OBJECTIVES: To investigate the effect of in vitro cultured calculus bovis (ICCB) on cerebral ischemia/reperfusion injury (CIRI) and its mechanism. METHODS: A CIRI rat model and a cell model were induced by middle cerebral artery occlusion (MCAO) in Sprague Dawley rats and oxygen glucose deprivation/reperfusion (OGD/R) in BV2 cells, respectively. The CIRI rat model was evaluated using the modified neurological severity score (mNSS), brain water content, and cerebral infarction volume after 1.5 h of ischemia followed by 72 h of reperfusion. Histopathological changes in the cortex and hippocampal CA1 region were observed with hematoxylin and eosin staining. Microglial polarization and NOD-like receptor thermal protein domain associated protein (NLRP) 3 inflammasome expression in the cortex were examined by immunofluorescence. BV2 cell viability was measured via MTT assay after treatment with ICCB and Nigericin. The expressions of NLRP3, ASC, caspase-1 proteins and inflammatory cytokines were detected with Western blotting in OGD/R treated BV2 cells (0.5 h OGD+24 h reperfusion) and in cells pretreated with Nigericin for 24 h. RESULTS: ICCB treatment significantly improved neurological function, reduced cerebral infarct volume and brain water content, and mitigated pathological damage in the cortical and hippocampal CA1 regions of rats subjected to CIRI (all P<0.05). ICCB promoted the transition of cortical microglia from M1 to M2 phenotypes and suppressed NLRP3 activation in microglial cells (all P<0.01). ICCB significantly down-regulated the expression of NLRP3, ASC, and caspase-1 proteins, and reduced the secretion of IL-18 and IL-1β in BV2 cells of OGD/R model (all P<0.01). In addition, Nigericin significantly reversed the salvage effect of ICCB on model cells (both P<0.01) and the modulation of inflammatory cytokines (P<0.05). CONCLUSIONS ICCB exerts a protective effect against CIRI by mitigating neuroinflammation, through the reduction of M1 microglial polarization, promotion of M2 conversion, and suppression of the NLRP3/ASC/caspase-1 signaling pathway.
Animals ; Rats, Sprague-Dawley ; Reperfusion Injury/prevention & control* ; Microglia/metabolism* ; Rats ; NLR Family, Pyrin Domain-Containing 3 Protein ; Brain Ischemia/metabolism* ; Male

This study aims to evaluate the application of flower petal-structured electrodes in pulsed field ablation (PFA) technology, with a particular focus on their performance in myocardial tissue ablation. Through a combination of simulation techniques and in vitro experiments, the study investigates the effects of different voltage levels, electrode-to-tissue contact distances, and their impact on ablation depth, continuity, and transmurality. The research methods include the construction of a myocardial tissue simulation model, electric field distribution simulation using COMSOL Multiphysics, and in vitro ablation experiments on potato tissue. The results indicate that as voltage increases, the ablation depth significantly increases. At a voltage of 2500 V, a transmural ablation depth of 4 mm can be achieved, and the ablation area remains relatively continuous. The in vitro experiments confirm the consistency of the simulation results, and pulsed field ablation does not induce significant temperature rise, confirming its non-thermal characteristic. The conclusion suggests that PFA technology requires less electrode contact and offers higher ablation efficiency, providing a new technological pathway for the clinical treatment of atrial fibrillation and effectively reducing the risk of complications associated with traditional ablation techniques.
Electrodes ; Catheter Ablation/instrumentation* ; Computer Simulation ; Flowers ; Atrial Fibrillation/surgery* ; Myocardium

