Objective:To observe any ability of pulsed electromagnetic field (PEMF) stimulation to regulate inflammation in degenerate nucleus pulposus cells.Methods:Primary nucleus pulposus cells from rats were cultured and divided into a control group, a model group, an A2AR-small interfering RNA group (A2AR-siRNA group), and a PEMF group. The control and model group cells were stimulated with tumor necrosis factor-α (TNF-α) in phosphate-buffered saline solution, those in the A2AR-siRNA and the PEMF groups were transfected with A2AR siRNA and given TNF-α stimulation. The PEMF group cells had four hours daily of PEMF irradiation beginning within 24 hours after the TNF-α stimulation for 2 days. After 48 hours, cell proliferation was detected by CCK8 assay, while the positive expression of A2AR in the nucleus pulposus cells was measured using immunofluorescence staining. The expression of A2AR, cyclic adenosine monophosphate (cAMP), protein kinase A (PKA), cAMP response element binding protein (CREB), nuclear factor-κB (NF-κB), nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), and interleukin-6 (IL-6) in the cells was observed using western blotting and real-time fluorescence quantitative polymerase chain reactions.Results:The rate of cell proliferation among the model group was approximately 60%, significantly lower than that of the control group (100%), but significantly higher than that of the A2AR-siRNA group (approximately 34%). The cell proliferation rate of the PEMF group was approximately 80%, significantly higher than that of the model and A2AR-siRNA groups. Compared with the control group, the positive expression of A2AR in the nucleus pulposus cells, A2AR protein and mRNA in the model group increased significantly in response to stress. The expression of A2AR protein and mRNA in the A2AR-siRNA group and the PEMF group was significantly lower than in the model group. Compared with the control group, the cAMP level in the nucleus pulposus cells of the model group had decreased significantly. The cAMP content in the A2AR-siRNA group further decreased compared with the model group, and that in the PEMF group increased compared with the model and the A2AR-siRNA groups. The average expression of PKA and CREB in the model group was significantly higher than among the control group, while that of PKA and CREB in the A2AR-siRNA group was significantly lower. The PKA and CREB protein levels in the PEMF group were slightly higher than in the A2AR-siRNA group, but the difference was not statistically significant. The expression of NF-κB, NLRP3, and IL-6 protein and mRNA in the cells of the model group was significantly higher than in the control group, but those of the A2AR-siRNA group were significantly higher than in the model group. The levels in the PEMF group were significantly lower than among the model and A2AR-siRNA groups, on average.Conclusions:Down-regulation of A2AR content further aggravates the inflammatory injury of degenerated nucleus pulposus cells. The mechanism may be related to the down-regulation of CREB activity, activation of the NF-κB signaling pathway, and subsequent up-regulation of inflammatory factors NLRP3 and IL-6. PEMF stimulation cannot significantly increase the A2AR level in degenerated nucleus pulposus cells, but it can promote the expression of cAMP and inhibit the downstream NF-κB signaling pathway, thereby exerting an anti-inflammatory effect.

Objective To investigate the predictive value of serum Nod-like receptor protein 3(NLRP3),calcitonin gene-related peptide(CGRP),and endothelial nitric oxide synthase(eNOS)for secondary intracranial infection and prognosis following craniotomy for cerebral hemorrhage.Meth-ods A case-control study was conducted.A total of 70 patients with secondary intracranial infection after craniotomy for cerebral hemorrhage in the hospital from January 2021 to May 2024 were selected as observation group,and 210 patients without secondary intracranial infection after craniotomy for cer-ebral hemorrhage were selected as control group.Surgical conditions and serum levels of NLRP3,CGRP,and eNOS were compared between the two groups,and clinical differences between died and surviving patients in the observation group were analyzed.Results The observation group had signifi-cantly higher rates of tracheal intubation and surgical duration exceeding 4 hours,along with significant elevated serum levels of NLRP3,CGRP,and eNOS compared to the control group(P＜0.001).Logistic regression analysis revealed that tracheal intubation,NLRP3,CGRP,and eNOS were influ-encing factors for secondary intracranial infection(P＜0.05).In the observation group,there were 18 deaths and 52 survivors.The death group had significantly higher age,the Acute Physiology and Chronic Health Evaluation Ⅱ(APACHE Ⅱ)score,and serum levels of NLRP3,CGRP,and eNOS compared to the surviving group(P＜0.05).Logistic regression analysis indicated that APACHE Ⅱ score and serum levels of NLRP3,CGRP,and eNOS were influencing factors for mor-tality in patients with secondary intracranial infection(P＜0.05).The areas under the curve of the receiver operating characteristic(ROC)curves for serum NLRP3,CGRP,and eNOS in predicting secondary intracranial infection were 0.805,0.784,and 0.735 respectively,with significant differ-ences(P＜0.001).The areas under the curve of ROC curves for serum NLRP3,CGRP,and eNOS in predicting mortality in patients with secondary intracranial infection were 0.684,0.763,and 0.763,respectively,with significant differences(P＜0.05).Conclusion Serum NLRP3,CGRP,and eNOS are influencing factors for secondary intracranial infection and poor prognosis fol-lowing craniotomy for cerebral hemorrhage,exhibiting certain predictive values.

