Objective To investigate the effect of MACC1 on RSL3-induced ferroptosis in colorectal cancer cells and explore its molecular mechanism.Methods MACC1 expression was detected in SW620,HCT116,LOVO and RKO cells using Western blotting.The effects of different concentrations of RSL3(an inducer of ferroptosis)or Fer-1(an inhibitor of ferroptosis)alone,or 10 μmol/L RLS3 combined with 10 μmol/L Fer-1,on viability of SW620 cells were examined using MTT assay.The survival of SW620 cells with mRNA interference of MACC1 was analyzed following treatment with RSL3,and RT-qPCR and Western blotting were performed to detect the changes in MACC1 expressions after RSL3 treatment at different concentrations and the changes in GPX4 expression after MACC1 knockdown.Flow cytometry and laser confocal microscopy were used to analyze the changes in ROS-induced lipid peroxidation in SW620 cells after MACC1 knockdown.Results SW620 cells had the highest MACC1 expression among the 4 colorectal cancer cell lines.Treatment with RSL3 significantly inhibited the viability of SW620 cells in a dose-dependent manner,while Fer-1 did not significantly affect the survival of SW620 cells.RSL3 alone reduced SW620 cell survival by 50%(P<0.01),and the combined treatment with RSL3 and Fer-1 caused no significant changes in cell survival(P>0.05).Treatment with RSL3 concentration-dependently suppressed MACC1 expressions at both the mRNA and protein levels in SW620 cells(P<0.01).MACC1 knockdown obviously enhanced the cytotoxic effect of RSL3,inhibited the expression of GPX4,and increased ROS-induced lipid peroxidation in SW620 cells(P<0.05).Conclusion MACC1 knockdown enhances RSL3-induced ferroptosis in cultured colorectal cancer cells by inhibiting the expression of GPX4.

Objective To investigate the effect of MACC1 on RSL3-induced ferroptosis in colorectal cancer cells and explore its molecular mechanism.Methods MACC1 expression was detected in SW620,HCT116,LOVO and RKO cells using Western blotting.The effects of different concentrations of RSL3(an inducer of ferroptosis)or Fer-1(an inhibitor of ferroptosis)alone,or 10 μmol/L RLS3 combined with 10 μmol/L Fer-1,on viability of SW620 cells were examined using MTT assay.The survival of SW620 cells with mRNA interference of MACC1 was analyzed following treatment with RSL3,and RT-qPCR and Western blotting were performed to detect the changes in MACC1 expressions after RSL3 treatment at different concentrations and the changes in GPX4 expression after MACC1 knockdown.Flow cytometry and laser confocal microscopy were used to analyze the changes in ROS-induced lipid peroxidation in SW620 cells after MACC1 knockdown.Results SW620 cells had the highest MACC1 expression among the 4 colorectal cancer cell lines.Treatment with RSL3 significantly inhibited the viability of SW620 cells in a dose-dependent manner,while Fer-1 did not significantly affect the survival of SW620 cells.RSL3 alone reduced SW620 cell survival by 50%(P<0.01),and the combined treatment with RSL3 and Fer-1 caused no significant changes in cell survival(P>0.05).Treatment with RSL3 concentration-dependently suppressed MACC1 expressions at both the mRNA and protein levels in SW620 cells(P<0.01).MACC1 knockdown obviously enhanced the cytotoxic effect of RSL3,inhibited the expression of GPX4,and increased ROS-induced lipid peroxidation in SW620 cells(P<0.05).Conclusion MACC1 knockdown enhances RSL3-induced ferroptosis in cultured colorectal cancer cells by inhibiting the expression of GPX4.