Objective:To investigate the mechanism by which hydrogen alleviates myocardial ischemia/reperfusion injury (IRI) through ultrasound-targeted destruction of hydrogen-loaded phospholipid microbubbles in the ischemic myocardium of rats.Methods:A total of 45 rats were randomly divided into three groups: sham operation group, IRI group (model group), and hydrogen treatment group (experimental group), with 15 rats in each group. A rat model of myocardial IRI was established. Rats in the sham group received 0.2 ml of normal saline via the tail vein, those in the model group received 0.2 ml of phospholipid microbubbles without hydrogen, and those in the treatment group received 0.2 ml of hydrogen-loaded phospholipid microbubbles. After 24 hours, cardiac function was assessed by left ventricular ejection fraction (LVEF) and left ventricular fractional shortening (LVFS). Serum levels of cardiac troponin I (cTnI), creatine kinase-myocardial band (CK-MB), lactate dehydrogenase (LDH), and interleukin-6 (IL-6) were measured by enzyme-linked immunosorbent assay (ELISA). Expression levels of Janus knase 2(JAK2), signal transducer and activator of transcription 3(STAT3), phosphorylated JAK2(p-JAK2), and phosphorylated STAT3(p-STAT3) proteins in myocardial tissue were detected by Western blot. Myocardial infarct size was evaluated by 2,3,5-triphenyltetrazolium chloride (TTC) staining, and myocardial histopathological changes were observed by hematoxylin-eosin (HE) staining.Results:After 24 hours of reperfusion, LVEF [(54.26±2.92) % vs (45.77±27%)] and LVFS [(24.11±1.68) % vs (19.50±1.19%)] were significantly higher in the treatment group than in the model group (both P<0.001). ELISA results showed that levels of CK-MB [(13.58±2.07) μg/L vs (20.07±1.57) μg/L], LDH [(47.76±8.32) μg/L vs (74.39±10.19) μg/L], cTnI [(7.50±0.26) μg/L vs (9.05±0.34) μg/L], and IL-6 [(121.34±8.97) ng/L vs (156.99±6.46) ng/L] were significantly lower in the treatment group compared with the model group (all P<0.001). TTC staining revealed a smaller infarct size in the treatment group [(48.77±2.68)%] than in the model group [(63.53±3.10)%, P<0.001]. Western blot analysis showed that the expression levels of JAK2 and p-STAT3 proteins were significantly lower in the treatment group than in the model group ( P<0.05). HE staining showed no pathological abnormalities in major organs (heart, liver, spleen, lung, and kidney) following hydrogen microbubble treatment. Conclusions:Ultrasound-targeted destruction of hydrogen microbubbles enables local hydrogen release in the myocardium, which downregulates IL-6 and inhibits activation of the JAK/STAT signaling pathway, thereby attenuating inflammation and reducing ischemia/reperfusion injury.

Objective In response to the clinical needs for personalized wrist orthoses,a topological optimization design method was proposed to achieve an integrated macro-and micro-structural optimization of a personalized,lightweight,and comfortable wrist orthosis.Methods A composite biomechanical finite element model of the wrist orthosis and upper limb was established to quantify the effects of the orthosis geometry on its fixation performance and comfort.A multi-condition topological optimization and microstructure design approach was employed to optimize the non-load-bearing areas of the orthosis.The orthosis was manufactured using three-dimensional(3D)-printed polyether ether ketone(PEEK),and the feasibility of the design was validated.Results While maintaining mechanical strength,the weight of the 3D-printed PEEK orthosis was reduced by 28％compared to the traditional orthoses.Both the pressure at the skin contact interface and the results of a subjective questionnaire indicated that test subjects experienced a high level of comfort wearing the orthosis.Conclusions The orthosis design achieved personalization,lightweight structure,and high comfort while ensuring mechanical strength and fixation performance.