Objective:To observe any ability of pulsed electromagnetic field (PEMF) stimulation to regulate inflammation in degenerate nucleus pulposus cells.Methods:Primary nucleus pulposus cells from rats were cultured and divided into a control group, a model group, an A2AR-small interfering RNA group (A2AR-siRNA group), and a PEMF group. The control and model group cells were stimulated with tumor necrosis factor-α (TNF-α) in phosphate-buffered saline solution, those in the A2AR-siRNA and the PEMF groups were transfected with A2AR siRNA and given TNF-α stimulation. The PEMF group cells had four hours daily of PEMF irradiation beginning within 24 hours after the TNF-α stimulation for 2 days. After 48 hours, cell proliferation was detected by CCK8 assay, while the positive expression of A2AR in the nucleus pulposus cells was measured using immunofluorescence staining. The expression of A2AR, cyclic adenosine monophosphate (cAMP), protein kinase A (PKA), cAMP response element binding protein (CREB), nuclear factor-κB (NF-κB), nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), and interleukin-6 (IL-6) in the cells was observed using western blotting and real-time fluorescence quantitative polymerase chain reactions.Results:The rate of cell proliferation among the model group was approximately 60%, significantly lower than that of the control group (100%), but significantly higher than that of the A2AR-siRNA group (approximately 34%). The cell proliferation rate of the PEMF group was approximately 80%, significantly higher than that of the model and A2AR-siRNA groups. Compared with the control group, the positive expression of A2AR in the nucleus pulposus cells, A2AR protein and mRNA in the model group increased significantly in response to stress. The expression of A2AR protein and mRNA in the A2AR-siRNA group and the PEMF group was significantly lower than in the model group. Compared with the control group, the cAMP level in the nucleus pulposus cells of the model group had decreased significantly. The cAMP content in the A2AR-siRNA group further decreased compared with the model group, and that in the PEMF group increased compared with the model and the A2AR-siRNA groups. The average expression of PKA and CREB in the model group was significantly higher than among the control group, while that of PKA and CREB in the A2AR-siRNA group was significantly lower. The PKA and CREB protein levels in the PEMF group were slightly higher than in the A2AR-siRNA group, but the difference was not statistically significant. The expression of NF-κB, NLRP3, and IL-6 protein and mRNA in the cells of the model group was significantly higher than in the control group, but those of the A2AR-siRNA group were significantly higher than in the model group. The levels in the PEMF group were significantly lower than among the model and A2AR-siRNA groups, on average.Conclusions:Down-regulation of A2AR content further aggravates the inflammatory injury of degenerated nucleus pulposus cells. The mechanism may be related to the down-regulation of CREB activity, activation of the NF-κB signaling pathway, and subsequent up-regulation of inflammatory factors NLRP3 and IL-6. PEMF stimulation cannot significantly increase the A2AR level in degenerated nucleus pulposus cells, but it can promote the expression of cAMP and inhibit the downstream NF-κB signaling pathway, thereby exerting an anti-inflammatory effect.

