal TXN inhibited the transcriptional activity of nuclear factor erythroid 2-related factor 2 (NRF2) and activator protein 1 (AP-1). NRF2 and AP-1 inhibition reduced endogenous TXN production that was promoted by exosomal TXN. Inhibition of NRF2 in vivo diminished the anti-senescence and regenerative effects of mExo. Conclusively, AMFR-mediated TXN ubiquitination promoted the sorting of TXN into mExo, allowing exosomal TXN to promote endogenous TXN production in NPSCs via TXN/NRF2/AP-1 feed-forward circuit to alleviate NPSCs senescence and disc degeneration.

BACKGROUND:Allogeneic tendons are often used in clinical treatment of tendon and ligament injuries,which need to be treated before clinical application.At present,allogeneic tendons will face multiple freeze-thaw cycles after cryopreservation,but there is no relevant research on the changes in the properties of the treated allogeneic tendon after several freeze-thaw cycles.OBJECTIVE:To explore the changes in mechanical properties and composition of allogeneic tendon after multiple freeze-thaw cycles.METHODS:The treated allogeneic tendon was frozen and thawed 3,6,and 10 times at-40 ℃(or-80 ℃),respectively.The mechanical test,DNA content,glycosaminoglycan sulfate,hydroxyproline content and total protein content of the allogeneic tendon after freezing and thawing were detected.RESULTS AND CONCLUSION:(1)There was no significant difference in the maximum load,maximum load elongation,tensile strength and elastic modulus between groups after freezing-thawing cycles for 3,6,and 10 times at-40 ℃(P＞0.05).There was no significant difference in the maximum load and maximum load elongation between the groups after freezing-thawing cycles for 3,6,and 10 times at-80 ℃(P＞0.05).The tensile strength of the group with 3 cycles of freezing and thawing was higher than that of the groups with 6 and 10 cycles of freezing and thawing(P＜0.05).The elastic modulus of the group with 3 cycles of freezing and thawing was higher than that of the group with 10 cycles of freezing and thawing(P＜0.05).(2)The freezing and thawing times did not affect the contents of DNA,hydroxyproline,total protein,and sulfated glycosaminoglycan in allogeneic tendon when stored at-40 ℃(or-80 ℃).(3)These findings suggest that in-40℃ storage,the performance of the allogeneic tendon should not be affected by taking out and putting back the tendon less than 10 times,while in-80 ℃ storage,the allogeneic tendon should be taken out and put back less than 6 times.

Energy metabolism disorders are a critical factor contributing to cognitive dysfunction.Astrocytes,as key suppliers of energy substrates to neurons,help to sustain normal cognitive function via their extensive energy metabolism activities in the brain,coordinated by various energy supply mechanisms.Imbalances in astrocyte-neuron energy metabolism coupling lead to abnormal neuronal activity,thereby driving the pathogenesis and progression of Alzheimer's disease(AD).It is therefore essential to investigate the pathological mechanisms and therapeutic targets associated with AD from the perspective of energy metabolism to support the future development of AD treatments and interventions.Traditional Chinese medicine(TCM),characterized by multi-component and multi-target effects,has shown potential efficacy for regulating cerebral energy metabolism,suggesting that modulating brain energy metabolism may represent a significant pathway by which TCM might ameliorate AD.This review systematically elucidates the mechanistic role of the dysregulation of astrocyte-neuron bioenergetic coupling in the pathogenesis and progression of AD,and critically evaluates recent advancements in therapeutic interventions mediated by TCM.

