AIM: To study the effects of disulfiram/copper complex (DSF/Cu) on ultrastructures and mechan-ical properties of human breast cancer and normal breast epithelial cells by atomic force microscopy (AFM) based on the nanoscale resolution and piconewton force measurement level.METHODS: The change of cell cycle and apoptotic rate of MCF-7 cells and MCF-10A cells induced by DSF/Cu were compared by flow cytometry.The cell surface morphology, ultra-structure, height, width and roughness were detected by AFM.The effects of DSF/Cu on the hardness (Young’s modu-lus) of the 2 kinds of cells were determined by AFM with indentation technique.RESULTS: DSF/Cu significantly in-duced apoptosis of the MCF-7 cells in a dose-dependent manner, whereas had little effect on the MCF-10A cells.The cell cycle analysis showed that DSF/Cu induced G2 /M arrest in the MCF-7 cells, but led to G0 /G1 arrest in the MCF-10A cells.The AFM images showed that the MCF-7 cells shrank and showed smaller and smoother morphology, and the filopo-dia were retracted obviously, even some became into lamellipodia, or disappeared completely after treated with DSF/Cu at concentrations of 400 and 800 nmol/L.The quantitative analysis indicated that the MCF-7 cells showed smaller width and
larger height, and the root mean square roughness and average roughness were decreased significantly in a dose-dependent manner after treated with DSF/Cu at concentrations of 400 and 800 nmol/L.However, little effect in the MCF-10A cells was observed.The biomechanics test at a single cell level demonstrated that the Young’s modulus of the MCF-7 cells and MCF-10A cells were both increased, yet the proportion increased in the MCF-7 cells was much higher than that in the MCF-10A cells after treated with DSF/Cu at concentrations of 400 and 800 nmol/L.CONCLUSION: DSF/Cu has strong antitumor effect on breast cancer with high efficiency and low toxicity by changing the properties of the biomechanics specifically.

We implemented tutorial system by relying on young teachers in hospitals and taking undergraduates in Capital Medical University as the main body aiming at cultivating students' clinical and scientific research abilities.Practice has proved that tutor system method is of great value to cultivate students' comprehensive clinical ability,scientific research ability and innovation ability.Meanwhile,this method is effective in improving clinical teaching effect.

Aim To explore the effects and mecha-nisms of choro-oxime derivatives on spatial learning and memory impairment in Kunming mice and SD rats induced by scopolamine and Aβ1-42 , respectively. Methods 40 Kunming mice were randomly divided into 5 groups: control group, model group, donepezil treatment group, arimoclomol treatment group and TCO-2 treatment group. There were 8 mice in each group. Mice of control group were established by intra-peritoneal injection of saline, and mice of other groups were injected with scopolamine and caused memory im-pairment. Both control group and model group were treated with solvent by intraperitoneal administration;donepezil treatment group received donepezil by intra-gastric administration; arimoclomol treatment group and TCO-2 treatment group were given the correspond-ing drugs by abdominal injection, respectively. The solvent and drugs were given at the same time every morning for 8 days. Spatial learning and memory abili-ty were tested by Morris water maze from the fifth day of the drugs administration. 40 SD rats were divided into 5 groups the same as the dementia model men-tioned above. Mice of control group were established by intracerebroventricular injection of saline, and mice of other groups were injected with insoluble Aβ1-42 to be induced of memory impairment. Solvent and drugs were also delivered as mentioned above. Morris water maze was carried out from the fifth day of the drug de-livery. After that, acetyl cholinesterase activity of hip-pocampus was tested with acetyl cholinesterase reagent kit; the content of Aβ1-42 in hippocampus was meas-ured by ELISA assay kit;the expression of phosphoryl-ated tau proteins was detected by Western Blot. Re-sults In both two dementia models, choro-oxime de-rivatives could improve the spatial learning and memory ability, shorten the escape latency and increase the times of crossing the former platform. Choro-oxime de-rivatives could also inhibit the acetyl cholinesterase ac-tivity in animal brain, decrease the concentration of Aβ1-42 and the expression of phosphorylated tau pro-teins in the dementia rats’ hippocampus. Conclusions Spatial learning and memory deficits induced by sco-polamine and Aβ1-42 could be reversed by choro-oxime derivatives. It may be concerned with enhancement of the cholinergic system functions and reduction of the levels of Aβ1-42 and phosphorylated tau proteins in the brain.