Objective To investigate the application value of MRI three-dimensional reconstruction techniques in the identification of offending vessels and affected nerves in cranial neurovascular compression syndrome.Methods A total of 91 patients with trigeminal neuralgia and 72 patients with hemifacial spasm underwent FIESTA and TOF-MRA scanning before microvascular decompression.Two-dimensional and three-dimensional reconstruction techniques(curved planar reconstruction and MR virtual endoscopy)were respectively performed using the original MRI data so as to evaluate the number and category of offending vessels and the degree of compression on cranial nerves.The results of imaging diagnosis were compared with the observations during neurosurgery.Results The correct identification rate of the offending vessels in trigeminal neuralgia was 95.6％by 3D reconstruction technique,with 94.5％diagnostic accuracy of neurovascular compression.In addition,the accurate identification rate was 91.7％on the detection of the offending vessels in hemifacial spasm,with 89.5％accuracy of the judgment on neurovascular compression.It should be noted that the diagnostic accuracy for the simple contact between the offending blood vessels and the cranial nerves using 3D reconstruction was significantly higher than that of the evaluation through 2D observation(P＜0.05).Conclusion 3D reconstruction technique can improve the accuracy of offending blood vessels identification and provide more reliable information on the degree of cranial nerve distortion or atrophy.Therefore,3D MRI images are more suitable for investigating the complex neurovascular compression.

Objective:To screen human monoclonal antibodies (mAbs) against the nuclear protein (NP) of severe fever with thrombocytopenia syndrome virus (SFTSV), identify their binding specificity to both recombinant NP and NP in viral particles, and determine their affinity constant and binding kinetics.Methods:Antibody genes were extracted from the blood of recovered individuals, and an antibody library was created using phage display. This library was panned by recombinant NP. The selected antibodies were expressed and purified. Enzyme linked immunosorbent assay (ELISA), western blot (WB), and indirect immunofluorescence assay (IFA) were used to assess the binding specificity of these mAbs to recombinant NP and NP in virions. Additionally, biolayer interferometry (BLI) was utilized to determine the antibody affinity constant.Results:An antibody library with a capacity of 7.24×10 7 was successfully constructed. Following three rounds of panning, 6 mAbs (named as NP-1, NP-10, NP-11, NP-20, NP-21, and NP-27) were isolated. The binding specificity of these 6 mAbs against recombinant NP was confirmed through indirect ELISA and WB analysis. Additionally, these mAbs were demonstrated specific in binding to NP in virions as evidenced by IFA detection. The affinity constant values of the 6 mAbs, determined by BLI assay, ranged from 0.47 nmol/L to 32 nmol/L. Conclusions:The 6 mAbs derived from the library are human mAbs that exhibit specificity to the NP of SFTSV and demonstrate a high affinity. These antibodies represent potential candidates for fundamental research and development of diagnostic reagents for SFTSV.

Objective:To document any effect of a pulsed electromagnetic field (PEMF) on the regulation of neuropeptide Y (NPY) in nucleus pulposus (NP) tissue, NP cell apoptosis and matrix degradation using rats with intervertebral disc degeneration (IDD).Methods:Eighteen Sprague-Dawley rats were randomly divided into a control group, an IDD model group (the model group), and a PEMF group. IDD was induced in both the model and PEMF groups. Right after the modeling, the PEMF group received 14 days of PEMF treatment, while the control group and model group were given no special treatment. Meanwhile, the primary rat nucleus pulposus cells (NPCs) were cultured using Dulbecco′s Modified Eagle Medium at 37℃ and 5% CO 2. When the fusion rate reached 90% after passage, the NPCs were divided into a control group, a TNF-α model group (referred to as model group) and TNF-α + PEMF group (referred to as PEMF group) and treated accordingly. Eight weeks after the modeling, safranin-o/fast green staining was used to assess any pathological morphology changes. The expression of NPY, neuropeptide Y receptor Y2 (NPY2R), bcl-2-associated X protein (Bax), B-cell lymphoma 2 (Bcl-2), collagen type II (Col-II) and matrix metalloproteinase-3 (MMP3) in the intervertebral disc and the cultivated nucleus pulposus cells of the 3 groups were determined. Results:The intervertebral disc cells in the model group were ruptured and folded, with significantly increased polysaccharide and protein components, and significantly increased bone fibers. In the PEMF group the cell boundaries were clearer, with less fibrin fracture and increased cartilage tissue. NPY was expressed in the fibrous annulus and the nucleus pulposus of the intervertebral disc in the model group. The average expression levels of NPY and NPY2R were significantly higher than in the control group and the model group. Compared with the control group, there was a significant increase in the level of Bax and a significant decrease in the expression of Bcl-2 in the model group, and there was a significant decrease in the level of Bax in the PEMF group. Compared with the control group, there was a significant decrease in the Col-II level but a significant increase in the MMP3 protein expression in the model group. The average Col-II mRNA expression was significantly higher in the PEMF group compared with the model group, but the average MMP3 protein expression was significantly less. Those results are consistent with observations in vivo.Conclusion:PEMF may reverse the imbalance of ECM metabolism and delay IDD degeneration by up-regulating the expression of NPY and Bcl-2, as well as blocking the Bax/Bcl-2 signaling pathway to inhibit apoptosis of nucleus pulposus cells.

