[摘 要] 目的：探究连翘苷（PHN）调节高迁移率族蛋白B1（HMGB1）/晚期糖基化终产物受体（RAGE）信号通路对胶质瘤U251细胞增殖、侵袭及上皮间质转化（EMT）的影响。方法：将人胶质瘤U251细胞分为PHN-0组（0 µmol/L PHN处理）、PHN低、中和高剂量组（PHN-50、PHN-100、PHN-200组，分别用50、100和200 µmol/L PHN处理）、PHN + pcDNA-NC组（转染pcDNA-NC质粒后200 µmol/L PHN处理）和PHN + HMGB1组（转染过表达HMGB1质粒后200 µmol/L PHN处理）。CCK-8法和克隆形成实验检测各组细胞的增殖能力，流式细胞术检测各组细胞的凋亡水平，Transwell实验检测各组细胞的迁移和侵袭能力，ELISA检测各组细胞分泌IL-8水平，免疫荧光法检测各组细胞中神经钙黏素（N-cadherin）和上皮钙黏素（E-cadherin）阳性率，WB法检测各组细胞中Toll样受体4（TLR4）、核因子-κB（NF-κB）、HMGB1、RAGE、N-cadherin、E-cadherin、细胞周期蛋白D1（cyclin D1）、细胞周期蛋白依赖性激酶2（CDK2）、B淋巴细胞瘤-2（Bcl-2）、Bcl-2相关X蛋白（BAX）蛋白的表达水平。结果：与PHN-0组相比，PHN-50、PHN-100、PHN-200组U251细胞增殖活力、克隆形成数、迁移和侵袭数、IL-8分泌水平、N-cadherin阳性率及其蛋白表达、TLR4、NF-κB、HMGB1、RAGE、cyclin D1、CDK2蛋白表达均显著降低（均P < 0.05），细胞凋亡率、E-cadherin阳性率及其蛋白表达、BAX/Bcl-2比值均显著升高（均P < 0.05）；同时过表达HMGB1则可逆转PHN对U251细胞增殖、迁移、侵袭及EMT的抑制作用和对凋亡的促进作用（均P < 0.05）。结论：PHN通过HMGB1/RAGE信号通路抑制胶质瘤U251细胞增殖、侵袭及EMT进程。

Parkinson's disease (PD) is a common neurodegenerative disorder mainly related to mitochondrial dysfunction of dopaminergic neurons in the midbrain substantia nigra. This study aimed to investigate the effects of exercise preconditioning on motor deficits and mitochondrial function in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mouse model. Eight-week-old male C57BL/6J mice were randomly divided into four groups: sedentary + saline (SS), sedentary + MPTP (SM), exercise + saline (ES), and exercise + MPTP (EM) groups. Mice in the ES and EM groups received 4 weeks of treadmill training, and then SM and EM groups were treated with MPTP for 5 days. Motor function was assessed by behavioral tests, and morphological and functional changes in dopaminergic neurons and mitochondria in the substantia nigra of the midbrain were evaluated using immunohistochemistry, Western blot, and transmission electron microscopy technology. The results showed that, compared with the SM group, the EM group exhibited significantly improved motor ability, up-regulated protein expression levels of tyrosine hydroxylase (TH) and dopamine transporter (DAT) in the midbrain, and down-regulated protein expression of α-synuclein (α-Syn) in the mitochondria of substantia nigra. Compared with the SM group, the EM group showed up-regulated protein expression levels of mitochondrial fusion proteins, including optical atrophy protein 1 (OPA1) and mitofusin 2 (MFN2), and biogenesis-related proteins, including peroxisome proliferator activated receptor gamma coactivator 1α (PGC-1α) and mitochondrial transcription factor A (TFAM), while the protein expression levels of dynamin-related protein 1 (DRP1) and mitochondrial fission protein 1 (FIS1) were significantly down-regulated. Compared with the SM group, the EM group showed significantly reduced damage to substantia nigra mitochondria, restored mitochondrial membrane potential and ATP production, and decreased levels of reactive oxygen species (ROS). These results suggest that 4-week treadmill pre-training can alleviate MPTP-induced motor impairments in PD mice by improving mitochondrial function, providing a theoretical basis for early exercise-based prevention of PD.
Animals ; Male ; Physical Conditioning, Animal/physiology* ; Mice ; Mice, Inbred C57BL ; Mitochondria/physiology* ; Dopaminergic Neurons ; MPTP Poisoning/physiopathology* ; Substantia Nigra ; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha ; 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine

