BACKGROUND:The mechanism of programmed death receptor-1(PD-1)effect on osteogenic differentiation of bone marrow mesenchymal stem cells in high glucose environment remains unclear. OBJECTIVE:To explore the effect of PD-1 on osteogenic differentiation of rat bone marrow mesenchymal stem cells in high glucose environment and its regulatory mechanism. METHODS:Rat bone marrow mesenchymal stem cells were randomly divided into normal glucose group(5.6 mmol/L),high glucose group(30 mmol/L),PD-1 overexpression group,PD-1 overexpression no-load group,PD-1 knockdown group,PD-1 knockdown no-load group,and PI3K/AKT pathway inhibitor group(PD-1 knockdown+5 μmol/L LY294002).Rat bone marrow mesenchymal stem cells were cultured in high glucose to simulate the diabetic environment in vitro.The mRNA expression of PD-1 and ligand PD-L1 and the mRNA expression of osteogenic markers Runx2 and OSX in rat bone marrow mesenchymal stem cells were detected by qRT-PCR.The osteogenic differentiation ability was observed by alkaline phosphatase staining and alizarin red staining.Cell proliferation was detected by CCK-8 assay.The protein expressions of PD-1,PD-L1,p-PI3K,and p-AKT were detected by western blot assay. RESULTS AND CONCLUSION:(1)The levels of PD-1 and PD-L1 were significantly increased in the high glucose environment in vitro,and the osteogenic differentiation ability of bone marrow mesenchymal stem cells was inhibited in the high glucose environment.(2)Knockdown of PD-1 expression could promote osteogenic differentiation of bone marrow mesenchymal stem cells,increase cell proliferation activity,and activate the PI3K/AKT pathway.(3)After addition of PI3K/AKT pathway inhibitor LY294002,the ability of bone marrow mesenchymal stem cells to differentiate into osteoblasts decreased.The results show that PD-1 is dependent on the PI3K/AKT signaling pathway to inhibit osteogenic differentiation of rat bone marrow mesenchymal stem cells under high glucose environment.

BACKGROUND:Programmed cell death protein 1 belongs to the immunoglobulin gene superfamily and can regulate the differentiation of osteoblasts and affect bone homeostasis.However,there are few studies on the regulatory role and mechanism of programmed cell death protein 1 in diabetic osteoporosis.OBJECTIVE:To investigate the regulatory role and mechanism of programmed cell death protein 1 on osteogenic differentiation of rat bone marrow mesenchymal stem cells under high-glucose environment.METHODS:(1)Animal experiment:A total of 12 Sprageu-Dawley rats were randomized into a control group(n=6)and a model group(n=6).The control group was fed routinely,whereas the model group was injected intraperitoneally with streptozotocin to establish a model of type 1 diabetes mellitus,and the high-fat feed was fed for 8 weeks to establish a model of type 1 diabetic osteoporosis.After 8 weeks of feeding,the femurs of rats in the two groups were taken and subjected to hematoxylin-eosin staining and micro-CT assay.The mRNA expression of programmed cell death protein 1 and programmed death ligand 1 was detected.(2)Cell experiment:Passage 3 rat bone marrow mesenchymal stem cells were randomly divided into four groups:normal control group,high-glucose model group cultured in low glucose medium,programmed cell death protein 1-silenced group transfected with programmed cell death protein 1 siRNA,and programmed cell death protein 1-silenced null group transfected with siRNA-NC.After 48 hours of transfection,the normal control group was cultured in a new low-glucose medium,and the other three groups were cultured in a high-glucose medium for another 48 hours of culture followed by osteogenic induction.After 21 days of osteogenic induction,alizarin red staining,and qRT-PCR(programmed cell death protein 1 and RUNX2 mRNA expression)and western blot(β-catenin,GSK-3β,p-GSK-3β and Axin2 protein expression)were performed.RESULTS AND CONCLUSION:In the animal experiment,hematoxylin-eosin staining and micro-CT assay showed successful modeling of type 1 diabetic osteoporosis in the model group.qRT-PCR assay showed that the mRNA expression of programmed cell death protein 1 and programmed cell death ligand 1 was higher in the model group than the control group(P<0.05).In the cell experiment,the results of alizarin red staining showed that the ability of mineralized nodule formation was lower in the high-glucose model group and the programmed cell death protein 1-silenced null group than in the control group and the programmed cell death protein 1-silenced group.Compared with the normal control group,the programmed cell death protein 1 mRNA expression and GSK3β and Axin2 protein expression were elevated in the high-glucose model group and the programmed cell death protein 1-silenced null group(P<0.05),and the RUNX2 mRNA expression and p-GSK3β and β-catenin protein expression were decreased(P<0.05).Compared with the high-glucose model group and the programmed cell death protein 1-silenced null group,programmed cell death protein 1 mRNA expression and GSK3β and Axin2 protein expression were decreased in the programmed cell death protein 1-silenced group(P<0.05),and RUNX2 mRNA expression and p-GSK3β and β-catenin protein expression were elevated(P<0.05).To conclude,programmed cell death protein 1 silencing can activate the Wnt/β-catenin and improve the osteogenic differentiation of rat bone marrow mesenchymal stem cells under high-glucose conditions.