Objective:To screen human monoclonal antibodies (mAbs) against the nuclear protein (NP) of severe fever with thrombocytopenia syndrome virus (SFTSV), identify their binding specificity to both recombinant NP and NP in viral particles, and determine their affinity constant and binding kinetics.Methods:Antibody genes were extracted from the blood of recovered individuals, and an antibody library was created using phage display. This library was panned by recombinant NP. The selected antibodies were expressed and purified. Enzyme linked immunosorbent assay (ELISA), western blot (WB), and indirect immunofluorescence assay (IFA) were used to assess the binding specificity of these mAbs to recombinant NP and NP in virions. Additionally, biolayer interferometry (BLI) was utilized to determine the antibody affinity constant.Results:An antibody library with a capacity of 7.24×10 7 was successfully constructed. Following three rounds of panning, 6 mAbs (named as NP-1, NP-10, NP-11, NP-20, NP-21, and NP-27) were isolated. The binding specificity of these 6 mAbs against recombinant NP was confirmed through indirect ELISA and WB analysis. Additionally, these mAbs were demonstrated specific in binding to NP in virions as evidenced by IFA detection. The affinity constant values of the 6 mAbs, determined by BLI assay, ranged from 0.47 nmol/L to 32 nmol/L. Conclusions:The 6 mAbs derived from the library are human mAbs that exhibit specificity to the NP of SFTSV and demonstrate a high affinity. These antibodies represent potential candidates for fundamental research and development of diagnostic reagents for SFTSV.

Objective:To explore any effect of pulsed electromagnetic field (PEMF) stimulation on intervertebral disc degeneration (IDD).Methods:Forty Sprague-Dawley rats were randomly divided into a control group, an IDD model group, a PEMF group and an observation group. An IDD model was induced in all except those in the control group. Both the PEMF and observation groups were given PEMF stimulation, while the latter was additionally injected with the A2AR agonist CGS-21680. Eight weeks after the modelling any pathological changes in the morphology of the rats′ intervertebral disc tissues were evaluated using saffron solid green staining. The expression of A2AR, cyclic adenosine phosphate (cAMP), protein kinase A (PKA), cysteine aspartate proteolytic enzyme-3 (Caspase-3), type II collagen (COL-II) and matrix metalloproteinase-3 (MMP3) in the intervertebral discs were evaluated.Results:The nucleus pulposus had shrunk, while fibrous tissues and chondrocytes had increased in the IDD model group. In the observation group the nucleus pulposus was intact and of basically normal shape. A2AR mRNA and protein levels were higher in the intervertebral disc tissue of the model group than among the control group, on average, while the levels in the observation group were significantly higher than in the other groups. In the PEMF and observation groups cAMP and PKA mRNA were significantly higher than in the IDD model group. The p38 MAPK and P-P38 MAPK levels of the IDD model group and its average P-P38 MAPK/p38 MAPK ratio were significantly higher than in the control group. In the PEMF and observation groups those indices had decreased to varying degrees, with those of the observation group significantly lower than among the model and PEMF groups on average, except for the p38 MAPK values. Caspase-3 and its mRNA were significantly higher in the model group than in the control group, on average, and those values were significantly lower in the PEMF and observation groups than in the IDD model group. The average MMP3 contents of the IDD model group had increased significantly compared with the control group, while the Col-Ⅱ level had decreased significantly. Compared with the IDD model group, the MMP3 level had decreased but Col-Ⅱ expression had increased in both the PEMF and observation groups, with significant differences between the IDD model and observation groups.Conclusions:The activation of the p38 MAPK signaling pathway by inflammatory factors to induce apoptosis is one of the important reasons for the aggravation of IDD lesions. PEMF combined with A2AR agonists can activate the A2AR/cAMP/PKA signaling pathway, inhibit p38 MAPK phosphorylation, reduce apoptosis of nucleus pulposus cells, and relieve IDD damage.