BACKGROUND:Natural biomaterials such as allogeneic materials are highly susceptible to bacterial contamination during the production process.Among them,bacterial endotoxin released by Gram-negative bacteria can cause human fever,coagulation,shock,and even death.There is no relevant research on the method of endotoxin removed from allogeneic materials while maintaining their natural structure and biological activity.OBJECTIVE:To explore the method and effect of endotoxin removed from allogeneic materials and other natural biological materials and to control the residual H2O2.METHODS:The endotoxin was removed from allogeneic bone materials by H2O2/ultrasound.The influence of ultrasound and the concentration of H2O2 on the endotoxin removal process was explored.The concentration of H2O2 was set as 1％,2％,3％,4％,and 5％by mass.The power of ultrasound was set as 80,120,140,160,and 200 W.The duration of ultrasound was 20,30,40,50,and 60 minutes.The temperature was 30,35,40,45,and 50 ℃.The optimal combination of conditions for treating allogeneic bone materials was selected by endotoxin content detection for subsequent experiments.The microstructure,H2O2 residue,and cytotoxicity of allogeneic bone materials after endotoxin removal were detected,and the effects of different concentrations of H2O2 residue on the survival rate of L929 mouse fibroblasts were explored.RESULTS AND CONCLUSION:(1)According to the results of the endotoxin removal process,when the concentration of H2O2 was 4％,the power of ultrasound was 200 W,with the duration of ultrasound of 50 minutes,and the temperature 50 ℃,it would effectively remove endotoxin from allogeneic bone materials.The optimal combination was selected to remove endotoxins from allogeneic bone materials.(2)After endotoxin removal,the residual H2O2 content of allogeneic bone materials was less than 47.551 mg/kg after being placed in a 60 ℃ water bath and changing the solution 6 times,which had no effect on the viability of L929 mouse fibroblasts.(3)Optical microscopy and scanning electron microscopy observations exhibited that the use of H2O2 solution in conjunction with ultrasonic treatment had no effect on the structure of allogeneic bone materials.(4)The results indicate that the H2O2 solution in conjunction with ultrasonic treatment has the characteristics of efficient and rapid removal of endotoxins,which can ensure the safety of allogeneic bone materials.

Energy metabolism disorders are a critical factor contributing to cognitive dysfunction.Astrocytes,as key suppliers of energy substrates to neurons,help to sustain normal cognitive function via their extensive energy metabolism activities in the brain,coordinated by various energy supply mechanisms.Imbalances in astrocyte-neuron energy metabolism coupling lead to abnormal neuronal activity,thereby driving the pathogenesis and progression of Alzheimer's disease(AD).It is therefore essential to investigate the pathological mechanisms and therapeutic targets associated with AD from the perspective of energy metabolism to support the future development of AD treatments and interventions.Traditional Chinese medicine(TCM),characterized by multi-component and multi-target effects,has shown potential efficacy for regulating cerebral energy metabolism,suggesting that modulating brain energy metabolism may represent a significant pathway by which TCM might ameliorate AD.This review systematically elucidates the mechanistic role of the dysregulation of astrocyte-neuron bioenergetic coupling in the pathogenesis and progression of AD,and critically evaluates recent advancements in therapeutic interventions mediated by TCM.

BACKGROUND:Natural biomaterials such as allogeneic materials are highly susceptible to bacterial contamination during the production process.Among them,bacterial endotoxin released by Gram-negative bacteria can cause human fever,coagulation,shock,and even death.There is no relevant research on the method of endotoxin removed from allogeneic materials while maintaining their natural structure and biological activity.OBJECTIVE:To explore the method and effect of endotoxin removed from allogeneic materials and other natural biological materials and to control the residual H2O2.METHODS:The endotoxin was removed from allogeneic bone materials by H2O2/ultrasound.The influence of ultrasound and the concentration of H2O2 on the endotoxin removal process was explored.The concentration of H2O2 was set as 1％,2％,3％,4％,and 5％by mass.The power of ultrasound was set as 80,120,140,160,and 200 W.The duration of ultrasound was 20,30,40,50,and 60 minutes.The temperature was 30,35,40,45,and 50 ℃.The optimal combination of conditions for treating allogeneic bone materials was selected by endotoxin content detection for subsequent experiments.The microstructure,H2O2 residue,and cytotoxicity of allogeneic bone materials after endotoxin removal were detected,and the effects of different concentrations of H2O2 residue on the survival rate of L929 mouse fibroblasts were explored.RESULTS AND CONCLUSION:(1)According to the results of the endotoxin removal process,when the concentration of H2O2 was 4％,the power of ultrasound was 200 W,with the duration of ultrasound of 50 minutes,and the temperature 50 ℃,it would effectively remove endotoxin from allogeneic bone materials.The optimal combination was selected to remove endotoxins from allogeneic bone materials.(2)After endotoxin removal,the residual H2O2 content of allogeneic bone materials was less than 47.551 mg/kg after being placed in a 60 ℃ water bath and changing the solution 6 times,which had no effect on the viability of L929 mouse fibroblasts.(3)Optical microscopy and scanning electron microscopy observations exhibited that the use of H2O2 solution in conjunction with ultrasonic treatment had no effect on the structure of allogeneic bone materials.(4)The results indicate that the H2O2 solution in conjunction with ultrasonic treatment has the characteristics of efficient and rapid removal of endotoxins,which can ensure the safety of allogeneic bone materials.