AIM:To investigate the molecular mechanism underlying ferroptosis induced by celastrol(Cel)in huamn pancreatic cancer cell line PANC-1.METHODS:The viability of PANC-1 cells was analyzed by MTT assay,and the effects of Cel on cell proliferation were analyzed using EdU and colony formation assays.Flow cytometry and fluores-cence microscopy were used to assess and observe changes in lipid reactive oxygen species(ROS)levels,respectively,while the levels of malondialdehyde(MDA),glutathione(GSH)and Fe2+were measured using specific kits.The protein expression of glutathione peroxidase 4(GPX4)was evaluated by Western blot,and GPX4 ubiquitination was measured by immunoprecipitation.RESULTS:It was found that the viability,proliferation and colony formation in PANC-1 cells de-creased gradually as the concentration of Cel increased.Addition of Cel alone to the cells reduced both cell rounding and viability,while treatment with ferrostatin-1(Fer-1)alone or in combination with Cel had no effect on either cell morpholo-gy or viability.Fluorescence staining of lipid ROS with BODIPYTM 581/591 C11 followed by flow cytometry analysis showed significantly increased levels of green fluorescence indicative of oxidized lipid ROS,which were further increased after treatment of the cells with Cel.Treatment of the cells with both Cel and Fer-1 reduced the green fluorescence and lip-id ROS levels.Treatment with Cel also increased the levels of MDA and Fe2+,relative to the controls,which reducing the levels of GSH,while addition of both Cel and Fer-1 to the cells restored the levels of MDA,Fe2+,and GSH to those of the control group.CONCLUSION:Treatment with Cel reduces the proliferation of pancreatic cancer cells by inducing fer-roptosis through promoting the ubiquitination and degradation of GPX4.

Objective To establish an animal model and evaluation system for lymph node metastasis of breast cancer.Methods A total of 60 female BALB/c mice(6～8 weeks old)were subjected,and then 6 models of lymph node metastasis(n=10)were constructed through injection at different parts in the mice with cell suspension of 4T1 breast cancer cells.Transgenic mice(n=5)of mouse mammary tumor virus-polyoma middle T antigen(MMTV-PyMT)were employed and served as model of spontaneous tumor metastasis.Then the advantages and disadvantages of different lymph node metastasis models were comprehensively evaluated from multiple aspects,such as operability,histomorphology and pathological detection,tumor growth rate and mouse survival.Results Among the 7 metastasis models,4 models of lymph node metastasis were successfully established.Among them,the PyMT mouse spontaneous tumorigenesis model showed the best clinical reproduction,with a tumorigenesis rate of up to 100％,but had a disadvantage of poor experimental standardization.The hind paw-popliteal lymph node model had the fastest lymph node metastasis,easy operation and high repeatability,and a tumorigenesis rate of 100％,indicating its suitable for lymph node metastasis related research.The thigh subcutaneous-inguinal lymph node model also successfully simulated lymph node metastasis,with simple operation and high repeatability,a tumorigenesis rate of up to 100％,but its metastasis time was slightly longer than the hind paw-popliteal lymph node model.The inguinal lymph node-contralateral lymph node model was also a successful lymph node metastasis model,but with difficult operation,only 50％tumor-bearing rate,and poor repeatability.Lymph node metastasis model was not successfully established in the other 3 tumor-bearing models(under the tongue-internal jugular scapular tongue muscle lymph node model,bone marrow-inguinal lymph node model and right upper back skin-axillary lymph node model)in a short time,with no tumor cells observed in the pathological sections.Conclusion Through the comprehensive comparison of multiple models,mouse hind paw-popliteal lymph node model is the most suitable for conducting related research.