This study investigated the effects and underlying mechanisms of vanillic acid(VA) against cardiac fibrosis(CF) induced by isoproterenol(ISO) in mice. Male C57BL/6J mice were randomly divided into control group, VA group(100 mg·kg~(-1), ig), ISO group(10 mg·kg~(-1), sc), ISO + VA group(10 mg·kg~(-1), sc + 100 mg·kg~(-1), ig), ISO + dynamin-related protein 1(Drp1) inhibitor(Mdivi-1) group(10 mg·kg~(-1), sc + 50 mg·kg~(-1), ip), and ISO + VA + Mdivi-1 group(10 mg·kg~(-1), sc + 100 mg·kg~(-1), ig + 50 mg·kg~(-1), ip). The treatment groups received the corresponding medications once daily for 14 consecutive days. On the day after the last administration, cardiac functions were evaluated, and serum and cardiac tissue samples were collected. These samples were analyzed for serum aspartate aminotransferase(AST), lactate dehydrogenase(LDH), creatine kinase-MB(CK-MB), cardiac troponin I(cTnI), reactive oxygen species(ROS), interleukin(IL)-1β, IL-4, IL-6, IL-10, IL-18, and tumor necrosis factor-α(TNF-α) levels, as well as cardiac tissue catalase(CAT), glutathione(GSH), malondialdehyde(MDA), myeloperoxidase(MPO), superoxide dismutase(SOD), total antioxidant capacity(T-AOC) activities, and cytochrome C levels in mitochondria and cytoplasm. Hematoxylin-eosin, Masson, uranium acetate and lead citrate staining were used to observe morphological and mitochondrial ultrastructural changes in the cardiac tissues, and myocardial injury area and collagen volume fraction were calculated. Flow cytometry was applied to detect the relative content and M1/M2 polarization of cardiac macrophages. The mRNA expression levels of macrophage polarization markers [CD86, CD206, arginase 1(Arg-1), inducible nitric oxide synthase(iNOS)], CF markers [type Ⅰ collagen(Coll Ⅰ), Coll Ⅲ, α-smooth muscle actin(α-SMA)], and cytokines(IL-1β, IL-4, IL-6, IL-10, IL-18, TNF-α) in cardiac tissues were determined by quantitative real-time PCR. Western blot was used to detect the protein expression levels of Coll Ⅰ, Coll Ⅲ, α-SMA, Drp1, p-Drp1, voltage-dependent anion channel(VDAC), hexokinase 1(HK1), NOD-like receptor protein 3(NLRP3), apoptosis-associated speck-like protein(ASC), caspase-1, cleaved-caspase-1, gasdermin D(GSDMD), cleaved N-terminal gasdermin D(GSDMD-N), IL-1β, IL-18, B-cell lymphoma-2(Bcl-2), B-cell lymphoma-xl(Bcl-xl), Bcl-2-associated death promoter(Bad), Bcl-2-associated X protein(Bax), apoptotic protease activating factor-1(Apaf-1), pro-caspase-3, cleaved-caspase-3, pro-caspase-9, cleaved-caspase-9, poly(ADP-ribose) polymerase-1(PARP-1), and cleaved-PARP-1 in cardiac tissues. The results showed that VA significantly improved cardiac function in mice with CF, reduced myocardial injury area and cardiac index, and decreased serum levels of AST, CK-MB, cTnI, LDH, ROS, IL-1β, IL-6, IL-18, and TNF-α. VA also lowered MDA and MPO levels, mRNA expressions of IL-1β, IL-6, IL-18, and TNF-α, and mRNA and protein expressions of Coll Ⅰ, Coll Ⅲ, and α-SMA in cardiac tissues, and increased serum levels of IL-4 and IL-10, cardiac tissue levels of CAT, GSH, SOD, and T-AOC, and mRNA expressions of IL-4 and IL-10. Additionally, VA ameliorated cardiac pathological damage, inhibited myocardial cell apoptosis, inflammatory infiltration, and collagen fiber deposition, reduced collagen volume fraction, and alleviated mitochondrial damage. VA decreased the ratio of F4/80~+CD86~+ M1 cells and the mRNA expressions of CD86 and iNOS in cardiac tissue, and increased the ratio of F4/80~+CD206~+ M2 cells and the mRNA expressions of CD206 and Arg-1. VA also reduced protein expressions of p-Drp1, VDAC, NLRP3, ASC, caspase-1, cleaved-caspase-1, GSDMD, GSDMD-N, IL-1β, IL-18, Bad, Bax, Apaf-1, cleaved-caspase-3, cleaved-caspase-9, cleaved-PARP-1, and cytoplasmic cytochrome C, and increased the expressions of HK1, Bcl-2, Bcl-xl, pro-caspase-3, pro-caspase-9 proteins, as well as the Bcl-2/Bax and Bcl-xl/Bad ratios and mitochondrial cytochrome C content. These results indicate that VA has a significant ameliorative effect on ISO-induced CF in mice, alleviates ISO-induced oxidative damage and inflammatory response, and its mechanism may be closely related to the inhibition of Drp1/HK1/NLRP3 and mitochondrial apoptosis signaling pathways, suppression of myocardial cell inflammatory infiltration and collagen fiber deposition, reduction of collagen volume fraction and CollⅠ, Coll Ⅲ, and α-SMA expressions, thus mitigating CF.
Animals ; Isoproterenol/adverse effects* ; Male ; Mice ; Signal Transduction/drug effects* ; Vanillic Acid/administration & dosage* ; Dynamins/genetics* ; Mice, Inbred C57BL ; Fibrosis/genetics* ; Apoptosis/drug effects* ; Mitochondria/metabolism* ; NLR Family, Pyrin Domain-Containing 3 Protein/genetics* ; Myocardium/metabolism* ; Humans