BACKGROUND:Allogeneic tendons are often used in clinical treatment of tendon and ligament injuries,which need to be treated before clinical application.At present,allogeneic tendons will face multiple freeze-thaw cycles after cryopreservation,but there is no relevant research on the changes in the properties of the treated allogeneic tendon after several freeze-thaw cycles.OBJECTIVE:To explore the changes in mechanical properties and composition of allogeneic tendon after multiple freeze-thaw cycles.METHODS:The treated allogeneic tendon was frozen and thawed 3,6,and 10 times at-40 ℃(or-80 ℃),respectively.The mechanical test,DNA content,glycosaminoglycan sulfate,hydroxyproline content and total protein content of the allogeneic tendon after freezing and thawing were detected.RESULTS AND CONCLUSION:(1)There was no significant difference in the maximum load,maximum load elongation,tensile strength and elastic modulus between groups after freezing-thawing cycles for 3,6,and 10 times at-40 ℃(P＞0.05).There was no significant difference in the maximum load and maximum load elongation between the groups after freezing-thawing cycles for 3,6,and 10 times at-80 ℃(P＞0.05).The tensile strength of the group with 3 cycles of freezing and thawing was higher than that of the groups with 6 and 10 cycles of freezing and thawing(P＜0.05).The elastic modulus of the group with 3 cycles of freezing and thawing was higher than that of the group with 10 cycles of freezing and thawing(P＜0.05).(2)The freezing and thawing times did not affect the contents of DNA,hydroxyproline,total protein,and sulfated glycosaminoglycan in allogeneic tendon when stored at-40 ℃(or-80 ℃).(3)These findings suggest that in-40℃ storage,the performance of the allogeneic tendon should not be affected by taking out and putting back the tendon less than 10 times,while in-80 ℃ storage,the allogeneic tendon should be taken out and put back less than 6 times.

OBJECTIVE To investigate the effects of borneol on pharmacodynamic and pharmacokinetic effects of Corydalis saxicola total alkaloids in depression model rats.METHODS Thirty male SD rats were divided into blank control group,negative control group,positive control group (fluoxetine 10 mg/kg,i.g.),single drug group (C.saxicola total alkaloids 210 mg/kg,i.g.) and combined drug group (C.saxicola total alkaloids 210 mg/kg+borneol 50 mg/kg,i.g.) according to the random number table method,with 6 rats in each group.By lipopolysaccharide (LPS) induction modeling,except blank control group (no model and no administration) received intraperitoneal injection of the same amount of normal saline,the rats in the other groups were intraperitoneally injected with LPS once a day to establish a rat model of depression.After 1 week of modeling,each administration group was given relevant drug intragastrically according to the corresponding dose,and blank control group and negative control group (without drug treatment) were administered intragastrically with an equal volume of solvent to dissolve the drug;continued modeling while administering the drug.After two weeks of continuous administration,the effects of C.saxicola total alkaloids versus the combination of C.saxicola total alkaloids and borneol on the behavior of depressed rats were tested by behavioral experiments;the levels of tumor necrosis factor-α,interleukin-1β and interleukin-6 in rats were determined;the histopathological changes of the hippocampus of rats were observed.Blood sample was collected from the orbit at different time points after administration on the 15th day,and the upper plasma was obtained.Ultra-performance liquid chromatography-triple quadrupole tandem mass spectrometry was established for the simultaneous determination of dehydrocarvedine,tetrahydropalmatine,coptisine,palmatine,jatrorrhizine,berberine,berberrubine and epiberberine in rat plasma.The average plasma concentration-time curve was depicted,the area under the curve (AUC) was calculated,and the pharmacokinetic parameters were analyzed by DAS 3.2.2 software.RESULTS Compared with blank control group,the negative control group had a decrease in body mass and sugar water preference rate,a decrease in the total distance of open field,a prolonged swimming immobility time,and a increased in the expression of inflammatory factors in serum (P<0.05);compared with negative control group,the single drug group and the combined drug group increased the preference rate of sugar water,increased the total distance of open field,shortened the time of swimming immobility,and decreased the expression of inflammatory factors in serum (P<0.05).There was no significant difference in the above indicators between the single drug group and the combined drug group in rats (P>0.05).Pharmacokinetic results showed that compared with single drug group,AUC0-t of coptisine,AUC0-t,AUC0-∞,tmax and cmax of jatrorrhizine,AUC0-t,AUC0-∞,t1/2 and cmax of berberrubine,and AUC0-t of epiberberine,cmax of dehydrocarvedine,cmax of palmatine were significantly increased in combined drug group,but there was no significant difference,indicating that borneol didn't have a significant effect on the efficacy of Corydalis saxicola nigra at this dose.CONCLUSIONS Both C.saxicola total alkaloids alone and in combination with borneol can improve depression-like behavior in depression model rats,reduce serum inflammatory cytokine levels,and protect hippocampal neurons.Compared with the use of Corydalis saxicola base alone,the combination with borneol do not show significant pharmacodynamic differences,bu can improve the absorption of coptisine,jatrorrhizine in model rats.