【Objective】 To explore the expressions of adipocyte enhancer binding protein 1 （AEBP1） gene and its isoforms in different types of gliomas， and the influence of AEBP1 gene on the prognosis of patients with different types of gliomas. 【Methods】 We used the GEPIA2 visual network analysis tool to analyze AEBP1 gene expression levels in the tumor tissues of glioblastomas （GBM， including classical， mesenchymal， neural， and proneural ones） and low-grade gliomas （LGG， including astrocytoma， oligoastrocytoma， oligodendroglioma） in the TCGA database and normal human tissue samples in the TCGA and GTEx databases by one-way ANOVA. The distribution trend of isoforms of AEBP1 gene in gliomas was analyzed using the violin plot. The Kaplan-Meier survival curve was drawn and the Logrank test was used to analyze the influence of AEBP1 gene expression in GBM and LGG tumor tissues on the prognosis of glioma patients. 【Results】 The expression of AEBP1 in the tumor tissues of overall GBM and the four types of GBM was higher than that in the normal control tissues （P<0.05）. The expression of AEBP1 in astrocytoma and oligodendrocyte astrocytoma tumor tissues was higher than that in normal control tissues （P<0.05）. There were nine isoforms of AEBP1 gene in GBM and LGG， and the expression level in GBM was higher. The overall survival （OS） of the AEBP1 low expression group of GBM patients and the proneuronal GBM patients was better than that of the high expression group （P<0.05）. The OS and progression-free survival of LGG patients and the AEBP1 low-expression group of astroglioma were better than those of the high-expression group （P<0.05）. 【Conclusion】 AEBP1 has an important clinical value in the pathogenesis and development of GBM and LGG， and thus can be used as a diagnostic marker and a candidate gene for targeted therapy.

【Objective】 To investigate the expression and biological role of miR-139-5p in glioblastoma and the regulatory effect of miR-139-5p on DNA methyltransferase 1 (DNMT1). 【Methods】 qRT-PCR was used to detect the expression of miR-139-5p in glioblastoma tumor tissue, paired paracancerous tissue, human normal glioma cell line HEB, and human glioma cell line U251. The expression of miR-139-5p in U251 cells was up-regulated by transfection of miR-139-5p mimetics, and the expression of DNMT1 was down-regulated by transfection of DNMT1-targeted siRNA (DNMT1-siRNA). The expression of DNMT1 and neurofibromatosis type 2 (NF2) in tissues and cells was detected by qRT-PCR, Western blotting, immunohistochemistry and immunofluorescence. The cell counting kit-8 (CCK-8), flow cytometry and Matrigel Transwell were used to evaluate the proliferation, apoptosis and invasion ability of U251 cells. 【Results】 Compared with paracancerous tissues or HEB cells, miR-139-5p expression in glioblastoma tissues and U251 cells was suppressed (P<0.05). Compared with control cells, transfection of miR-139-5p mimic significantly down-regulated the expression of DNMT1 and up-regulated the expression of NF2 (P<0.05). Compared with control cells, transfection of DNMT1-siRNA significantly promoted the expression of NF2 (P<0.05). Transfection of miR-139-5p mimetics or DNMT1-siRNA significantly induced U251 cell apoptosis and inhibited cell invasion (P<0.05). 【Conclusion】 miR-139-5p plays an anti-cancer role in glioblastoma, and it inhibits tumor proliferation and metastasis by targeting negative regulation of DNMT1.