Eyes are important human organs, and retinal degenerative diseases can cause irreversible blindness. Up to date, cell transplantation is an important method for the treatment of degenerative eye diseases. Because the number of adult eye stem cells is rather limited,it is still very difficult to activate endogenous adult stem cells as a way to treat degenerative eye diseases. Therefore,photoreceptor cells ( photoreceptor) and retinal pigment epithelium (RPE) cells generated from pluripotent stem cells include embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) can be an effective source of cells for transplantation. This article described the major types of retinal degenerative diseases including retinitis pigmentosa ( RP) and age-related macular degeneration (AMD) by focusing on their pathological process and development of stem cell transplantation. We reviewed the related stem cell therapies and research status of two different vulnerable cells, RPE and photoreceptor cell in degenerative ophthalmopathy. We also discussed the value and limitations of cell transplantation therapy and the prospect of future treatment.

Aim To study the mechanism of DSF-Cu induced apoptosis of human nasopharyngeal carcinoma CNE-2 Z cells by affecting the function of mitochondria and cytoskeleton. Methods The cell cycle,the rate of apotosis,the levels of intracellular ROS and MMP in CNE-2 Z cells were tested by flow cytometry after trea-ted with different concentration of DSF-Cu. The chan-ges of the cell surface morphology, ultrastructure, cell height, width and roughness were detected by AFM. The distribution and reorganization of cytoskeleton F-actin were observed by Laser scanning confocal micro-scope. Results Cells were incubated with different concentration of DSF-Cu ( 0 ~200 nmol · L-1 ) for 24 h, the apoptotic ratio increased significantly and the treatment of DSF-Cu resulted in a concentration-de-pendent accumulation of CNE-2Z cells in G2/M phase. Furthermore,the treatment of DSF-Cu was able to in-crease the production of intracellular ROS and decrease the MMP in CNE-2Z cells. In addition,AFM imaging showed that compared to the control group,with the in-crease of DSF-Cu concentration,the CNE-2Z cells be-came smaller, cytoplasm condensed, the height in-creased,and the surface roughness reduced. Moreover, the filopodia became shorter, shrinked and even com-pletely destroyed after treated with different concentra-tion of DSF-Cu. At last,the LSCM image showed that the fluorescence intensity of F-actin networks was de-creased, then the structure was rearranged and de-stroyed obviously by treated with DSF-Cu. Conclusion DSF-Cu can induce apoptosis and arrest cell cycle at G2/M phase in CNE-2Z cell through a mitochondria-dependent pathway. Above findings highlight the appli-cations of AFM at the single cell level for the investiga-tion of antineoplastic drug in nasopharyngeal carcinoma therapy.