Objective:To investigate the effect of DEAD-box helicase (DDX) 21 on myocardial ischemia-reperfusion (I/R) injury and its potential mechanisms.Methods:In vivo, adult male Bama pigs and C57BL/6J mice were used to establish a myocardial I/R injury model by ligating the left anterior descending coronary artery, with sham-operated groups set as controls. The expression of DDX21 in myocardium after I/R injury was assessed by quantitative real-time PCR (qRT-PCR), Western blot, and immunofluorescence staining. Following the establishment of the myocardial I/R injury model in mice, AAV9 vectors with cardiac-specific expression were injected in situ into the peri-infarct region (The I/R+DDX21 group, I/R+negative control (NC) group, I/R+sh-NC group and I/R+sh-DDX21 group were injected with AAV9:cTnT-DDX21, AAV9:cTnT-NC, AAV9:cTnT-sh-NC and AAV9:cTnT-sh-DDX21, respectively). Additionally, the I/R+A-485 group received intraperitoneal injections of the cAMP response element-binding protein (CREB) binding protein inhibitor A-485, while the I/R+PBS group was injected with an equivalent volume of phosphate-buffered saline (PBS) as the control. Echocardiography was performed on postoperative days 1 and 28 to evaluate cardiac function (left ventricular ejection fraction and fractional shortening). At 28 days post-surgery, mice were euthanized and heart tissues were harvested for histological sectioning. Myocardial fibrosis was evaluated using Masson′s trichrome staining. In vitro, primary cardiomyocytes were isolated from neonatal day 1 C57BL/6J mice using enzymatic digestion method. Cardiomyocytes were transfected with plasmids or small interfering RNA (siRNA). The cardiomyocytes transfected with DDX21-siRNA were assigned to the siDDX21 group, those transfected with the DDX21 plasmid were assigned to the DDX21 group, and those transfected with the corresponding empty plasmid or siRNA were assigned to the NC group. Additionally, cardiomyocytes were treated with A-485 (A-485 group) or PBS (PBS group). An oxygen-glucose deprivation/reoxygenation (OGD/R) model was used to simulate cellular injury. Transcriptome sequencing was performed to identify downstream mechanisms of DDX21. Differential gene expression analysis was conducted using software such as DESeq2, and alternative splicing events in the mRNA transcriptome were analyzed using rMATS software. Mitochondrial superoxide, mitochondrial membrane potential, ATP content, and mitochondrial respiratory chain complex enzyme activity in cardiomyocytes were detected using immunofluorescence staining and commercial assay kits. The oxidative phosphorylation level of the cells was assessed by the Seahorse extracellular flux analyzer. Acetylated DDX21 levels were measured using co-immunoprecipitation and Western blot assays.Results:The expression levels of DDX21 in myocardium from the Bama pigs and mice in the I/R injury model were significantly higher than those in the sham group (all P<0.001). Echocardiographic results showed that at 28 days post-surgery, compared to the I/R+NC group, the I/R+DDX21 group exhibited higher left ventricular ejection fraction and fractional shortening, while the I/R+sh-DDX21 group showed lower values; Masson staining results demonstrated that, compared to the I/R+NC group, the myocardial fibrosis area in the I/R+DDX21 group was significantly reduced, whereas it was significantly increased in the I/R+sh-DDX21 group (all P<0.001). Transcriptomic sequencing results suggested that DDX21 may influence myocardial injury by regulating mitochondrial metabolic activity. In vitro, compared to the OGD/R+NC group, the OGD/R+DDX21 group exhibited lower mitochondrial superoxide levels, higher polymer/monomer ratio, maximal oxygen consumption, reserve capacity, and ATP content. In contrast, the OGD/R+siDDX21 group showed the opposite results, with reduced activity of mitochondrial respiratory chain complex V (all P<0.05). Mechanistically, rMATS software and other analyses indicated that knockdown of DDX21 affected the alternative 3′ splicing sites of ATP5J precursor mRNA, inhibiting the splicing of certain exonic sequences. Overexpression of DDX21 upregulated both mRNA and protein levels of ATP5J. Co-immunoprecipitation experiments showed that, compared to the PBS group, acetylated DDX21 levels were reduced in the A-485 group. Further in vivo experiments showed that, compared to the I/R+PBS group, the I/R+A-485 group exhibited higher left ventricular ejection fraction and fractional shortening, and a lower proportion of left ventricular fibrosis (all P<0.001). Conclusions:DDX21 improves cardiomyocyte energy metabolism and alleviates I/R injury by regulating the alternative splicing of ATP5J. A-485 holds potential as a novel small molecule candidate for the treatment of myocardial injury.

