BACKGROUND:Psoralen has a strong anti-osteoporotic activity and may have a restorative effect on chemotherapy-induced osteoporosis. OBJECTIVE:To explore the restorative effect of psoralen on the osteogenic differentiation of bone marrow mesenchymal stem cells in mice inhibited by cyclophosphamide and its mechanism. METHODS:C57BL/6 mouse bone marrow mesenchymal stem cells were isolated and cultured.Effect of psoralen on viability of bone marrow mesenchymal stem cells was detected by MTT assay.Osteogenic induction combined with alkaline phosphatase staining was used to determine the optimal dose of psoralen to restore the osteogenic differentiation of bone marrow mesenchymal stem cells inhibited by cyclophosphamide.The mRNA expression levels of Runx2,alkaline phosphatase,Osteocalcin,osteoprotegerin,and Wnt/β-catenin signaling pathway-related genes Wnt1,Wnt4,Wnt10b,β-catenin,and c-MYC were measured by RT-qPCR at different time points under the intervention with psoralen.The protein expression of osteogenic specific transcription factor Runx2 and Wnt/β-catenin signaling pathway related genes Active β-catenin,DKK1,c-MYC,and Cyclin D1 was determined by western blot assay at different time points under the intervention with psoralen. RESULTS AND CONCLUSION:(1)There was no significant effect of different concentrations of psoralen on the viability of bone marrow mesenchymal stem cells.The best recovery of the inhibition of osteogenic differentiation of bone marrow mesenchymal stem cells caused by cyclophosphamide was under the intervention of psoralen at a concentration of 200 μmol/L.(2)Psoralen reversed the reduction in osteogenic differentiation marker genes Runx2,alkaline phosphatase,Osteocalcin and osteoprotegerin mRNA expression and Runx2 protein expression in bone marrow mesenchymal stem cells caused by cyclophosphamide conditioned medium.(3)Psoralen reversed the decrease in Wnt/β-catenin pathway-related genes Wnt4,β-catenin,c-MYC mRNA and Active β-catenin,c-MYC,and Cyclin D1 protein expression and the increase in DKK1 protein expression in bone marrow mesenchymal stem cells caused by cyclophosphamide conditioned medium.(4)The results showed that cyclophosphamide inhibited osteogenic differentiation of bone marrow mesenchymal stem cells in mice,and psoralen had a restorative effect on it.The best intervention effect was achieved at a concentration of 200 μmol/L psoralen,and this protective effect might be related to the activation of Wnt4/β-catenin signaling pathway by psoralen.

BACKGROUND:Anthocyanin is one of the most important active components in Lycium ruthenicum Murr,which has antioxidant and immunomodulatory effects.CD146+human adipose-derived pericytes/perivascular cells(CD146+hAD-PCs)are the progenitors of bone marrow mesenchymal stem cells,which can promote the proliferation and differentiation of hematopoietic stem/progenitor cells in vitro.The support effect of anthocyanin in combination with CD146+hAD-PCs on umbilical cord blood hematopoietic stem/progenitor cells remains to be studied. OBJECTIVE:To investigate the supporting effect of anthocyanins in Lycium ruthenicum Murr(ALRM)combined with CD146+hAD-PCs on umbilical cord blood CD34+hematopoietic stem/progenitor cells(UCB CD34+HSPCs)in vitro. METHODS:The CCK-8 assay was used to detect the effect of different concentrations(0,200,400,600,800,1 000 mg/L)of ALRM on the proliferation of CD146+hAD-PCs.Flow cytometry was used to detect the effect of ALRM on the cell cycle of CD146+hAD-PCs.The co-culture experiments were divided into blank group,ALRM group,CD146+hAD-PCs group,and ALRM+CD146+hAD-PCs group to analyze the in vitro supporting effect of ALRM combined with CD146+hAD-PCs on UCB CD34+HSPCs.The number of expanded cells and the number of colony-forming units were compared at 1,2,and 4 weeks of co-culture.The immunophenotype of cells was detected by flow cytometry.The level of cytokines was detected by enzyme-linked immunosorbent assay. RESULTS AND CONCLUSION:(1)The cell viability of CD146+hAD-PCs was highest at an ALRM concentration of 200 mg/L,the proportion of G0/G1 phase cells decreased and the proportion of S and G2/M phase cells increased in CD146+hAD-PCs(P<0.01).(2)The change in number of UCB CD34+HSPCs cells in the ALRM+CD146+hAD-PCs group was higher than that in the ALRM group at 1,2,and 4 weeks of co-culture(all P<0.05),and higher than that in CD146+hAD-PCs group at 2 and 4 weeks of co-culture(all P<0.05).The number of cells in the ALRM group and blank group decreased gradually with the extension of co-culture time.(3)Colony forming capacity and immunophenotype analysis:The number of colony-forming units in the ALRM+CD146+hAD-PCs group was higher than that in the CD146+hAD-PCs group and ALRM group at 1 and 2 weeks of co-culture(P<0.05).The proportion of CD45+and CD34+CD33-cells in the ALRM+CD146+hAD-PCs group was higher than that in the CD146+hAD-PCs group at 1 and 2 weeks of co-culture(all P<0.01).(4)Changes in cytokines:Interleukin-2 level in the ALRM+CD146+hAD-PCs group was higher than that in the ALRM and CD146+hAD-PCs groups(P<0.05).The interleukin-3 content of the ALRM+CD146+hAD-PCs group was higher than that of the CD146+hAD-PCs group at 2 and 4 weeks(P<0.05).The expression level of granulocyte colony-stimulating factor in the ALRM+CD146+hAD-PCs group was higher than that in the CD146+hAD-PCs group at 1 week,and higher than that in the ALRM group and CD146+hAD-PCs group at 2 weeks(P<0.01).Interferon-γ content in the ALRM group and ALRM+CD146+hAD-PCs group was lower than that in the CD146+hAD-PCs group at 1,2,and 4 weeks of co-culture(P<0.01).(5)Due to the absence of stromal cells in the blank group,UCB CD34+HSPCs could not be counted after 1 week of co-culture and were not subjected to immunophenotyping,colony analysis,or cytokine assays.(6)In summary,ALRM can promote the expansion of UCB CD34+HSPCs in vitro by promoting CD146+hAD-PCs proliferation and cell cycle transformation,which is of great value in hematopoietic stem cell transplantation.