BACKGROUND:In cartilage degeneration in osteoarthritis,cytokines and signaling pathways that target chondrocytes play an important role. OBJECTIVE:To review the latest research progress of osteoarthritis related cytokines and signaling pathways in recent years,such as the mechanism of action and treatment modalities,in order to provide a basis for future exploration of new therapeutic targets and modalities. METHODS:Literature search was conducted on CNKI,WanFang,VIP,PubMed,Web of Science,and Medline databases.Chinese search terms were"osteoarthritis,cytokines,signal pathway,chondrocyte,inflammation,treatment".Finally,60 papers were included for review. RESULTS AND CONCLUSION:(1)In current studies,it is believed that the specific mechanism of osteoarthritis is not clear,and a large number of studies have shown that osteoarthritis is strongly associated with cytokines and signaling pathways,which is a complex process of action.Relevant studies taking cytokines and signaling pathways as therapeutic breakthroughs are also the current hot spot.(2)The receptor antagonists of pro-inflammatory factors such as interleukin 1 are not effective in the treatment of osteoarthritis,and more studies turn to gene therapy.(3)The therapeutic methods of transforming growth factor β,recombinant factors of Wnt signaling pathway,gene therapy and mesenchymal stem cells have obtained positive research results.However,basic and clinical studies on safety and efficacy are likely to be conducted in future studies.(4)At present,relevant therapeutic methods such as platelet-rich plasma have been widely used in clinical practice,while recombinant factor,gene therapy and mesenchymal stem cell therapy are all in the research stage,among which mesenchymal stem cell therapy and gene therapy are expected to make breakthroughs in the field of cartilage repair and regeneration,and are worthy of expectation in the future.However,more clinical and basic studies are needed to verify its effectiveness and safety,explore its mechanism of action and scope of application,and set standards for its clinical use.

Objective:To evaluate the relationship between silence information regulator 1 (SIRT1) and signal transducers and activators of transcription 3 (STAT3) acetylation during high glucose-induced cardiac microvascular endothelial cell injury.Methods:Cardiac microvascular endothelial cells of Sprague-Dawley rats were cultured.The cells at the logarithmic growth phase were selected and divided into 3 groups ( n=24 each) using a random number table method: control group (C group), high glucose group (HG group) and high glucose+ SIRT1 agonist SRT1720 group (HG+ SRT group). The cardiac microvascular endothelial cells were seeded in a 6- or 96-well cell culture plate at a density of 2×10 5 cells/ml.When the cell density reached 50%, the culture medium was then replaced with high-glucose (glucose 33 mmol/L) DMEM culture medium containing with 10% fetal bovine serum and 1% double antibody in HG and HG+ SRT groups.In group HG+ SRT, 20 μmol/L SRT1720 was added simultaneously, and the cells were cultured at 37 ℃ in an incubator with 5% CO 2 for 24 h. The cell viability was determined by CCK-8 assay, the activity of superoxide dismutase (SOD) was detected using a spectrophotometer, the levels of lactic dehydrogenase (LDH), interleukin-6 (IL-6) and tumor necrosis factor-β (TNF-β) in the supernatant were detected by enzyme-linked immunosorbent assay, and the expression of SIRT1, acetylated STAT3 (ac-STAT3) and phosphorylated STAT3 (p-STAT3) was determined by Western blot. Results:Compared with C group, the cell viability and SOD activity were significantly decreased, levels of LDH, IL-6 and TNF-β in the supernatant were increased, expression of SIRT1 was down-regulated, and expression of ac-STAT3 and p-STAT3 was up-regulated in group HG and group HG+ SRT ( P<0.05). Compared with group HG, the cell viability and SOD activity were significantly increased, levels of LDH, IL-6 and TNF-β in the supernatant were decreased, expression of SIRT1 was up-regulated, and expression of ac-STAT3 and p-STAT3 was down-regulated in group HG+ SRT ( P<0.05). Conclusion:SIRT1 can alleviate high glucose-induced cardiac microvascular endothelial cell injury by promoting STAT3 deacetylation.