Objective:To investigate the biological role and molecular mechanism of checkpoint kinase 1 (CHK1) in delaying cardiac aging in mice.Methods:In vitro, a senescence model of H9C2 cells (a cardiomyocyte line) was induced using H 2O 2. A control group (without H 2O 2 treatment) and three H 2O 2-treated groups (at concentrations of 10, 30, and 50 μmol/L) were set up. The CCK-8 assay was used to evaluate the proliferative activity of cells in each group; Western blot analysis was employed to detect the expression level of CHK1; and quantitative real-time polymerase chain reaction (qRT-PCR) was utilized to determine the messenger RNA (mRNA) expression levels of P16 and interleukin-1β (IL-1β). In vivo, C57BL/6 wild-type mice aged 2 months ( n=15) and 24 months ( n=40), as well as myocardial-specific CHK1-overexpressing (CHK1-TG) mice aged 2 months ( n=15) and 24 months ( n=40), were selected. The mice were divided into four groups based on age and genotype: 2-month-old wild-type (WT-2M), 24-month-old wild-type (WT-24M), 2-month-old CHK1-TG (CHK1-TG-2M), and 24-month-old CHK1-TG (CHK1-TG-24M). Echocardiography was used to evaluate cardiac function of mice in the WT-24M and CHK1-TG-24M groups. Western blot analysis was conducted to measure the protein expression levels of CHK1, total Ras-related protein 1 (Rap1), NADPH oxidase 4 (Nox4), and Rap1-guanosine triphosphate (Rap1-GTP, the active form of Rap1) in the cardiac tissue of mice in each group. qRT-PCR was used to detect the messenger RNA (mRNA) expression levels of CHK1, collagen type Ⅰ (Coll1), matrix metalloproteinase-2 (Mmp2), alpha-smooth muscle actin (α-SMA), P53, P21, P16, thioredoxin 1 (Trx1), thioredoxin reductase (TrxR), glutathione recluctase (GR), Rap1, and Nox4. Immunofluorescence staining was employed to determine the protein expression levels of P53, P21, and P16, as well as the proportion of histone H2AX phosphorylation-positive cells. Dihydroethidium (DHE) staining was used to detect the relative intensity of DHE. Wheat germ agglutinin staining, HE staining, Masson staining and Sirius red staining were applied to measure the cross-sectional area of cardiomyocytes, cardiac morphology, and myocardial fibrosis area. Mice in the WT-24M and CHK1-TG-24M groups were intraperitoneally injected with the Rap1 activity inhibitor GGTI298 (25 μmol/kg). After injection, the oxidative stress damage in the cardiac tissue of the mice was detected, along with the mRNA expression levels of fibrosis-related indicators (Coll1, Mmp2, and α-SMA) and cell cycle inhibitory proteins (P16, P21, and P53). Results:A concentration of 30 μmol/L was determined as the optimal concentration for establishing an H 2O 2-induced senescence model of myocardial cells in vitro. The expression level of CHK1 in H9C2 cells of the 30 μmol/L H 2O 2 group was lower than that in the control group ( P<0.05). Echocardiographic examination showed that the left ventricular ejection fraction ((61.08±1.13)% vs. (52.55±2.02)%) and fractional shortening ((31.80±1.27)% vs. (25.18±1.59)%) of mice in the CHK1-TG-24M group were higher than those in the WT-24M group (both P<0.05). qRT-PCR and Western blot analysis revealed that, compared with the WT-24M group, mice in CHK1-TG-24M group had higher expression levels of CHK1 and its mRNA, lower expression levels of Nox4 and its mRNA, and higher expression level of Rap1-guanosine triphosphate (Rap1-GTP) (all P<0.05). However, there were no statistically significant differences in the total expression level of Rap1 and its mRNA between the two groups (both P>0.05). In addition, the mRNA expression levels of Coll1, Mmp2, and α-SMA in myocardial tissue of mice in the CHK1-TG-24M group were lower than those in the WT-24M group (all P<0.05). Immunofluorescence staining results showed that the expression levels of P53, P21, and P16 proteins, as well as the proportion of phosphorylated histone H2AX-positive cells in myocardial tissue of mice in the WT-24M group were higher than those in the CHK1-TG-24M group (all P<0.05). qRT-PCR further confirmed that the mRNA expression levels of the above-mentioned proteins in cardiac tissue of mice in the WT-24M group were higher than those in the CHK1-TG-24M group (all P<0.05). DHE staining results indicated that the relative intensity of DHE in cardiac tissue of mice in the CHK1-TG-24M group was lower than that in the WT-24M group ( P<0.05). Meanwhile, the left ventricular internal diameter, cross-sectional area of cardiomyocytes, and myocardial fibrosis area of mice in the CHK1-TG-24M group were all smaller than those in the WT-24M group (all P<0.05). Furthermore, the degree of DNA damage in cardiac tissue as well as the mRNA levels of fibrosis-related indicators (Coll1, Mmp2, and α-SMA) and cell cycle inhibitory proteins (P53, P21, P16) in mice of the WT-24M+GGTI298 group were higher than those in the WT-24M group and the CHK1-TG-24M+GGTI298 group (all P<0.05). Conclusion:CHK1 alleviates oxidative stress-induced damage in mouse cardiomyocytes by activating the Rap1/Nox4 signaling pathway, thereby delaying cardiac aging in mice.

Migraine disorders, a common disabling headache condition, is the second largest contributor to global neurological disability-adjusted life-years. Since 2000, migraine surgery has been pioneered and elaborated by plastic surgeons. For certain patients, migraine episodes are associated with "trigger sites" caused by compression of the peripheral nerves in the head and neck region. Botulinum toxin type A injections or nerve blocks, clinical manifestations and image studies can help confirm the trigger sites. Surgical deactivation of the trigger sites can effectively alleviate or eliminate migraine disorders, with the optimal result incorporated with experience and principles of plastic and cosmetic surgery.

This article reviews a study from Cell Metabolism, titled "Transketolase promotes MAFLD by limiting inosine-induced mitochondrial activity" [Tong L, Chen Z, Li Y, et al. Cell Metab. 2024, 36(5): 1013-1029.e5], and discusses its scientific implications. The study explores selective hepatic insulin resistance in metabolic dysfuction-associated fatty liver disease(MAFLD), where insulin stimulates lipid synthesis but not gluconeogenesis in resistant livers. It reveals the key role of transketolase(TKT) in MAFLD progression by disrupting pentose metabolism and nucleoside homeostasis, leading to inhibited mitochondrial activity. Targeting TKT with N-acetylgalactosamine-conjugated small interfering RNA(GalNAc-siTKT) improved metabolic dysfunction-associated steatohepatitis(MASH) and hepatic fibrosis.