BACKGROUND:It has been found that abnormal apoptosis of bone tissue cells induced by abnormal iron metabolism plays an important role in the progression of osteoporosis. OBJECTIVE:To investigate the effect of naringin on iron metabolism and cell apoptosis in bone tissue of rats with osteoporosis. METHODS:Fifty 2-month-old female Sprague-Dawley rats were randomly divided into five groups with 10 rats in each group:sham group,osteoporosis group,naringin low-dose group,naringin high-dose group,and naringin high-dose+DKK-1 group.Except for the sham group,rat models of osteoporosis were established by removing bilateral ovarian tissues in the other groups.At 8 weeks after modeling,rats in the naringin low-and high-dose groups were given 100 and 400 mg/kg/d naringenin by gavage,respectively,and rats in the naringenin high dose+DKK-1 group were given 400 mg/kg/d naringin by gavage and subcutaneous injection of 25 mg/kg/d DKK-1,an inhibitor of the Wnt1 signaling pathway,for 7 consecutive days.Relevant indexes were detected after administration. RESULTS AND CONCLUSION:Compared with the osteoporosis group,naringin could enhance the bone mineral density and serum calcium and superoxide dismutase levels in rats(P<0.05),and reduce the serum levels of osteocalcin,malondialdehyde,and phosphorus(P<0.05),while DKK-1 could partially inhibit the interventional effect of naringin(P<0.05).Results from Micro-CT scanning,hematoxylin-eosin and TUNEL staining showed that compared with the osteoporosis group,naringin significantly improved bone microstructure and reduced the rate of cell apoptosis,while DKK-1 partially inhibited the interventional effect of naringin.Immunofluorescence staining results showed that compared with the osteoporosis group,naringin could reduce the oxygen content,anti-tartaric acid phosphatase expression,and elevate the expression of alkaline phosphatase in active tibia tissues(P<0.05),while DKK-1 could partially inhibit the interventional effect of naringin(P<0.05).Results from Prussian blue staining and immunohistochemical staining showed that compared with the osteoporosis group,naringin reduced iron deposition in bone and liver tissues as well as the expression of transferrin receptor 1(P<0.05),and elevated the protein expression of ferroportin 1(P<0.05)in bone tissue,and DKK-1 partially inhibited the intervention of naringin(P<0.05).PCR and western blot assay of tibia specimens showed that compared with the osteoporosis group,naringin decreased the expression of anti-tartrate acid phosphatase,transferrin receptor 1 and Bax(P<0.05),and elevated the expression of alkaline phosphatase,ferroportin 1,Bcl-2,Wnt1 and β-catenin(P<0.05),while DKK-1 partially inhibited the interfering effect of naringin(P<0.05).To conclude,naringin inhibits the progression of osteoporosis by reducing iron deposition and apoptosis rate in bone tissue,which may be related to the activation of the Wnt1 signaling pathway.