Objective To establish a dynamic prediction model of fatal massive hemorrhage in trauma based on the vital signs time series data and machine learning algorithms.Methods Retrospectively analyze the vital signs time series data of 7522 patients with trauma in the Medical Information Mart for Intensive Care-Ⅳ(MIMIC-Ⅳ)database from 2008 to 2019.According to the occurrence of posttraumatic fatal massive hemorrhage,the patients were divided into two groups:fatal massive hemorrhage group(n=283)and non-fatal massive hemorrhage group(n=7239).Six machine learning algorithms,including logistic regression(LR),support vector machine(SVM),random forests(RF),adaptive boosting(AdaBoost),gated recurrent unit(GRU),and GRU-D were used to develop a dynamic prediction models of fatal massive hemorrhage in trauma.The probability of fatal massive hemorrhage in the following 1,2,and 3 h was dynamically predicted.The performance of the models was evaluated by accuracy,sensitivity,specificity,positive predictive value,negative predictive value,Youden index,and area under receiver operating characteristic curve(AUC).The models were externally validated based on the trauma database of the Chinese PLA General Hospital.Results In the MIMIC-Ⅳ database,the set of dynamic prediction models based on the GRU-D algorithm was the best.The AUC for predicting fatal major bleeding in the next 1,2,and 3 h were 0.946±0.029,0.940±0.032,and 0.943±0.034,respectively,and there was no significant difference(P=0.905).In the trauma dataset,GRU-D model achieved the best external validation effect.The AUC for predicting fatal major bleeding in the next 1,2,and 3 h were 0.779±0.013,0.780±0.008,and 0.778±0.009,respectively,and there was no significant difference(P=0.181).This set of models was deployed in a public web calculator and hospital emergency department information system,which is convenient for the public and medical staff to use and validate the model.Conclusion A set of dynamic prediction models has been successfully developed and validated,which is greatly significant for the early diagnosis and dynamic prediction of fatal massive hemorrhage in trauma.

In this study, fluvoxamine maleate sustained-release pellet system tablets were prepared and were used to evaluate their release behaviors in vitro. Fluvoxamine maleate pellets were prepared using centrifugal-spherization method and coated by fluidized bed as bottom-spray. The multi-unit sustained-release pellets and appropriate excipients for prescription volumes were mixed uniformly and then compressed to tablets. Screening and determining the optimal formulation of drug loaded pellets through L8 (2⁴) Taguchi experiment. Using Minitab software to design a DOE experiment with 2⁴ partial factors, including material temperature, fan speed, atomization pressure, and spray rate to optimize the bottom spray coating process. Taking monostearate glycerol ester with a particle size of 24-40 mesh as the main diluent for tableting to relieve the delamination phenomenon between pellets and excipients during tablet pressing and reduce mechanical damage to the coating film. By examining the powder fluidity indexes such as angle of repose, bulk density, tapped density, and Hausner ratio of mixed particles, it was found that the flowability and compressibility are good and suitable for direct compression. Evaluate the basic properties of the sustained-release tablets, investigate the in vitro release behavior and study the release mechanism. The results of in vitro release test showed that the self-made sustained-release tablets could disintegrate into independent pellet units in phosphate buffer at pH 6.8 and release slowly within 24 h, which conformed to the first-order drug release model. The fluvoxamine maleate sustained-release pellet system tablets meet the requirements of preparation design and has a great commercial prospect.