AIM:This study aimed to explore the induction of ferroptosis in non-small-cell lung cancer(NSCLC)cells by tubeimoside II(TBMS II)and to elucidate the underlying molecular mechanisms.METHODS:H460 NSCLC cells were cultured in vitro.Cell survival rates were assessed by using MTT assays,and doses of TBMS II resulting in below 50%survival were selected for further experimentation.Cell migration was evaluated using Transwell assays and the effects of TBMS II on H460 cell proliferation were assessed by colony formation assays.Flow cytometry and fluores-cence microscopy were used to assess changes in lipid peroxidation(lipid ROS),and the levels of GSH,T-AOC,MDA,and Fe2+were measured using commercial kits.Protein levels of GPX4,SLC7A11,FTH1,NCOA4,P62,and LC3 were examined using Western blot.Changes in mitochondrial structure were detected by transmission electron microscopy,and immunofluorescence was used to assess LC3 co-localization of FTH1 and NCOA4,as well as co-localization of LC3 and NCOA4 with lysosomes.RESULTS:Compared with the control group,TBMS II dose-dependently reduced H460 cell via-bility,migration,and clone formation,accompanied by the appearance of vacuoles within the cells.TBMS II treatment al-so led to decreased GSH and T-AOC levels,while increasing the cellular contents of MDA,indicating oxidative stress.Ad-ditionally,there was a decrease in the expression of the antioxidant proteins SLC7A11 and GPX4 in the cells,while lipid ROS and Fe2+levels were increased in proportion to the TBMS II concentration.The ferroptosis inhibitor ferrostatin-1 re-versed cell death caused by TBMS II,suggesting ferroptosis induction.Furthermore,increasing the TBMS II concentra-tion resulted in an upregulation of the autophagy marker proteins LC3 II/LC3 I and P62,indicative of increased autopha-gy.TBMS II also affected mitochondrial morphology in the cells,as seen in reduced mitochondrial fluorescence intensity.Protein expression of NCOA4 increased with higher TBMS II concentrations,while that of FTH1 decreased.Co-localiza-tion of LC3 II with FTH1 and NCOA4,as well as the lysosomal association of LC3 II and FTH1,also increased in a dose-dependent manner.CONCLUSION:TBMS II induces ferritinophagy in H460 cells,leading to decreased cell viability and increased ferroptosis.

Objective:To investigate the characteristics of 18F-FDG and 18F-9-fluoropropyl-(+ )-dihydrotetrabenazine (FP-(+ )-DTBZ; short for DTBZ) brain vesicular monoamine transporter type 2 (VMAT2) PET/CT imaging and analyze its clinical diagnostic value in Parkinson′s disease (PD) patients with or without rapid eye movement sleep behaviour disorder (RBD). Methods:From July 2022 to June 2023, 50 patients clinically confirmed with primary PD who underwent 18F-FDG and 18F-DTBZ PET/CT imaging in the First Affiliated Hospital of Zhengzhou University were prospectively collected. Among them, 18 patients with PD accompanied by RBD (PD-RBD(+ ) group; 16 males, 2 females, age (59.2±9.3) years); 32 patients without RBD (PD-RBD(-) group; 16 males, 16 females, age (57.7±10.2) years). Moreover, 10 healthy controls matched with the age of PD patients were included (5 males, 5 females, age (60.3±9.6) years). 18F-DTBZ specific uptake ratio (SUR) of bilateral striatum, caudate nucleus, anterior putamen, posterior putamen, nucleus accumbens, substantia nigra and other brain regions were obtained with occipital cortex as the reference region. Striatal anterior-posterior gradient and other related semi-quantitative indicators were calculated according to the corresponding formula. One-way analysis of variance (the least significant difference (LSD)- t test), Kruskal-Wallis rank sum test (Bonferroni correction), independent-sample t test and Mann-Whitney U test were used to analyze the data. Pearson correlation and Spearman rank correlation analyses were used to evaluate the correlations. ROC curve analysis was also performed. The differences in global glucose metabolism in two groups were compared using statistical parametric mapping (SPM). Results:PD-RBD(+ ) group had a significantly lower Mini-Mental State Examination (MMSE) or PD Sleep Scale (PDSS) score than PD-RBD(-) group ( z values: -3.12, -3.08, both P<0.01), and its contralateral striatal anterior-posterior gradient of the predominantly affected limbs was significantly lower than that in PD-RBD(-) group ( t=-2.73, P=0.009). SPM analysis showed that the glucose metabolism in the contralateral prefrontal lobe was higher than that in the PD-RBD (-) group ( t values: 3.11-3.57, all P<0.001). 18F-DTBZ SUR in the bilateral striatum, caudate nucleus, anterior putamen, posterior putamen, nucleus accumbens, substantia nigra were considerably lower in both groups compared to the healthy control group ( F values: 6.24-147.61, H values: 8.66-24.43, all P<0.05; post-hoc: LSD- t test, Bonferroni correction, all P<0.01). In the PD-RBD(-) group, contralateral striatal anterior-posterior gradient were negatively correlated with unified PD Rating Scale (UPDRS) score and modified Hoehn-Yahr (mH-Y) stage ( r=-0.35, P=0.048; rs=-0.39, P=0.026). The AUC for distinguishing PD-RBD(+ ) and PD-RBD(-) with a contralateral striatal anterior-posterior gradient was 0.706 (95% CI: 0.562-0.851, P=0.016), with the sensitivity and specificity of 59.4%(19/32) and 16/18, respectively. Conclusions:The decrease of contralateral striatal anterior-posterior gradient of VMAT2 is more obvious in patients with PD-RBD(+ ), and there are differences in brain metabolism between the two groups, suggesting that there may be different neuropathological changes and different pathophysiological mechanisms between PD patients with and without RBD. 18F-DTDZ PET/CT can provide imaging basis for the differential diagnosis of the disease subtypes.

