Objective:To investigate the effects of microRNA-325-3p(miR-325-3p)over-expression on the invasion and migration of colon cancer cells,and to clarify their mechanisms.Methods:The tumor tissue and the corresponding adjacent normal tissue of 25 patients clearly diagnosed with colon cancer were collected.The expression levels of miR-325-3p and relevant evolutionary and lymphoid interest domain containing protein 1(PRELID1)mRNA were detected by real-time fluorescence quantitative PCR(RT-qPCR)method.The expression levels of miR-325-3p and PRELID1 mRNA and the expression levels of PRELID1 protein in human normal colon cells NCM460 and colon cancer cells SW480,HCT116 and HT-29 were detected by RT-qPCR and Western blotting methods.The SW480 cells were divided into control group,NC-mimics group,miR-325-3p mimics group,miR-325-3p mimics+oe-NC group and miR-325-3p mimics+oe-PRELID1 group.The invasion and migration abilities of cells in various groups were detected by Transwell chamber assay and scratch healing assay,respectively.The expression levels of E-Cadherin,N-Cadherin and Vimentin in the epithelial-mesenchymal transition(EMT)of cells in various groups were detected by Western blotting method.The downstream target genes of miR-325-3p were predicted using the Targetscan bioinformatics website,and the targeted regulatory relationship between miR-325-3p and PRELID1 was verified by the dual-luciferase assay.Results:Compared with adjacent normal tissue,the expression level of miR-325-3p in cancer tissue was significantly decreased(P＜0.05),while the expression level of PRELID1 mRNA was significantly increased(P＜0.05).The expression levels of miR-325-3p and PRELID1 mRNA was negatively correlated in colon cancer tissue(r＜0,R2=0.392,P＜0.001).Compared with NCM460 cells,the expression levels of miR-325-3p in SW480,HCT116 and HT-29 cells were significantly decreased(P＜0.05),and the expression levels of PRELID1 mRNA were significantly increased(P＜0.05).Compared with control group and NC-mimics group,the number of invasive SW480 cells in miR-325-3p mimics group was significantly reduced(P＜0.05),the rate of scratch healing was decreased(P＜0.05),and the expression level of E-Cadherin protein was increased,while the expression levels of N-Cadherin and Vimentin proteins were decreased(P＜0.05).The dual-luciferase assay confirmed that PRELID1 was the direct target of miR-325-3p.Compared with miR-325-3p mimics+oe-NC group,the number of invasive SW480 cells in miR-325-3p mimics+oe-PRELID1 group was significantly increased(P＜0.05),the rate of scratch healing was increased(P＜0.05),the expression level of E-Cadherin protein was decreased,and the expression levels of N-Cadherin and Vimentin proteins were increased(P＜0.05).Conclusion:Over-expression of miR-325-3p can inhibit EMT process by down-regulating PRELID1 expression,thereby inhibiting SW480 cell invasion and migration.