Objective To investigate the value of magnetic resonance imaging(MRI)T2-mapping in evaluating the activity of Graves ophthalmopathy(GO).Methods A total of 64 patients with GO in the Department of Endocrinology,the First Affiliated Hospital of Chongqing Medical University from July 2019 to January 2021 were collected.Simple random grouping was performed by computer,with 49 cases as observation subjects,and 15 patients for diagnostic test.According to clinical activity score(CAS),49 GO patients were divided into active group(CAS≥3 points,48 eyes)and inactive group(CAS<3 points,50 eyes).Normal control group(NC group)included 31 patients(62 eyes).All subjects underwent 3.0T orbital MRI T2-mapping.Measuring the T2 relaxation time(T2RT)of superior rectus,inferior rectus,medial rectus,and lateral rectus on five layers behind the eyeball on T2-mapping coronal images,and select the maximum value of T2RT in the five layers for each extraocular muscle to represent the T2RT of this extraocular muscle.Finally,select the maximum T2RT values of the four extraocular muscles,expressed as extraocular muscle maximum T2RT.Compare the differences of the above 5 indicators(superior rectus T2RT,inferior rectus T2RT,medial rectus T2RT,lateral rectus T2RT,extraocular muscle maximum T2RT)between active group,inactive group and NC group.ROC curve was used to analyze the diagnostic value of the above 5 indicators for GO activity assessment,and the diagnostic threshold was obtained.Then,another 15 GO patients were performed for diagnostic tests evaluation to determine the indicators of high diagnostic efficacy and the threshold of diagnostic activity.Results The T2RT of all extraocular muscles in active group were significantly higher than those in inactive group and NC group,the difference was statistically significant(P<0.001).The threshold value of the five indicators were obtained by ROC curve analysis.The maximum T2RT cut-off values of superior rectus muscle,inferior rectus muscle,medial rectus muscle,lateral rectus muscle and extraocular muscles for judging activity were 80.200 ms,97.045 ms,94.355 ms,85.750 ms and 101.385 ms respectively.Another 15 GO patients were performed for diagnostic tests,the indexes with relatively high sensitivity,specificity,positive predictive value and negative predictive value were inferior rectus T2RT and extraocular muscle maximum T2RT,the cut-off values of GO activity were 97.045 ms and 101.385 ms,respectively;the sensitivity were 91.7％and 93.8％,respectively;the specificity all were 80.0％.Conclusions MRI T2-mapping sequence has a good value in assessment of GO activity.The inferior rectus T2RT and extraocular muscle maximum T2RT can be choosed to evaluate the activity of GO.

ObjectiveIn recent years, the negative impact of microgravity on astronauts’ nervous systems has received widespread attention. The repetitive transcranial magnetic stimulation (rTMS) technology has shown significant positive effects in the treatment of neurological and psychiatric disorders. The potential benefits of combined frequency stimulation (CFS) which combines different frequency stimulation patterns in ameliorating neurological dysfunctions induced by the microgravity environment, still require in-depth investigation. Exploring the therapeutic effects and electrophysiological mechanisms of CFS in improving various neurological disorders caused by microgravity holds significant importance for neuroscience and the clinical application of magnetic stimulation. MethodsThis study employed 40 C57BL/6 mice, randomly divided into 5 groups: sham group, hindlimb unloading (HU) group, 10 Hz group, 20 Hz group, and combined frequency stimulation (10 Hz+20 Hz, CFS) group. Mice in all groups except the sham group received 14 d of simulated microgravity conditions along with 14 d of repetitive transcranial magnetic stimulation. The effects of CFS on negative emotions and spatial cognitive abilities were assessed through sucrose preference tests and water maze experiments. Finally, patch-clamp techniques were used to record action potentials, resting membrane potentials, and ion channel dynamics of granule neurons in the hippocampal dentate gyrus (DG) region. ResultsCompared to the single-frequency stimulation group, behavioral results indicated that the combined frequency stimulation (10 Hz+20 Hz) significantly improved cognitive impairments and negative emotions in simulated microgravity mice. Electrophysiological experiments revealed a decrease in excitability of granule neurons in the hippocampal DG region after HU manipulation, whereas the combined frequency stimulation notably enhanced neuronal excitability and improved the dynamic characteristics of voltage-gated Na⁺ and K⁺ channels. ConclusionThe repetitive transcranial magnetic stimulation with combined frequencies (10 Hz+20 Hz) effectively ameliorates cognitive impairments and negative emotions in simulated microgravity mice. This improvement is likely attributed to the influence of combined frequency stimulation on neuronal excitability and the dynamic characteristics of Na⁺ and K⁺ channels. Consequently, this study holds the promise to provide a theoretical basis for alleviating cognitive and emotional disorders induced by microgravity environments.

