Objective To investigate whether exposure pathways influence the distribution pattern and toxicity of polystyrene nanoplastics(PSNPs)in hepatic cells.Methods Male C57BL/6J wild-type healthy mice aged 6 to 8 weeks old and weighed 18 to 22 g were administered with PSNPs via gavage or tail vein injection.Then,we tracked PSNPs distribution in the major organs of mice via an in vivo imaging system(IVIS).After that,we analyzed the cellular accumulation patterns in hepatic cell subpopulations(hepatocytes and Kupffer cells)using immunofluorescence and transmission electron microscopy(TEM).300 nm PSNPs were administered via gastric gavage or tail vein injection,and 70 nm PSNPs were injected via the portal vein.The cellular localization of PSNPs in the liver was analyzed using immunofluorescence.Subsequently,using AML-12 cells,a normal mouse liver cell line,as the parenchymal hepatocyte model,the uptake of PSNPs in AML-12 cells was analyzed by confocal laser scanning microscope(CLSM).Flow cytometry was performed to observe and quantify PSNPs uptake,and to analyze the underlying endocytosis mechanisms.IVIS was used to analyze PSNPs uptake features in vivo.Finally,using mouse macrophage line RAW264.7 as a Kupffer cell model and AML-12 cells as a parenchymal hepatocyte model,the cell-type-specific toxic effects induced by 100 μg/ml PSNPs were examined through transcriptomics and metabolomics analyses.Results IVIS revealed predominant hepatic accumulation of PSNPs regardless of exposure pathways via intragastric gavage or tail vein injection.Immunofluorescence/TEM demonstrated exposure pathway-dependent cellular distribution:intragastric PSNPs were localized mainly in hepatocytes,while intravenous PSNPs were accumulated in Kupffer cells.Changes in particle size(300 nm vs.70 nm)did not alter the cellular distribution pattern,while 70 nm PSNPs injected via the portal vein accumulated in Kupffer cells,which suggested that the cell-type-specific distribution of PSNPs in the liver was independent of PSNPs size and might be related to the transport of PSNPs in the gastrointestinal tract.Flow cytometry showed that PSNPs uptake by AML-12 was time-dependent and that the underlying endocytosis mechanism involved pathways mediated by clathrin(P<0.000 1),macropinocytosis(P=0.002 6),and lipid rafts(P<0.000 1).Findings on PSNPs distribution in blood revealed that the uptake of PSNPs by hepatocytes exhibited a rate saturation phenomenon.Multi-omics analysis identified distinct toxicity patterns:PSNPs disrupted lipid metabolism and neurotransmitter homeostasis in AML-12 cells and induced inflammation and oxidative stress in Kupffer cells.Conclusion Exposure pathways mediate the hepatic cell-type-specific distribution of PSNPs,thereby altering the downstream toxicological consequences induced by exposure to PSNPs.

AIM:This study aims to investigate the molecular mechanism by which tubeimoside I(TBMS1)inhibits Snail expression in pancreatic cancer cells(PANC-1).METHODS:Human pancreatic cancer PANC-1 cells were cultured in vitro.The inhibitory effect of TBMS1 on PANC-1 cells was assessed using the MTT assay,and the data were analyzed based on the IC50 value of TBMS1.The impact of TBMS1 on the clonal formation ability of PANC-1 cells was evaluated through colony formation assays.The Transwell assay was employed to assess the effect of TBMS1 on the migrato-ry capability of PANC-1 cells.Apoptosis and cell cycle alterations in PANC-1 cells were analyzed using acridine orange staining and flow cytometry.The expression of Snail protein in pancreatic cancer and its relationship with survival of the patients were analyzed using the GEPIA database and Kaplan-Meier Plotter data.Immunofluorescence staining was con-ducted to investigate the effect of TBMS1 on Snail expression,while Western blot was used to evaluate the expression of poly(ADP-ribose)polymerase(PARP),E-cadherin and Snail in the cells.The ubiquitination of Snail protein was mea-sured using immunoprecipitation techniques.RESULTS:As the concentration of TBMS1 increased,the survival rate and number of clones formed by PANC-1 cells progressively decreased,leading to apoptosis,cleavage of PARP,and cell cycle arrest in the G1 phase.There was also a reduction in the proportion of cells in the S phase and a decrease in cell migration ability.The expression of Snail protein,a critical factor in cell migration,was inhibited,while E-cadherin protein levels were increased.Treatment with the proteasome inhibitor MG132 was able to reverse the suppression of Snail protein ex-pression caused by TBMS1.Immunoprecipitation results indicated that TBMS1 enhances the ubiquitination and subse-quent degradation of Snail protein.CONCLUSION:TBMS1 effectively inhibits the malignant progression of pancreatic cancer cells by promoting the ubiquitination and degradation of Snail protein in PANC-1 cells.