BACKGROUND:Previous studies have found that neohesperidin can delay bone loss in ovariectomized mice and has the potential to treat osteoporosis,but its specific mechanism of action remains to be explored. OBJECTIVE:To explore the key targets and possible mechanisms of neohesperidin in the treatment of osteoporosis based on bioinformatics and cell experiments in vitro. METHODS:The gene expression dataset related to osteoporosis was obtained from GEO database,and the differentially expressed genes were screened and analyzed in R language.The osteoporosis-related targets were screened from GeneCards and DisGeNET databases,and the neohesperidin-related targets were screened from ChEMBL and PubChem databases,and the common targets were obtained by intersection of the three.The String database was used to construct the PPI network of intersection genes,and the key targets were screened.The DAVID database was used for GO and KEGG enrichment analysis.The AutoDock software was used to verify the molecular docking between the neohesperidin and the target protein.The effect of neohesperidin on osteogenic differentiation of C57 mouse bone marrow mesenchymal stem cells was detected.Complete medium was used as blank control group;osteogenic induction medium was used as the control group;and osteogenic induction medium containing different concentrations of neohesperidin(25,50 μmol/L)was used as experimental group.The expression of alkaline phosphatase,the degree of mineralization,the expression of osteogenic-related genes and target genes during osteogenic differentiation of cells were measured at corresponding time points. RESULTS AND CONCLUSION:(1)9 253 differentially expressed genes,2 161 osteoporosis-related targets,and 326 neohesperidin-related targets were screened.There were 53 common targets among the three.All 53 genes were up-regulated in osteoporosis samples.The PPI network screened the target gene PRKACA of research significance.GO function and KEGG pathway enrichment analysis showed that neohesperidin's treatment of osteoporosis through PRKACA target mainly depended on biological processes such as protein phosphorylation and protein autophosphorylation,acting on endocrine resistance,proteoglycan in cancer,and estrogen signaling pathway to play a therapeutic role.Molecular docking results showed that neohesperidin had a certain binding ability to the protein corresponding to the target PRKACA.(2)The results of alkaline phosphatase staining showed that neohesperidin could promote the expression of alkaline phosphatase in the early stage of osteogenic differentiation of mesenchymal stem cells.Alizarin red staining showed that neohesperidin could promote the mineralization of osteogenic differentiation of mesenchymal stem cells.RT-qPCR results showed that neohesperidin could increase the mRNA expression of alkaline phosphatase,PRKACA,and osteocalcin.(3)These results indicate that neohesperidin may promote osteogenic differentiation through PRKACA target on the estrogen signaling pathway to prevent and treat osteoporosis.

BACKGROUND:Physiotherapy is very important for the recovery after anterior cruciate ligament reconstruction.In recent years,many doctors are optimizing the physical rehabilitation program after anterior cruciate ligament reconstruction.However,there is still a lack of efficient rehabilitation training after anterior cruciate ligament reconstruction. OBJECTIVE:To investigate the effect of vibration therapy combined with suspension training on movement and knee joint function after anterior cruciate ligament reconstruction. METHODS:A total of 80 patients undergoing first unilateral anterior cruciate ligament reconstruction at the Affiliated Sport Hospital,Shanghai University of Sport were randomly divided into vibration therapy group(n=40)and vibration therapy+suspension training group(n=40).In the vibration therapy group,vibration therapy(10 minutes each,once a day,6 times per week)was performed at the 13th week after anterior cruciate ligament reconstruction.Patients in the vibration therapy+suspension training group were treated with vibration therapy(10 minutes each,once a day,6 times per week)and suspension training(twice a week)at the 13th week after anterior cruciate ligament reconstruction.Training in each group was performed for 8 weeks.Knee joint function was evaluated by knee joint Lysholm score before and 8 weeks after training.The symmetry index was evaluated by the isokinetic muscle strength evaluation training system.The balance test system was used to evaluate the average trace error difference of the bilateral multi-axes. RESULTS AND CONCLUSION:Compared with those before training,the knee Lysholm score and the knee extension and flexion symmetry indexes increased(P<0.05),and the average trace error difference decreased after training(P<0.05).Compared with the vibration therapy group,the knee Lysholm score in the vibration therapy+suspension training group increased(P<0.05),the knee extension and knee flexion symmetry index increased(P<0.05),and the average trace error difference decreased(P<0.05).To conclude,compared with vibration therapy training alone,vibration therapy combined with suspension training can significantly improve knee joint function,increase muscle strength and symmetry,and improve balance stability in patients undergoing anterior cruciate ligament reconstruction.