Objective To identify the potential pathological mechanisms of tuberculous spondylitis(TS).Methods Spinal specimens were collected from 13 TS patients and 13 controls who received treatment at a hospital from March 2021 to March 2023.Specimens were randomly selected from 3 TS patients and 3 controls to perform high-throughput lncRNAs and mRNAs sequencing with Illumina NovaSeq 6000.Differentially expressed lncRNAs(DELncRs)and mRNAs(DEmRs)in TS specimens were identified and analyzed through differential expression,and enrichment analysis was performed.The top 20 DEmRs with high connectivity were identified through protein-protein interaction(PPI)network.Regulatory network of DElncRs and DEmRs was built.Finally,gene expression of the remaining specimens was analyzed using qRT-PCR detection.Results A total of 1 243 DEmRs and 262 DElncRs were identified.Enrichment analysis revealed that muscle contraction,muscle system processes,muscle structural development,PI3K Akt signaling pathway,calcium signaling pathway,and cAMP signaling pathway were activated in TS,while responses to cytokines,cytokine-mediated signaling pathways,regulation of immune system processes,cytokine-cytokine receptor interactions,human T-cell leukemia virus type 1 infection,and phago-somes were inhibited in TS.Three sub-networks were identified in PPI,among which MYL1,TTN,LOC102723407,HLA-A,interleukin(IL)-6,and IL-1β had the highest connectivity and were identified as key DEmRs.MYL1,TTN,and IL-6 were regulated by DElncRs.qRT-PCR validated the differential expression of key DEmRs in TS.Conclusion DEmRs are regulated by lncRNAs and participate in the pathological process of TS,and the immune responses are inhibited in diseases condition.This study reveals key molecules and signaling pathways in TS,providing new insights into the pathological mechanisms of TS,and suggest scientific basis for developing new therapeutic targets.

Objective To identify the potential pathological mechanisms of tuberculous spondylitis(TS).Methods Spinal specimens were collected from 13 TS patients and 13 controls who received treatment at a hospital from March 2021 to March 2023.Specimens were randomly selected from 3 TS patients and 3 controls to perform high-throughput lncRNAs and mRNAs sequencing with Illumina NovaSeq 6000.Differentially expressed lncRNAs(DELncRs)and mRNAs(DEmRs)in TS specimens were identified and analyzed through differential expression,and enrichment analysis was performed.The top 20 DEmRs with high connectivity were identified through protein-protein interaction(PPI)network.Regulatory network of DElncRs and DEmRs was built.Finally,gene expression of the remaining specimens was analyzed using qRT-PCR detection.Results A total of 1 243 DEmRs and 262 DElncRs were identified.Enrichment analysis revealed that muscle contraction,muscle system processes,muscle structural development,PI3K Akt signaling pathway,calcium signaling pathway,and cAMP signaling pathway were activated in TS,while responses to cytokines,cytokine-mediated signaling pathways,regulation of immune system processes,cytokine-cytokine receptor interactions,human T-cell leukemia virus type 1 infection,and phago-somes were inhibited in TS.Three sub-networks were identified in PPI,among which MYL1,TTN,LOC102723407,HLA-A,interleukin(IL)-6,and IL-1β had the highest connectivity and were identified as key DEmRs.MYL1,TTN,and IL-6 were regulated by DElncRs.qRT-PCR validated the differential expression of key DEmRs in TS.Conclusion DEmRs are regulated by lncRNAs and participate in the pathological process of TS,and the immune responses are inhibited in diseases condition.This study reveals key molecules and signaling pathways in TS,providing new insights into the pathological mechanisms of TS,and suggest scientific basis for developing new therapeutic targets.