ObjectiveTo investigate the pharmacodynamics， pharmacokinetics and brain distribution of seven main components of Jiaotaiwan alone and Jiaotaiwan combined with borneol in lipopolysaccharide（LPS）-induced depression rats. MethodRats were randomly divided into the control group， model group， fluoxetine group（10 mg·kg^-1）， Jiaotaiwan group（1.5 g·kg^-1） and combination group（1.5 g·kg^-1 of Jiaotaiwan+0.05 g·kg^-1 of borneol）， with 8 rats in each group. Except for the control group， the depression model was established by intraperitoneal injection of LPS for 7 consecutive days， and each group was given the corresponding drugs or the same volume of pure water by gavage for 14 consecutive days. The behavioral indicators and levels of serum inflammatory factors［interleukin-1β（IL-1β）， IL-6 and tumor necrosis factor-α（TNF-α）］ of rats in each group were compared. The morphological changes of hippocampal neurons were observed by hematoxylin-eosin（HE） and Nissl staining. After the behavioral tests of sucrose preference， open field and forced swimming were completed， blood samples were collected from Jiaotaiwan group and combination group at the set time points after gavage， the contents of seven components in plasma were determined by ultra performance liquid chromatography-triple quadrupole tandem mass spectrometry（UPLC-QqQ-MS/MS）， and the pharmacokinetic parameters of each component were analyzed by DAS 3.2.2. Brains were rapidly removed on ice after blood collection was completed， and UPLC-QqQ-MS/MS was used to compare the content changes of the 7 components in brain tissue. ResultCompared with the control group， rats in the model group showed decreased sucrose preference rate and total distance of open field， prolonged swimming immobility time， and increased expression of inflammatory factors in serum（P<0.01）. Compared with the model group， the sucrose preference rate and total distance of open field were increased， and the swimming immobility time was shortened in the rats of each administration group， and the expression of inflammatory factors in serum was decreased in rats from Jiaotaiwan group and combination group（P<0.05， P<0.01）. The results of HE and Nissl staining showed that Jiaotaiwan group and combination group had a certain protective effect on hippocampal neurons. The pharmacokinetic results showed that compared with Jiaotaiwan group， the area under the curve（AUC_0-t， AUC_0-∞）， peak concentration（C_max） and the average steady-state plasma concentration（C_av） of berberine and epiberberine in the combination group were increased（P<0.05， P<0.01）， AUC_0-t， AUC_0-∞， mean residence time（MRT_0-∞） and C_av of coptisine were significantly increased（P<0.05， P<0.01）， C_max of jatrorrhizine increased significantly（P<0.05）， but the pharmacokinetic changes of the seven alkaloids were consistent. The results of brain tissue distribution showed that compared with Jiaotaiwan group， the contents of jatrorrhizine， epiberberine， coptisine， palmatine and berberine in the brain tissue of combination group were increased（P<0.05， P<0.01）， the content of magnoflorine increased and that of berberrubine decreased， but the difference was not statistically significant. ConclusionJiaotaiwan alone or combined with borneol can improve the depression-like behavior of rats， reduce the levels of serum inflammatory factors， improve the pathological damage of hippocampus， and have antidepressant effect. Compared with Jiaotaiwan alone， the combination of Jiaotaiwan and borneol can increase the exposure of multiple active components of Jiaotaiwan in the plasma and brain tissue of rats， improve its bioavailability， promote its absorption， and improve the brain targeting of the drug， which is more conducive to the antidepressant effect of Jiaotaiwan.