Objective:To evaluate the relationship between protein kinase RNA-like endoplasmic reticulum kinase (PERK) pathway-mediated endoplasmic reticulum stress and the reduction of cerebral ischemia-reperfusion (I/R) injury by dexmedetomidine in mice by the in vivo experiment and the cell experiment. Methods:In the in vivo experiment, 20 healthy clean-grade male mice, aged 6-8 weeks, weighing 20-30 g, were divided into 4 groups ( n=5 each) using a random number table method: sham operation group (group S), sham operation+ dexmedetomidine group (group SD), cerebral I/R group (group IR) and cerebral I/R+ dexmedetomidine group (group IRD). Cerebral I/R was established by two-vessel occlusion plus hypotension.Dexmedetomidine 25 μg/kg was intraperitoneally injected at 10 min of ischemia in group IRD and at the corresponding time point in group SD.Neurological function was assessed using modified neurological severity score at 1 h of reperfusion.The animals were then sacrificed and brain tissues were taken for determination of the expression of endoplasmic reticulum stress-related proteins such as immunoglobulin heavy chain-binding protein (BIP), eukaryotic translation initiation factor 2α (EIF-2α), phosphorylated EIF-2α (p-EIF-2α), PERK and phosphorylated PERK (p-PERK) (by Wester blot). In the cell experiment, a mouse hippocampal neuronal cell line was selected and divided into 4 groups ( n=12 each) using a random number table method: control group (group C), oxygen-glucose deprivation/restoration (OGD/R) group (group OGD/R), OGD/R+ dexmedetomidine group (group OGD/R+ D) and OGD/R+ ISRIB (PERK pathway inhibitor) group (group OGD/R+ ISRIB). Cells were exposed to 94%N 2-5%CO 2-1%O 2 and incubated in a low-glucose DMEM medium for 6 h followed by restoration to establish OGD/R model.At 30 min before OGD, dexmedetomidine (final concentration 5 mmol/L) was added in group OGD/R+ D, and ISRIB (final concentration 10 mmol/L) was added in group OGD/R+ ISRIB.After 12-h restoration was completed, the cell survival rate was detected by CCK-8 assay.At 24 of restoration, the expression of endoplasmic reticulum stress-related proteins was determined by Wester blot. Results:In the in vivo experiment, compared with group S, neurobehavioral score was significantly increased and the expression of BIP, p-EIF-2α and p-PERK in brain tissues was up-regulated in group IR ( P<0.05). Compared with group IR, neurobehavioral score was significantly decreased and the expression of BIP, p-EIF-2α and p-PERK in brain tissues was down-regulated in group IRD ( P<0.05). In the cell experiment, compared with group C, the expression of BIP, p-EIF-2α, PERK and p-PERK was significantly up-regulated, and the cell survival rate was decreased in group OGD/R ( P<0.05). Compared with group OGD/R, the expression of BIP, p-EIF-2α, PERK and p-PERK was significantly down-regulated, and the cell survival rate was increased in OGD/R+ D, OGD/R+ ISRIB groups ( P<0.05). Compared with group OGD/R+ ISRIB, the expression of PERK was significantly down-regulated ( P<0.05) and no significant change was found in the other parameters in group OGD/R+ D ( P>0.05). Conclusion:The mechanism by which dexmedetomidine reduces cerebral I/R injury may be related to inhibiting PERK pathway-mediated endoplasmic reticulum stress in mice.