BACKGROUND:Our previous study found that Modified Erxian Pill could alleviate inflammation in collagen-induced arthritis rats,but its mechanism needs to be further verified. OBJECTIVE:To analyze the components absorbed in the blood of Modified Erxian Pill,and observe the effect of the drug-containing serum of Modified Erxian Pill on pyroptosis of J774A.1 macrophages. METHODS:(1)Analysis of components absorbed in the blood of Modified Erxian Pill:Ultra-high performance liquid chromatography-high resolution mass spectrometry was used to detect and identify Modified Erxian Pill and its components absorbed in the blood.(2)Effect of the drug-containing serum of Modified Erxian Pill on pyroptosis of J774A.1 macrophages:Molecular docking technology was used to initially verify the sesquiterpenoids and NLRP3 in components absorbed in the blood of Modified Erxian Pill.J774A.1 macrophages were randomly divided into blank control group,lipopolysaccharide+adenosine triphosphate group,and lipopolysaccharide+adenosine triphosphate+Modified Erxian Pill with low(2.5%),medium(5%),and high(10%)dose groups.The release of lactate dehydrogenase in the cell supernatant of each group was detected according to the kit instructions.The levels of interleukin-1β and interleukin-18 in cell supernatant were detected in each group by ELISA.The cell membrane damage was detected by Hoechst/PI staining.The expression levels of NLRP3,Caspase-1,GSDMD,and GSDMD-N protein in the cells of each group were detected by western blot assay. RESULTS AND CONCLUSION:(1)A total of 32 active components of Modified Erxian Pill were identified,and 21 components entered the blood.The main components into blood included a variety of sesquiterpenoids.(2)Molecular docking results showed that 3-O-Acetyl-13-deoxyphomenone,Incensol oxide,Atractylenolide III,Rupestonic acid,and 3,7-Dihydroxy-9,11-eremophiladien-8-one had good binding activity with NLRP3.(3)Compared with the blank control group,lactate dehydrogenase activity and the expression levels of interleukin-1β and interleukin-18 were significantly increased in cell supernatant of lipopolysaccharide+adenosine triphosphate group(P<0.001).Hoechst/PI staining showed that the number of PI-positive cells was significantly increased.After the intervention of lipopolysaccharide+adenosine triphosphate+Modified Erxian Pill group,all of them showed different degrees of reduction.(4)Compared with the blank control group,NLRP3,Caspase-1,GSDMD,and GSDMD-N protein expression levels were significantly increased in the lipopolysaccharide+adenosine triphosphate group(P<0.05).Compared with lipopolysaccharide+adenosine triphosphate group,the protein expressions of NLRP3,Caspase-1,GSDMD,and GSDMD-N were significantly decreased in the lipopolysaccharide+adenosine triphosphate+Modified Erxian Pill group(P<0.05),and had a certain dose dependence.These findings verify that the drug-containing serum of Modified Erxian Pill may inhibit the pyroptosis of J774A.1 macrophages by regulating the NLRP3/Caspase-1/GSDMD pathway.

The Huangdi Neijing proposes the " using bitter herbs to nourish or purge" theory to guide clinical prescription and formulation of herbal remedies based on the physiological characteristics and functions of the five zang viscera, along with the properties and flavors of medicinal herbs. This study explored diabetic kidney disease pathogenesis and treatment based on the " using bitter herbs to nourish or purge" theory. Kidney dryness is a key pathological factor in diabetic kidney disease, and the disharmony of kidney dryness is an essential aspect of its pathogenesis. Strengthening is the primary therapeutic principle, and kidney dryness is a persistent factor throughout the occurrence and progression of diabetic kidney disease. In the early stage, the pathogenesis involves heat-consuming qi and injuring yin, leading to kidney dryness. In the middle stage, the pathogenesis manifests as qi deficiency and blood stasis in the collaterals, resulting in turbidity owing to kidney dryness. In the late stage, the pathogenesis involves yin and yang deficiency, with kidney dryness and disharmony. This study proposes the staging-based treatment based on the " need for firmness" characteristic of the kidney. The aim is to provide new insights for clinical diagnosis and treatment in traditional Chinese medicine by rationally using pungent, bitter, and salty medicinal herbs to nourish and moisturize the kidney. This approach seeks to promote precise syndrome differentiation and personalized treatment for different stages of diabetic kidney disease, thereby enhancing clinical efficacy.