ObjectiveTo investigate the effect of total alkaloids of Corydalis saxicola on a rat model of lipopolysaccharide（LPS）-induced depression， as well as the pharmacokinetic characteristics of 8 of its major components. MethodTwenty-four male SD rats were randomly divided into normal group， model group， fluoxetine group（10 mg·kg^-1） and total alkaloids of C. saxicola group（210 mg·kg^-1）， with 6 rats in each group. In addition to the normal group， the rats were injected intraperitoneally with LPS to establish the inflammation model of depression， and the drug administration was started 1 week after modeling， and the administration groups were gavaged according to the corresponding dose， and the normal and model groups were intragastric administration with equal volume of distilled water， and the administration was performed along with the modeling. After two weeks of continuous administration， the effect of total alkaloids of C. saxicola on the behavior of depressed rats were tested by sucrose preference， forced swimming and open field experiments， the levels of tumor necrosis factor-α（TNF-α）， interleukin（IL）-1β and IL-6 in serum of rats were determined by enzyme-linked immunosorbent assay（ELISA）， the histopathological changes of rat hippocampus were observed by hematoxylin-eosin（HE） staining. After the last administration， blood was collected from orbit according to the set time， and ultra-high performance liquid chromatography-triple quadrupole tandem mass spectrometry（UPLC-QqQ-MS） was established to simultaneously detect the concentrations of dehydrocavidine， tetrahydropalmatine， coptisine， palmatine， jatrorrhizine， berberine， berberrubine and epiberberine in plasma， and drug-time curves were drawn. The pharmacokinetic parameters were analyzed by DAS 2.0 software. ResultCompared with the normal group， the model group exhibited a decrease in sucrose preference rate， total distance traveled in the open field， as well as an increase in swimming immobility time and serum inflammatory factor expression（P<0.01）. In contrast， compared with the model group， rats in each administration group showed an increase in sucrose preference rate and total distance traveled in the open field， a decrease in swimming immobility time， and a reduction in serum inflammatory factor expression（P<0.05， P<0.01）. Additionally， HE staining results revealed that neurons in the hippocampus of rats from the model group were characterized by loss， disorganization and residual vacuoles， whereas those from the total alkaloids of C.saxicola group displayed an increase in number with orderly arrangement and clear cytoplasm. Pharmacokinetic results showed that the time to peak（t_max） and half-life（t_1/2） of the 8 active ingredients were 0.19-2.06 h and 3.71-8.70 h after continuous administration of total alkaloids of C. saxicola. Among them， the area under the curve（AUC_0-∞） of tetrahydropalmatine was the highest and the t_1/2 was the shortest， and the AUC_0-∞ of coptisine， palmatine， jatrorrhizine， berberine， berberrubine and epiberberine were low. The curves of dehydrocavidine， coptisine， palmatine， berberine and epiberberine showed obvious double peak phenomenon. ConclusionTotal alkaloids of C. saxicola can improve the depression-like behavior of rats， inhibit the expression of inflammatory factors in serum， improve the pathological injury of hippocampus， and has the antidepressant effect. Meanwhile， the effective site is absorbed quickly and eliminated slowly in the depressed model rats， and the efficacy is maintained for a long time.