BACKGROUND:Programmed cell death protein 1 belongs to the immunoglobulin gene superfamily and can regulate the differentiation of osteoblasts and affect bone homeostasis.However,there are few studies on the regulatory role and mechanism of programmed cell death protein 1 in diabetic osteoporosis.OBJECTIVE:To investigate the regulatory role and mechanism of programmed cell death protein 1 on osteogenic differentiation of rat bone marrow mesenchymal stem cells under high-glucose environment.METHODS:(1)Animal experiment:A total of 12 Sprageu-Dawley rats were randomized into a control group(n=6)and a model group(n=6).The control group was fed routinely,whereas the model group was injected intraperitoneally with streptozotocin to establish a model of type 1 diabetes mellitus,and the high-fat feed was fed for 8 weeks to establish a model of type 1 diabetic osteoporosis.After 8 weeks of feeding,the femurs of rats in the two groups were taken and subjected to hematoxylin-eosin staining and micro-CT assay.The mRNA expression of programmed cell death protein 1 and programmed death ligand 1 was detected.(2)Cell experiment:Passage 3 rat bone marrow mesenchymal stem cells were randomly divided into four groups:normal control group,high-glucose model group cultured in low glucose medium,programmed cell death protein 1-silenced group transfected with programmed cell death protein 1 siRNA,and programmed cell death protein 1-silenced null group transfected with siRNA-NC.After 48 hours of transfection,the normal control group was cultured in a new low-glucose medium,and the other three groups were cultured in a high-glucose medium for another 48 hours of culture followed by osteogenic induction.After 21 days of osteogenic induction,alizarin red staining,and qRT-PCR(programmed cell death protein 1 and RUNX2 mRNA expression)and western blot(β-catenin,GSK-3β,p-GSK-3β and Axin2 protein expression)were performed.RESULTS AND CONCLUSION:In the animal experiment,hematoxylin-eosin staining and micro-CT assay showed successful modeling of type 1 diabetic osteoporosis in the model group.qRT-PCR assay showed that the mRNA expression of programmed cell death protein 1 and programmed cell death ligand 1 was higher in the model group than the control group(P<0.05).In the cell experiment,the results of alizarin red staining showed that the ability of mineralized nodule formation was lower in the high-glucose model group and the programmed cell death protein 1-silenced null group than in the control group and the programmed cell death protein 1-silenced group.Compared with the normal control group,the programmed cell death protein 1 mRNA expression and GSK3β and Axin2 protein expression were elevated in the high-glucose model group and the programmed cell death protein 1-silenced null group(P<0.05),and the RUNX2 mRNA expression and p-GSK3β and β-catenin protein expression were decreased(P<0.05).Compared with the high-glucose model group and the programmed cell death protein 1-silenced null group,programmed cell death protein 1 mRNA expression and GSK3β and Axin2 protein expression were decreased in the programmed cell death protein 1-silenced group(P<0.05),and RUNX2 mRNA expression and p-GSK3β and β-catenin protein expression were elevated(P<0.05).To conclude,programmed cell death protein 1 silencing can activate the Wnt/β-catenin and improve the osteogenic differentiation of rat bone marrow mesenchymal stem cells under high-glucose conditions.