Objective : To explore the mechanism of isorhamnetin (Iso) in the treatment of oral squamous cell carcinoma (OSCC) using network pharmacology and molecular docking methods and to verify it in vitro. Methods : The key targets were obtained by constructing the PPI protein interaction network based on the common intersection targets of Iso-OSCC. At the same time, gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) were used to analyze the related signaling pathways of the intersection targets. Iso and core targets were also analyzed through molecular docking and visualization. Colony formation assay and Transwell assay were used to identify the effect of Iso on the proliferation and invasion of Cal-27 cells. Western blot was used to analyze the regulatory effects of different concentrations of Iso on estrogen receptor-1 (ESR1), phosphoinositide-3-kinase regulatory subunit-1 (PIK3R1), Src tyrosine kinase (SRC), and phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway proteins. Results: A total of 269 potential intersection targets of Iso-regulated OSCC were obtained. According to the degree obtained by topological analysis, PIK3R1, AKT1, SRC, ESR1, and other core targets were screened out. KEGG analysis showed that 165 signaling pathways were enriched in the intersection targets of Iso-OSCC, among which the PI3K/AKT signaling pathway played an important role in the treatment of OSCC with Iso. Molecular docking results showed that the absolute value of binding energy between target proteins PIK3R1, AKT1, SRC, ESR1, and Iso was high. After Cal-27 cells were treated with Iso, the number of cell colony formations, the number of transmembrane cells, and the expression of PIK3R1, ESR1, SRC, p-PI3K, and p-AKT were negatively correlated with the increase in Iso concentration (P < 0.05). Conclusion Iso can inhibit PI3K/AKT signal transduction and influence the expression of PIK3R1, AKT1, SRC, and ESR1 proteins, thereby inhibiting the occurrence and development of OSCC.

Depression is a psychiatric disorder whose main symptoms include low mood， loss of interest， anxiety， sleep disturbances， and changes in appetite. Orexin， a neuropeptide located in hypothalamic neurons， has a wide range of projections throughout the central nervous system and is involved in various behavioral modulations related to depression. This study systematically reviewed the effects of orexin and its receptor-related drugs on depression and found that orexin could exert complex regulatory effects on multiple brain regions by binding to related receptors， affecting emotions， sleep， anxiety， etc. The abnormal state of expression of plasma orexin in patients with depression was found. Exogenous orexin-A， selective orexin receptor 1 antagonists （SORA1s）， selective orexin receptor 2 antagonists （SORA2s）， and dual orexin receptor antagonists （DORAs） have demonstrated antidepressant-like effects in various animal models of depression. Among them， clinical trials involving exogenous orexin-A are relatively scarce. Drugs related to SORA1s and SORA2s， such as JNJ-61393215 and Setorexant， have made significant progress in the treatment of depression. DORAs， such as Suvorexant， Lemborexant， and Daridorexant， are primarily used to treat insomnia. Notably， Suvorexant has also shown potential in alleviating symptoms of anxiety and depression.

AIM:To analyze differential proteins associated with the pathogenesis of dry eye(DE)using bioinformatics methods, in order to reveal their potential molecular mechanisms.METHODS: Articles published in PubMed and EMBASE databases from the inception of the database to August 31, 2023, that used proteomic methods to detect protein expression in clinical samples of dry eye were searched. Differential proteins were selected and further analyzed using the STRING database and Cytoscape software for hub gene screening and module analysis. Protein-protein interaction(PPI)analysis, gene ontology(GO)functional annotation, and Kyoto encyclopedia of genes and genomes(KEGG)pathway enrichment analysis were performed.RESULTS: A total of 21 articles were included, identifying 74 differentially expressed proteins. The most frequently occurring differential proteins were calgranulin A(SA1008), lipocalin-1(LCN1), lysozyme C(LYZ), mammaglobin-B(SCGB2A1), proline-rich protein 4(PRR4), transferrin(TF), and calgranulinB(S100A9). The top 10 hub genes were serum albumin(ALB), tumor necrosis factor(TNF), interleukin 6(IL6), IL1B, IL8, matrix metalloproteinase 9(MMP9), alpha-1-antitrypsin(SERPINA1), IL10, complement component 3(C3), and lactotransferrin(LTF). Module analysis suggested MMP9 and PRR4 as seed genes. KEGG analysis showed that differential proteins were mainly enriched in the IL17 signaling pathway(61.9%).CONCLUSION: The results reveal potential molecular targets and pathways for DE and confirm the association between the pathogenesis of DE and inflammation. Further in-depth research is needed to confirm the significance of these biomarkers in clinical practice.