In June 2018,sows at a pig farm in Jiamusi,Heilongjiang Province,suffered a large num-ber of miscarriages,and subsequently weaned piglets at the farm began to show persistent high fe-ver symptoms at around 35 days of age,with some pigs having a fever of more than 41.5 ℃.In or-der to determine the cause of this outbreak,63 clinical samples from this farm were tested.The re-sults showed that 60 out of 63 samples were positive for the porcine reproductive and respiratory syndrome virus(PRRSV)antigen.Subsequently,PRRSV antigen-positive plasmid was transfected into African green monkey embryonic kidney cells(Marc-145),and after three generations of blind transmission,indirect immunofluorescence assay(IFA)was performed.The results showed that one PRRSV strain,named HLJ38,was successfully isolated.Then the whole genome of HLJ38 strain was sequenced and then analyzed in detail by bioinformatics software.Sequence analysis showed that there were three deletions of 131 amino acids(323-433 aa,483 aa and 504-522 aa)in the derived sequence of Nsp2 gene of HLJ38 strain,which was consistent with the molecular ge-netic marker of NADC30 PRRSV.The phylogenetic tree analysis showed that HLJ38 and NADC30 PRRSV in GenBank belong to lineage 1 subgroup,and the nucleotide homology of HLJ38 and NADC30 PRRSV in GenBank was only 85.2％and 84.6％.Recombinant analysis showed that HLJ38 was a recombinant NADC30-like PRRSV,and the recombinant gene fragments were de-rived from multiple strains,among which the fragment of 1-201 nt was provided by VR2332 strain and fragment of 6 641-8 061 nt derived from the HP-PRRSV strain.In summary,the re-sults showed that the outbreak in this pig farm may be caused by the recombination of PRRSV strains among different lineages,and the recombinant circulating strains not only have certain pathogenicity but also suggest that the existing commercial vaccines provide limited cross-protec-tion against them.Recombination between different lineages increases the genetic diversity of PRRSV and aggravates the difficulty of prevention and control of PRRS in pig farms.Therefore,it is necessary to continuously monitor the epidemic dynamics of PRRSV in pig farms and take effec-tive measures in time to curb the spread of PRRS.

BACKGROUND:Degeneration of nucleus pulposus cells is a key component of intervertebral disc degeneration.Ferroptosis,a novel form of programmed cell death,is closely associated with the onset and progression of intervertebral disc degeneration;however,its precise mechanisms remain unclear.OBJECTIVE:To establish an oxidative stress model in vitro by inducing ferroptosis in nucleus pulposus cells using tert-butyl hydroperoxide and to investigate the mechanisms of tert-butyl hydroperoxide-induced ferroptosis in nucleus pulposus cells,thereby elucidating the role of ferroptosis in the pathogenesis of intervertebral disc degeneration.METHODS:Nucleus pulposus cells were treated with varying concentrations of tert-butyl hydroperoxide(0,25,50,100,and 200 μmol/L),and cell morphology and viability were assessed using fluorescence microscopy and the cell counting kit-8 assay.Interventions with 100 μmol/L tert-butyl hydroperoxide,10 μmol/L RSL3,or dimethylsulfoxide were applied to nucleus pulposus cells,and cell proliferation was evaluated using the EdU assay.The expression levels of ferroptosis-related proteins(glutathione peroxidase 4,ferritin heavy chain 1,PTGS2,and ACSL4)and intervertebral disc degeneration marker proteins(matrix metalloproteinase 13 and Col2A)were analyzed via western blot and immunofluorescence.Additionally,reactive oxygen species and lipid peroxidation levels were quantified using the reactive oxygen species detection kit and C11-BODIPY probe.Mitochondrial morphological changes were observed under transmission electron microscopy.RESULTS AND CONCLUSION:(1)tert-Butyl hydroperoxide treatment significantly reduced the viability and proliferation of nucleus pulposus cells.(2)tert-Butyl hydroperoxide induced typical ferroptosis-related morphological changes in nucleus pulposus cells.(3)tert-Butyl hydroperoxide exposure led to a decrease in the expression of ferroptosis-suppressing proteins glutathione peroxidase 4 and ferritin heavy chain 1,while increasing the expression of ferroptosis-promoting factors ACSL4 and PTGS2.(4)tert-Butyl hydroperoxide elevated intracellular reactive oxygen species production and lipid peroxidation levels in nucleus pulposus cells.(5)Transmission electron microscopy revealed ferroptosis-specific mitochondrial changes in nucleus pulposus cells treated with tert-butyl hydroperoxide,including contraction,reduced cristae,and increased membrane density.(6)tert-Butyl hydroperoxide treatment also resulted in the increased expression of matrix metalloproteinase 13 and decreased expression of Col2A in nucleus pulposus cells.In conclusion,tert-butyl hydroperoxide induces ferroptosis in nucleus pulposus cells,contributing to the development of intervertebral disc degeneration.This process may represent a key pathological mechanism in intervertebral disc degeneration and offers potential targets for developing novel therapeutic strategies.