[Objective]To compare the differences of neuromyelitis optica spectrum disorder(NMOSD)and multiple sclerosis (MS) on pregnancy ,and analyze the mutual impact of pregnancy on the diseases.[Methods]Prospectively collected clinical information of 235 NMOSD patients and 125 MS patients ,including the annualized relapse rate (ARR),the Expanded Disability Status Scale(EDSS)and pregnancy outcomes. 70 NMOSD patients and 30 MS patients were screened out as information patients. The ARR and EDSS score in two groups were compared during the year before pregnancy,during pregnancy and after 1 year postpartum, respectively. 50 cases of normal pregnant women for the same period as the control group ,then to compared the difference of three groups on pregnancy outcomes.[Results]Attacks occurring during pregnancy or one year after childbirth/abortion in NMOSD and MS were 53.25%(41/77)and 20.00%(7/35)(P=0.001);The ARR during the first 3 months postpartum periods of NMOSD and MS group(2.65,2.51)was significantly higher than during the year before pregnancy(0.27,0.49,P < 0.001)and during pregnancy (0.32,0.2,P<0.001);The EDSS score of two groups increased after 1 year postpartum(3.06 ± 2.16,2.19 ± 1.28)than that during the year before pregnancy(1.58 ± 0.48,1.92 ± 1.29,P < 0.001)and during pregnancy(1.92 ± 1.35,1.67 ± 0.70,P < 0.001). There was no difference on ARR and EDSS score between NMOSD and MS group. NMOSD ,MS and normal control group had no dif?ference on pregnancy outcomes and neonatal weight.[Conclusions]Compared with MS,the attack of NMOSD had more closer relation?ship with pregnancy;both NMOSD and MS would increase the risk of disease relapsing and disability after pregnancy;the diseases had no effect on pregnancy outcomes.

BACKGROUNDGreen tea has been shown to improve cholesterol metabolism in animal studies, but the molecular mechanisms underlying this function have not been fully understood. Long non-coding RNAs (lncRNAs) have recently emerged as a major class of regulatory molecules involved in a broad range of biological processes and complex diseases. Our aim was to identify important lncRNAs that might play an important role in contributing to the benefits of epigallocatechin-3-gallate (EGCG) on cholesterol metabolism.

METHODSMicroarrays was used to reveal the lncRNA and mRNA profiles in green tea polyphenol(-)-epigallocatechin gallate in cultured human liver (HepG2) hepatocytes treated with EGCG and bioinformatic analyses of the predicted target genes were performed to identify lncRNA-mRNA targeting relationships. RNA interference was used to investigate the role of lncRNAs in cholesterol metabolism.

RESULTSThe expression levels of 15 genes related to cholesterol metabolism and 285 lncRNAs were changed by EGCG treatment. Bioinformatic analysis found five matched lncRNA-mRNA pairs for five differentially expressed lncRNAs and four differentially expressed mRNA. In particular, the lncRNA AT102202 and its potential targets mRNA-3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR) were identified. Using a real-time polymerase chain reaction technique, we confirmed that EGCG down-regulated mRNA expression level of the HMGCR and up-regulated expression of AT102202. After AT102202 knockdown in HepG2, we observed that the level of HMGCR expression was significantly increased relative to the scrambled small interfering RNA control (P < 0.05).

CONCLUSIONSOur results indicated that EGCG improved cholesterol metabolism and meanwhile changed the lncRNAs expression profile in HepG2 cells. LncRNAs may play an important role in the cholesterol metabolism.