Objective To investigate the impact of silymarin(SM)on the malignant growth of glioma cells and the regulatory mechanism on the miR-124-3p/WEE1 axis.Methods Glioma U87 cells were grouped into control,SM low,medium,and high concentration groups,and SM high concentration + miR-124-3p inhibitor group(SM high + miR-124-3p inhibitor group).CCK-8 was used to measure the proli-feration rate of cells;Transwell? assay was applied to assay the migration and invasion of cells;cell cycle progression was detected by flow cytometry;Western blotting was applied to measure the expression of cyclin D1 and apoptosis-related proteins;the levels of miR-124-3p and WEE1 mRNA were determined by qRT-PCR;and a luciferase activity test was applied to verify the targeting relationship between miR-124-3p and WEE1;in addition,the establishment,administration,and analysis of a NOD/SCID mouse model of intracranial trans-planted tumor were conducted.Results Compared with the control group,the cell proliferation,the numbers of migrating and invading cells,the expression of cyclin D1,and the level of WEE1 mRNA in the various SM treatment groups decreased,the number of cells in G0/G1 phase,the expression of cleaved caspase-8,cleaved caspase-9,cleaved caspase-3 and miR-124-3p increased(P＜0.05);furthermore,transfection of miR-124-3p inhibitor reversed the inhibitory effect of SM on the malignant behavior of glioma cells.In vivo experiments with mice showed that the weights and volumes of tumors in the SM treatment group were lower than those in the model group(P＜0.05),and there was no discernible change in the weight of the mice(P＞0.05).Conclusion SM can inhibit the malignant growth of glioma cells by upregulating miR-124-3p and downregulating WEE1.

Inguinal hernia (IH) is a common complication in prostate cancer patients undergoing radical prostatectomy (RP), which seriously affects their quality of life.The repair of IH is often challenging due to the adhesion in the inguinal area caused by previous surgery.This article focuses on the risk factors for IH after RP, including surgical approach, postoperative anatomical changes, low body mass index (BMI), muscle loss, advanced age, international prostate symptom score (IPSS), and patent sheath process, and prevention strategies for IH after RP, including robot assisted laparoscopic radical prostatectomy (RARP) with IH repair, preservation of the posterior pubic space in RARP, treatment of the spermatic cord, extraperitoneal RARP, and intraoperative management of the sheath like process, aiming to provide reference for urologists and to improve patients' quality of life.

This study aimed to investigate the therapeutic effects of Morinda officinalis iridoid glycosides (MOIG) on bone loss of rheumatoid arthritis (RA) rats, and the mechanism of osteoclast function and activity induced by lipopolysaccharide (LPS). RA rats were established by injecting bovin type II collagen. The Bio-ethic Committee of Zhejiang Chinese Medical University approved all experimental protocols associated with this study (IACUC-20180410-03). The collagen-induced arthritis (CIA) rats were administered drug by gavage for 8 weeks; the femoral trabecular micro-structure changes were observed in CIA rats by micro-CT; the LPS-induced osteoclasts model further observed the effect and mechanism of anti-inflammatory osteoporosis in vitro. The results indicated that MOIG could markedly increase bone mineral density (BMD) in CIA rats, improve trabecular micro-structure. In vitro studies demonstrated that MOIG could significantly inhibit osteoclastogensis and differentiation, suppress tartrate resistant acid phosphatase (TRAP) activity, F-actin ring formation, TNF receptor associated factor 6 (TRAF6) recruitment, and inhibitor of nuclear factor kappa-Bα (IκBα) degradation as well as p65 phosphorylation, thereby repressing nuclear factor kappa-B (NF-κB) signaling pathway activation. Subsequently, MOIG effectively inhibited osteoclast nuclear factor of activated T-cells c1 (NFATc1) and cellular oncogene Fos (c-Fos) expression, as well as bone resorption related protein activity including matrix metalloprotein 9 (MMP-9) and cathepsin K (CtsK). Meanwhile, MOIG also repressed the phosphorylation expression of Janus activating kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3), thereby inhibiting JAK2/STAT3 signaling pathway activation. Moreover, further studies found that MOIG could suppress glycogen synthase kinase-3β (GSK-3β) activity, and GSK-3β gene silencing could markedly inhibit oetsoclast F-actin ring formation as well as the phosphorylation expression of p65 and STAT3. Of note, compared with GSK-3β gene silencing group, there was no significant difference in the group treated with both MOIG with GSK-3β gene silencing simultaneously. Thus, the results suggested that MOIG may inhibit NF-κB signaling pathway and JAK2/STAT3 signaling pathway activation via regulating GSK-3β, thereby alleviating bone destruction in RA.