AIM:This study aims to investigate the molecular mechanism by which tubeimoside I(TBMS1)inhibits Snail expression in pancreatic cancer cells(PANC-1).METHODS:Human pancreatic cancer PANC-1 cells were cultured in vitro.The inhibitory effect of TBMS1 on PANC-1 cells was assessed using the MTT assay,and the data were analyzed based on the IC50 value of TBMS1.The impact of TBMS1 on the clonal formation ability of PANC-1 cells was evaluated through colony formation assays.The Transwell assay was employed to assess the effect of TBMS1 on the migrato-ry capability of PANC-1 cells.Apoptosis and cell cycle alterations in PANC-1 cells were analyzed using acridine orange staining and flow cytometry.The expression of Snail protein in pancreatic cancer and its relationship with survival of the patients were analyzed using the GEPIA database and Kaplan-Meier Plotter data.Immunofluorescence staining was con-ducted to investigate the effect of TBMS1 on Snail expression,while Western blot was used to evaluate the expression of poly(ADP-ribose)polymerase(PARP),E-cadherin and Snail in the cells.The ubiquitination of Snail protein was mea-sured using immunoprecipitation techniques.RESULTS:As the concentration of TBMS1 increased,the survival rate and number of clones formed by PANC-1 cells progressively decreased,leading to apoptosis,cleavage of PARP,and cell cycle arrest in the G1 phase.There was also a reduction in the proportion of cells in the S phase and a decrease in cell migration ability.The expression of Snail protein,a critical factor in cell migration,was inhibited,while E-cadherin protein levels were increased.Treatment with the proteasome inhibitor MG132 was able to reverse the suppression of Snail protein ex-pression caused by TBMS1.Immunoprecipitation results indicated that TBMS1 enhances the ubiquitination and subse-quent degradation of Snail protein.CONCLUSION:TBMS1 effectively inhibits the malignant progression of pancreatic cancer cells by promoting the ubiquitination and degradation of Snail protein in PANC-1 cells.

Background Heavy metals may play an important role in environmental risk factors associated disorders of activities of daily living (ADL) in older adults. Objective To investigate the associations between plasma levels of six heavy metals (zinc, arsenic, cadmium, lead, manganese, and copper) and ADL disorders in older adults. Methods A cross-sectional survey was conducted from 2018 to 2019 among 1412 rural elderly people in Gongcheng Yao Autonomous County, Guangxi Zhuang Autonomous Region. The plasma metal concentrations were detected by inductively coupled plasma mass spectrometry (ICP-MS), and subsequently classified into three groups (T1-T3) based on tertiles, with the T1 group as the reference. The samples were assessed for ADL disorders using the ADL scale. Logistic regression and restricted cubic spline model (RCS) were used to estimate the odds ratio (OR) and 95% confidence interval (CI) to assess the associations between heavy metals and the prevalence of reporting ADL disorders. Results The mean age of the study population was (68.52 ± 5.92) years, 825 (58.4%) female and 587 (41.6%) male. Of these, 372 (26.34%) subjects reported ADL disorders, and most of them were female (74.30%). The results of logistic regression showed that the participants in the cadmium T2 group (0.15-0.25 µg·L⁻¹) had a higher risk of ADL disorders compared to the T1 group (≤0.15 μg·L⁻¹) (OR=1.552, 95%CI: 1.086, 2.134). After stratification by sex, the relative risk of ADL disorders was lower in the plasma copper T3 group (>1019.58 µg·L⁻¹) compared to the T1 group (≤868.12 μg·L⁻¹) in men (OR=0.481, 95%CI: 0.232, 0.998). The relative risk of ADL disorders was higher in the plasma cadmium T2 group (0.15-0.25 µg·L⁻¹) compared to the T1 group (≤0.15 μg·L⁻¹) in women (OR=1.758, 95%CI: 1.182, 2.616). The RCS results showed that the risk of ADL disorders in men was nonlinearly associated with copper (P_nonlinear=0.011, P_overall<0.05). Conclusion High levels of cadmium are positively associated with the risk of reporting ADL disorders, while high levels of copper are negatively associated with the risk of reporting ADL disorders in men.