To summarize the nursing experience of an adolescent with early repolarization syndrome who had implanted a subcutaneous implantable cardioverter defibrillator(ICD)and experiences frequent ventricular fibrillation.The main nursing care were as follows:recognizing a ventricular fibrillation attack and emergency care;risk prevention and handling in transportation;prevention of iatrogenic infection and strengthening protective isolation;promoting the recovery of brain function;personalized health education;pre-hospital first aid skills training for families.After careful treatment and nursing,the patient did not recur arrhythmia,and heart function had improved significantly,and the pulmonary inflammation disappeared.No medical-acquired infections occurred during hospitalization.The patient's motor ability and language expression ability recovered to the baseline,and was discharged on the 20th day of admission.A month after discharge,the patient recovered well and could study and live normally.

Objective:To investigate the relationship between C3, C4, Th1/Th2 levels and the Myasthenia Gravis Daily Living Scale (MG-ADL) score in patients with myasthenia gravis (MG) and its efficacy in predicting the transition of ocular muscle type to systemic type.Methods:A retrospective study of 94 patients with ophthalmic MG admitted to Haikou People's Hospital from April 2017 to April 2020 was conducted. According to whether they had converted to systemic MG within 6 months, they were divided into transformation group ( n=35) and non-transformation group ( n=59). The levels of C3, C4 and Th1/Th2, as well as the score of MG-ADL and Quantitative Myasthenia Gravis (QMG) were compared between the two groups before and 1 and 3 months after treatment. The correlation between C3, C4 and Th1/Th2 levels and MG-ADL and OMG scores, as well as the related influencing factors of the transformation from ocular muscle type to systemic type was analyzed. The efficiency of each index in predicting the transformation from ocular muscle type to systemic type was analyzed. Results:At 1 and 3 months after treatment, the C3 and C4 in both groups were significantly higher than before treatment, and Th1/Th2 was significantly lower than before treatment; the C3 and C4 in the non-transformation group were higher than that in the transformation group, while Th1/Th2 was lower than that in the transformation group (all P<0.05). The MG-ADL and QMG scores in 2 groups at 1 and 3 months after treatment were significantly lower than those before treatment, and those in the non-transformation group were lower than those in the transformation group (all P<0.05). C3 and C4 levels were negatively correlated with MG-ADL and QMG scores (all P<0.05), while Th1/Th2 levels were positively correlated with MG-ADL and QMG scores (all P<0.05). At 1 and 3 months after treatment, C3, C4 and Th1/Th2 were the influencing factors for the transformation from ocular muscle type to systemic type (all P<0.05). The area under the curve (AUC) of C3, C4 and Th1/Th2 combined to predict the transformation from ocular muscle type to systemic type at 3 months after treatment was 0.939, and the best predictive sensitivity and specificity were 91.43% and 88.14%, respectively. Conclusions:There is a good linear relationship between C3, C4, Th1/Th2 levels and MG-ADL scores in MG patients, and it has a high efficiency in predicting the transition of ocular muscle type to systemic type.