Objective:To evaluate the role of protein O-linked beta-N-acetylglucosaminylation (O-GlcNAcylation) modification in oxidative stress injury in nerve cells of mice subjected to oxygen-glucose deprivation and restoration (OGD/R).Methods:The standard mouse hippocampal neuron cell line was inoculated on a culture plate or dish at a density of 5×10 4 cells/ml and divided into 4 groups ( n=20 each) using a random number table method: normal group (N group), O-(connection)N-acetylglucosamine hydrolase (OGA) inhibitor Thiamet G group (T group), OGD/R group (D/R group) and Thiamet G+ OGD/R complex sugar group (T-D/R group). The cells were exposed to a mixed gas of 94% N 2-5% CO 2-1% O 2 for 6 h in a low-glucose medium, then medium was replaced with a common medium for restoring oxygen and glucose, and the cells were cultured for 12 h. Thiamet G at a final concentration of 1 mmol/L was added to the culture medium at 4 h before OGD/R in T-D/R group, and the medium was replaced with a medium containing Thiamet G at a final concentration of 1 mmol/L at 4 h before extraction of cellular proteins.After oxygen and glucose restoration was completed, the accumulation of cellular ROS was measured using DCFH-DA staining, mitochondrial membrane potential was measured using Jc-1 staining, O-GlcNAc modification was determined by immunofluorescence, and the expression of nuclear factor E2-related factor 2 (Nrf2), c-Jun N-terminal kinase (JNK), phosphorylated JNK (p-JNK), and p53 tumor suppressor gene (p53) was detected using Western blot. Results:Compared with group N, the expression of O-GlcNAc in nerve cells was significantly up-regulated in group T, and the accumulation of ROS in nerve cells was significantly increased, JC-1 monomer was increased, JC-1 polymer was decreased, Nrf2 expression was down-regulated, and the expression of p-JNK and p53 was up-regulated in group D/R, and the expression of O-GlcNAc in nerve cells was up-regulated, the accumulation of ROS was increased, the polymerization of JC-1 monomer and JC-1 was increased, Nrf2 expression was down-regulated, and the expression of p-JNK and p53 was up-regulated in group T-D/R ( P<0.05). Compared with group D/R, the expression of O-GlcNAc in nerve cells was significantly up-regulated, the accumulation of ROS was decreased, JC-1 monomer was decreased, JC-1 polymer was increased, the expression of Nrf2 was up-regulated, and the expression of p-JNK and p53 was down-regulated in group T-D/R ( P<0.05). Conclusion:When mouse nerve cells are subjected to OGD/R, the protein O-GlcNAc modification as an endogenous protective mechanism is enhanced, which can reduce oxidative stress injury, and the mechanism may be related to regulating the Nrf2-mediated JNK pathway.

BACKGROUND:Studies have shown that posterior cruciate ligament injuries are associated with the anatomical morphology of the knee joint. OBJECTIVE:To explore anatomical morphological risk factors for posterior cruciate ligament injury. METHODS:The imaging data of 142 patients who visited Affiliated Hospital of Binzhou Medical University for knee joint problems from January 2015 to August 2022 were retrospectively analyzed.They were divided into posterior cruciate ligament injury group(n=71,including 49 males and 22 females)and posterior cruciate ligament intact group(n=71,including 49 males and 22 females).Intercondylar notch width,intercondylar notch height,bicondyle width,notch width index,angle of intercondylar notch,Blumensaat's line inlication angle,medial tibial spine height,lateral tibial spine height,tibial spine width,tibiofemoral consistency index,tibial plateau anterior-posterior diameter,medial tibial depth and patellar tendon-tibial shaft angle were measured on MRI images.Posterior tibial slope was measured on X-ray images.The above indicators were included in the logistic regression analysis for investigation. RESULTS AND CONCLUSION:(1)Univariate logistic regression analysis in men showed that tibial spine width,tibiofemoral consistency index,medial tibial depth,and posterior tibial slope were associated with posterior cruciate ligament injury(P<0.05).Multivariate binary logistic regression analysis showed that tibiofemoral consistency index and medial tibial depth were associated with posterior cruciate ligament damage(P<0.05).(2)Univariate logistic regression analysis in women showed that medial tibial spine height,lateral tibial spine height,tibial spine width,and posterior tibial slope were associated with posterior cruciate ligament injury(P<0.05).Multivariate binary logistic regression analysis showed that posterior tibial slope was associated with posterior cruciate ligament damage(P<0.05).(3)The receiver operating characteristic curve showed that tibiofemoral consistency index,medial tibial depth and posterior tibial slope had a certain predictive value on posterior cruciate ligament damage.(4)These findings suggest that anatomical morphological risk factors for posterior cruciate ligament injury differ between men and women,and tibial spine width and posterior tibial slope are common risk factors.In the male population,abnormal tibial spine width,tibiofemoral consistency index,medial tibial depth,and posterior tibial slope are easy to induce posterior cruciate ligament injury.In the female population,abnormal medial tibial spine height,lateral tibial spine height,tibial spine width,and posterior tibial slope are easy to induce posterior cruciate ligament injury.Clinicians can use the above risk factors to identify abnormal knee morphology,assess people at risk of posterior cruciate ligament injury,and provide preventive advice and guidance for treatment.