In June 2018,sows at a pig farm in Jiamusi,Heilongjiang Province,suffered a large num-ber of miscarriages,and subsequently weaned piglets at the farm began to show persistent high fe-ver symptoms at around 35 days of age,with some pigs having a fever of more than 41.5 ℃.In or-der to determine the cause of this outbreak,63 clinical samples from this farm were tested.The re-sults showed that 60 out of 63 samples were positive for the porcine reproductive and respiratory syndrome virus(PRRSV)antigen.Subsequently,PRRSV antigen-positive plasmid was transfected into African green monkey embryonic kidney cells(Marc-145),and after three generations of blind transmission,indirect immunofluorescence assay(IFA)was performed.The results showed that one PRRSV strain,named HLJ38,was successfully isolated.Then the whole genome of HLJ38 strain was sequenced and then analyzed in detail by bioinformatics software.Sequence analysis showed that there were three deletions of 131 amino acids(323-433 aa,483 aa and 504-522 aa)in the derived sequence of Nsp2 gene of HLJ38 strain,which was consistent with the molecular ge-netic marker of NADC30 PRRSV.The phylogenetic tree analysis showed that HLJ38 and NADC30 PRRSV in GenBank belong to lineage 1 subgroup,and the nucleotide homology of HLJ38 and NADC30 PRRSV in GenBank was only 85.2％and 84.6％.Recombinant analysis showed that HLJ38 was a recombinant NADC30-like PRRSV,and the recombinant gene fragments were de-rived from multiple strains,among which the fragment of 1-201 nt was provided by VR2332 strain and fragment of 6 641-8 061 nt derived from the HP-PRRSV strain.In summary,the re-sults showed that the outbreak in this pig farm may be caused by the recombination of PRRSV strains among different lineages,and the recombinant circulating strains not only have certain pathogenicity but also suggest that the existing commercial vaccines provide limited cross-protec-tion against them.Recombination between different lineages increases the genetic diversity of PRRSV and aggravates the difficulty of prevention and control of PRRS in pig farms.Therefore,it is necessary to continuously monitor the epidemic dynamics of PRRSV in pig farms and take effec-tive measures in time to curb the spread of PRRS.

BACKGROUND:Degeneration of nucleus pulposus cells is a key component of intervertebral disc degeneration.Ferroptosis,a novel form of programmed cell death,is closely associated with the onset and progression of intervertebral disc degeneration;however,its precise mechanisms remain unclear.OBJECTIVE:To establish an oxidative stress model in vitro by inducing ferroptosis in nucleus pulposus cells using tert-butyl hydroperoxide and to investigate the mechanisms of tert-butyl hydroperoxide-induced ferroptosis in nucleus pulposus cells,thereby elucidating the role of ferroptosis in the pathogenesis of intervertebral disc degeneration.METHODS:Nucleus pulposus cells were treated with varying concentrations of tert-butyl hydroperoxide(0,25,50,100,and 200 μmol/L),and cell morphology and viability were assessed using fluorescence microscopy and the cell counting kit-8 assay.Interventions with 100 μmol/L tert-butyl hydroperoxide,10 μmol/L RSL3,or dimethylsulfoxide were applied to nucleus pulposus cells,and cell proliferation was evaluated using the EdU assay.The expression levels of ferroptosis-related proteins(glutathione peroxidase 4,ferritin heavy chain 1,PTGS2,and ACSL4)and intervertebral disc degeneration marker proteins(matrix metalloproteinase 13 and Col2A)were analyzed via western blot and immunofluorescence.Additionally,reactive oxygen species and lipid peroxidation levels were quantified using the reactive oxygen species detection kit and C11-BODIPY probe.Mitochondrial morphological changes were observed under transmission electron microscopy.RESULTS AND CONCLUSION:(1)tert-Butyl hydroperoxide treatment significantly reduced the viability and proliferation of nucleus pulposus cells.(2)tert-Butyl hydroperoxide induced typical ferroptosis-related morphological changes in nucleus pulposus cells.(3)tert-Butyl hydroperoxide exposure led to a decrease in the expression of ferroptosis-suppressing proteins glutathione peroxidase 4 and ferritin heavy chain 1,while increasing the expression of ferroptosis-promoting factors ACSL4 and PTGS2.(4)tert-Butyl hydroperoxide elevated intracellular reactive oxygen species production and lipid peroxidation levels in nucleus pulposus cells.(5)Transmission electron microscopy revealed ferroptosis-specific mitochondrial changes in nucleus pulposus cells treated with tert-butyl hydroperoxide,including contraction,reduced cristae,and increased membrane density.(6)tert-Butyl hydroperoxide treatment also resulted in the increased expression of matrix metalloproteinase 13 and decreased expression of Col2A in nucleus pulposus cells.In conclusion,tert-butyl hydroperoxide induces ferroptosis in nucleus pulposus cells,contributing to the development of intervertebral disc degeneration.This process may represent a key pathological mechanism in intervertebral disc degeneration and offers potential targets for developing novel therapeutic strategies.