Catechin ; analogs & derivatives ; metabolism ; Cell Line, Tumor ; Cholesterol ; metabolism ; Hep G2 Cells ; Humans ; RNA, Long Noncoding ; genetics

BACKGROUND:Spine minimaly invasive technique through foraminal mirror is the method to treat lumbar disc herniation with minimal wound. This technique can be conducted under local anesthesia, and does not need to resect the smal joint or destroy the vertebral plate, and has smal damage to the spine. OBJECTIVE: To explore the short-period effects of transforaminal endoscopic spine system for adjacent-segment degenerative changes-caused low back pain after lumbar fixation and fusion. METHODS:A total of 31 patients with degenerative changes after posterior lumbar bone graft fusion fixation, who required secondary surgery, were enroled in this study, including 23 males and 8 females, at the age of 45-81 years old. The postoperative time was 1.1-5.7 years. There were 3 cases of L3-4 single segment, 15 cases of L4-5 single segment, 8 cases of L5S1 single segment, and 5 cases of multi-segment. These patients were treated with transforaminal endoscopic spine system, and folowed up for 6 months. Visual Analogue Scale score and lumbar function Japanese Orthopedic Association score were observed. RESULTS AND CONCLUSION: Lumbar and leg pain symptoms were relieved noticeably during the operation. The patient could walk immediately after the surgery, and the postoperative recovery was quite satisfactory. Visual Analogue Scale score was lower immediately, 1, 3 and 6 months after treatment compared with pre-treatment. Lumbar function Japanese Orthopedic Association score was higher immediately, 1, 3 and 6 months after treatment compared with pre-treatment. Results verify that transforaminal endoscopic spine system for degenerative changes after posterior lumbar bone graft fusion fixation has some advantages such as high safety, short operation time, less hemorrhage, less complications, rapid restoration and easily accepted by patients.

BACKGROUND:Ferula sinkiangensis K.M. Shen is composed of volatile oil, resin and gum that have the anti-inflammatory, anti-alergic, antispasmodic and analgesic effects. But its analgesic mechanism is unclear. OBJECTIVE: To observe the effect ofFerula sinkiangensis K.M. Shen on heat pain, mechanical pain, Fos protein expression and astrocyte activation in spinal cord of rats with neuropathic pain. METHODS: Eighty adult Sprague-Dawley rat models of chronic sciatic nerve injury were randomly divided into five groups and then intragasticaly administeredFerula sinkiangensis K.M. Shen at low, moderate and high doses (0.075, 0.15, 0.30 g/kg), celecoxib or physiological saline. Heat pain and mechanical pain were measured at 1 day before operation and at 1, 2, 3, 5, 7, 14 days after operation. The spinal cord tissue at S4-5 segments was harvested and Fos protein expression and astrocyte activation in the spinal cord of rats were observed by immunohistochemical staining method. RESULTS AND CONCLUSION: After 1 and 5 days of medication, behavioral pain scores of rats in the low-, moderate-, and high-doseFerula sinkiangensis K.M. Shen groups were significantly higher than that in the physiological saline group (P < 0.01). The largest reduction in heat pain threshold was measured in the moderate-doseFerula sinkiangensis K.M. Shen group compared to the other groups (P < 0.01). The most significant reduction in rat mechanical pain threshold was measured in the high-doseFerula sinkiangensis K.M. Shen group than in the other groups (P < 0.01). At each time point post-operation, the number of Fos protein-positive cels in the low-, moderate- and high-doseFerula sinkiangensis K.M. Shen and celecoxib groups was significantly lower than that in the physiological saline group (P < 0.05); the number of Fos protein-positive cels in the moderate- and high-doseFerula sinkiangensis K.M Shen groups was significantly higher than that in the celecoxib group (P< 0.05). At each time point post-operation, the number of astrocytes in the spinal cord tissue of rats in the high-doseFerula sinkiangensis K.M. Shen and celecoxib groups was significantly lower than that in the physiological saline group (P< 0.05). There was significant difference in the number of astrocytes between the moderate- and high-doseFerula sinkiangensis K.M shen groups and celecoxib group (P< 0.05). These results confirm thatFerula sinkiangensis K.M. Shen may effectively aleviate the neuropathic pain of rats, and the mechanism of which may be related to the activation of Fos protein and astrocytes in the spinal cord.

Airway Obstruction ; diagnosis ; therapy ; Child, Preschool ; Humans ; Infant ; Infant, Newborn