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 investigate the effect of tocilizumab (TCZ) on ventricular arrhythmias (VAs) after myocardial infarction (MI) in Sprague-Dawley rats and explore its potential mechanism.Methods:The random number table method was used to divide 32 adult male Sprague-Dawley rats into 4 groups: Sham group, TCZ group, MI group and MI+TCZ group, with 8 rats in each group. The MI model was established by ligation of the left anterior descending branch of the coronary artery in the MI and MI+TCZ groups, and only sutured without ligation in the Sham and TCZ groups. TCZ was injected into the left superior cervical ganglion (SCG) of rats in the TCZ and MI+TCZ groups after successful modeling or sham operation, and the same amount of normal saline was injected in the Sham and MI groups. 24 h after successful modeling, ECG of rats in each group was recorded, heart rate variability (HRV, including low frequency power (LF), high frequency power (HF), LF/HF ratio), QT interval, QTc interval were calculated, and left ventricular effective refractory period (ERP) and VA inducibility were measured. Myocardial infarct size and tissue changes were observed with triphenyl tetrazolium chloride staining and HE staining. Real-time PCR analysis was used to detect the messager RNA (mRNA) expression of interleukin-6 (IL-6) and signal transducer and activator of transcription (STAT) 3 in SCG and potassium voltage-gated channel subfamily D member 2 (Kcnd2) in myocardial infarction periphery. The expression of c-fos in SCG was detected by immunofluorescence staining.Results:Compared with Sham group and MI+TCZ group, rats in MI group had higher LF and LF/HF ratio, longer QT interval and QTc interval, more VAs induced, lower HF and shorter ERP ( P all<0.05). Triphenyl tetrazolium chloride staining and HE staining showed that rats in the Sham and TCZ groups had normal myocardial tissue structure, those in the MI group had severe myocardial injury, and those in the MI+TCZ group had less myocardial injury than those in the MI group. Real-ime PCR analysis showed that compared with Sham group and MI+TCZ group, mRNA expression levels of IL-6 and STAT3 in SCG of rats in MI group were higher, and mRNA expression level of myocardial Kcnd2 was lower ( P all<0.05). Immunofluorescence staining showed that the content of c-fos in SCG of rats in MI group was higher than that of Sham group and MI+TCZ group ( P all<0.05). Conclusions:TCZ may reduce neural activity of the SCG after MI by inhibiting the IL-6/STAT3 signaling pathway, thereby alleviating myocardial injury and inhibiting VAs.

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.

Objective To investigate the effect of renal denervation on cardiac function and sympa-thetic nerve remodeling in spontaneously hypertensive rats(SHR).Methods Sixteen SPF male SHRs were randomly and equally divided into renal denervation(RDN)group and sham operation group,and 8 SPF Wistar-Kyoto(WKY)rats served as the normotensive control group.Blood pressure and heart rate of each group were non-invasively recorded every two weeks.In 8 weeks after the RDN procedure,mean arterial pressure was recorded after carotid artery catheterization,and heart rate variability,including low-frequency power(LF),high-frequency power(HF),and LF/HF ratio,was detected with electrocardiography.Cardiac function was assessed using echocar-diography(ECG),with indicators such as left ventricular ejection fraction(LVEF),left ventricu-lar fractional shortening(LVFS),left ventricular end-diastolic diameter(LVEDD),and left ven-tricular end-systolic diameter(LVESD).HE staining was used to assess myocardial injury,while immunohistochemical staining was employed to detect myocardial tyrosine hydroxylase(TH)density.RT-PCR was performed to measure the mRNA expression of IL-1β and TNF-α.Results The SBP before surgery,as well as at 2,4,6,and 8 weeks post-surgery,and the mean arterial pressure(MAP)were significantly higher in the sham operation and RDN groups than the control group,while,the MAP in the RDN group at 2,4,6,and 8 weeks post-surgery was significantly lower than that in the sham operation group(P＜0.05).Compared with the control group,the sham operation group showed significant increases in LVEDD,LVESD,heart weight-to-body weight ratio,myocardial TH,LF and LF/HF ratio,and myocardial IL-1β and TNF-α expression,while HF,LVEF,and LVFS were significantly reduced(P＜0.05,P＜0.01).Compared with the sham operation group,the RDN group showed significant increases in LVEF[(83.32±2.34)％vs(75.33±2.46)％,P＜0.05],LVFS[(45.57±2.42)％vs(38.42±1.64)％,P＜0.05],and HF(66.73±2.33 vs 60.23±1.54,P＜0.01).Meanwhile,LVESD[(3.56±0.34)mm vs(4.33±0.36)mm,P＜0.05],LVEDD[(6.43±0.38)mm vs(7.23±0.42)mm,P＜0.05],heart weight-to-body weight ratio(3.52±0.16 vs 3.82±0.22,P＜0.05),myocardial TH(0.15±0.01 vs 0.19±0.02,P＜0.05),LF[(15.55±1.08)％vs(19.91±1.79)％,P＜0.01]and LF/HF ratio(0.23±0.01 vs 0.33±0.03,P＜0.01),and the mRNA levels of myocardial IL-1β(2.47±0.15 vs 3.12±0.18,P＜0.05)and TNF-a(3.15±0.21 vs 3.79±0.17,P＜0.05)were significantly reduced.Conclusion RDN can reduce blood pressure in SHR,improve cardiac structure and function,and inhibit cardi-ac sympathetic nerve remodeling,which may be through alleviating neuroinflammation.

Insufficient myocardial perfusion is a common pathological feature in the early stage of various heart diseases,and quantitative evaluation of myocardial perfusion is of great significance for diagnosing and treating diseases and evaluating prognosis.Cardiac MR(CMR)perfusion imaging is non radiative,having high spatial resolution and being sensitive to local myocardial ischemia.Numerical simulation technology combined with computational biomechanics could simulate the process of myocardial blood flow perfusion.The research progresses of CMR and numerical simulation for quantitative evaluation of myocardial perfusion were reviewed in this article.