Sarcomatoid carcinoma of the renal pelvis accounts for a very low percentage of malignant tumors in the renal pelvis and has a poor prognosis. This article reported a patient with sarcomatoid carcinoma of the renal pelvis. The patient presented with macroscopic hematuria as the first symptom, and CT suggested left renal occupancy, unilateral nephrectomy was performed, and pathology suggested sarcomatoid carcinoma of the renal pelvis. Three weeks after surgery, a follow-up CT showed tumor recurrence. Programmed death 1(PD-1)inhibitor was given once every 3 weeks. Repeated CT examination after 24 weeks of continuous treatment suggested that the recurrent tumor disappeared. The patients was followed-up for 42 months without tumor recurrence or metastasis.

Objective:To construct a health education program for enterostomy patients based on the guideline of Facilitating Client Centered Learning. Methods:From July to October 2022, based on the adaptability survey and literature research of the previous guidelines, the first draft of the health education program for enterostomy patients was formed. A total of 13 experts were selected using purposive sampling method, and two rounds of expert consultation were conducted through the Delphi method on the first draft of the program to build the final health education program for enterostomy patients. We calculated the authority, enthusiasm, coordination, and concentration of experts.Results:The health education program for enterostomy patients included 3 first-level indicators, 9 second-level indicators, and 31 third-level indicators. The effective response rates of the first and second rounds of consultation questionnaires were both 100.0%, and the authority coefficients of experts were both greater than 0.7. In the second round of consultation, the Kendal coordination coefficients of the importance of the first, second and third level indicators were 0.231, 0.154 and 0.182 ( P<0.05), and the Kendal coordination coefficients of the feasibility of the first, second and third level indicators were 0.216, 0.154 and 0.129 ( P<0.05), with coefficients of variation < 0.25. Conclusions:The health education program for enterostomy patients based on the guideline of Facilitating Client Centered Learning is practical and scientific, and can provide guidance for clinical practice.

Objective To evaluate the effect of over-expression of phosphatase and tensin homolog does on chromosome ten (PTEN) in podocytes on kidney under high fat diet (HFD) in vivo and clarify the mechanism how PTEN regulates scavenger receptor A (SR-A) expression exposed to oxidized low density lipoprotein (ox-LDL) in podocytes in vitro.Methods The podocyte-specific PTEN knockin (PPKI) mice were fed with HFD to establish mouse model of lipid-induced renal injury.Mice were divided into four groups:ND+Ctrl group,ND+PPKI group,HFD+Ctrl group and HFD+PPKI group.After 24 weeks of dietary intervention,all mice were tested for clinical and biochemical parameters,including serum creatinine (Scr) as well as urine albumin excretion rate (UAER);renal lipid content was measured by oil red O staining and cholesterol quantitative analysis;the pathological changes of glomeruli were observed by PAS staining and electron microscope.Podocyte injury was induced by ox-LDL in vitro.Western blotting was used to detect the changes of SR-A expression induced by ox-LDL after YAP-siRNA interfering (si-YAP),as well as YAP phosphorylation induced by ox-LDL after interfering by PTEN-siRNA (si-PTEN) and PTEN phosphatase inhibitor (Bpv-PTEN),and overexpressing by recombinant adenovirus (ad-PTEN).Results Compared with ND+Ctrl group,HFD+ Ctrl group significantly aggravated the levels of Scr and UAER,the expression of SR-A in podocytes,renal lipid content,mesangial matrix expansion,effacement of podocyte foot processes,and incrassation of glomerular basement membrane (all P ＜ 0.05).Conversely,compared with HFD+Ctrl group,HFD+ PPKI group obviously alleviated the above lipid-induced renal damage (all P ＜ 0.05).In vitro,the expression of SR-A in podocytes was up-regulated when stimulated with ox-LDL (P ＜ 0.05),and the knockout of YAP significantly down-regulated the expression of SR-A induced by ox-LDL (P ＜ 0.05).Exposed to ox-LDL,the expression of p-YAP increased in podocytes (P ＜ 0.05);over-expression of PTEN inhibited p-YAP up-regulation induced by ox-LDL (P ＜ 0.05),while either knockdown of PTEN or inhibition of PTEN phosphatase activity displayed opposite effect (all P ＜ 0.05).Conclusions Over-expression of PTEN in podocytes protected the kidney against damage from HFD in vivo and PTEN might suppress SR-A mediated lipid uptake via dephosphorylating p-YAP to prevent podocyte injury from ox-LDL.

Objective: To explore the role of ROCK1 in oxidized low-density lipoprotein (ox-LDL) induced podocyte injury and its possible mechanism. Methods: The conditionally immortalized mouse podocyte cells were cultured in vitro and exposed to 20 μg/ml ox-LDL for 24 h. Western blotting was used to analyze the expression level of p-MYPT, nephrin, LC3-Ⅱ, p62, p-ULK1 in groups of control, ox-LDL, ROCK1 siRNA with ox-LDL, wtROCK1 with ox-LDL. Podocytes were incubated with DiI labeled ox-LDL for 4 h and fluorescence microscope was used to analyze lipid distribution. Results: Compared with control group, ox-LDL increased cell cholesterol accumulation, activated ROCK along with decreased nephrin, LC3-Ⅱ(P<0.05), and increased p62, and p-ULK1 expression (P<0.05). Over-expression of ROCK1 significantly decreased the expression of nephrin and LC3-Ⅱ, but up-regulated the levels of p62, p-ULK1 and cell cholesterol accumulation in ox-LDL stimulated podocytes (P<0.05). In contrast, Inhibition of ROCK1 protected podocyte by improved lipophagy. Conclusion ROCK1 mediated disfunction of lipophagy contributes to the ox-LDL induced podocyte injury.

Objective To compare the diuretic efficacy of torasemide as a 2-hour continuous infusion and as a bolus injection of equal dose in patients with nephrotic syndrome,and to investigate a preferable administration mode of torasemide for these patients.Methods Twenty-three hospitalized patients were randomized to receive torasemide 20 mg or 40 mg per day by either 2-hour intravenous infusion or bolus injection,and interchanged after 48 hours of washout.Results Patients received torasemide by 2-hour intravenous infusion exhibited significantly higher daily urinary volume,chloride excretion,sodium excretion and fractional excretion of sodium (FENa) within 24 hours than those by bolus injection (P ＜ 0.05).Significantly lower bound-state torasemide excretion,higher ratio of urinary volume to torasemide excretion and a markedly larger area under the curve in the plasma concentrationtime profiles were also observed in the infusion group (P ＜ 0.05).Conclusion 2-hour continuous infusion delivers a better diuretic effect compared with a bolus injection of equal dose of torasemide in patients with nephrotic syndrome.