This study explored the effects and underlying mechanism of Raf kinase inhibitory protein(RKIP)on apoptosis in mast cells sensitized by Echinococcus granulosus cyst fluid.Bone marrow-derived mast cells(BMMCs)were isolated and cultured from RKIP knockout(KO)and wild-type(WT)C57BL/6 mice.Cells were divided into control and sensitized groups.The sensitized group was incubated for 24 h in RPMI1640 medium containing 10%serum from mice infected with E.granulosus,then activated for 3 h or 6 h with E.granulosus cyst fluid.The control group was incubated for 24 h in RPMI1640 medium,and then received an equal vol-ume of PBS.Cells and supernatants were collected for analysis.Flow cytometry was used to detect the expression of CD117 and FcεRⅠα on BMMCs.The levels of β-hexosaminidase,IL-4,and TNF-α in the supernatant were quantified with ELISA.Western blot analy-sis was used to assess expression changes in RKIP,apoptosis-related proteins,and pathway proteins in BMMC before and after sensi-tization.Flow cytometry analysis revealed that after 4 weeks of induction,the CD117 and FcεRⅠα double-positivity rates on both WT and KO BMMC exceeded 90%.ELISA indicated that the E.granulosus cyst fluid resulted in significantly greater β-hexosaminidase re-lease(F=16.88,P<0.05),and levels of IL-4(F=16.51,P<0.05)and TNF-α(F=9.78,P<0.05)in the KO sensitized group than the WT sensitized group.With respect to the WT control group,the WT sensitized group showed significantly down-regulated pro-tein expression levels of RKIP(F=8.20,P<0.05)and Bcl-2(F=101.40,P<0.01)after 3 h,but significantly up-regulated levels of p-PI3K(F=8.04,P<0.05),p-Akt(F=32.52,P<0.01),p-P65(F=13.29,P<0.05),and cleaved-caspase-3(F=46.34,P<0.01).With respect to the WT sensitized group,the KO sensitized group showed significantly up-regulated protein expression of p-PI3K(F=8.45,P<0.05),p-Akt(F=8.58,P<0.05),p-P65(F=11.02,P<0.05),and Bcl-2(F=84.50,P<0.001)after 3 h,but significantly down-regulated expression of cleaved-caspase-3(F=15.66,P<0.05).In conclusion,RKIP may inhibit the PI3K/Akt/NF-κB pathway,thereby inducing apoptosis in mast cells sensitized by E.granulosus cyst fluid.This process may help ease aller-gic reactions caused by mast cells in echinococcosis,thus offering a promising new approach for preventing and treating such reactions.

Objective:To explore the effectiveness and safety of different anticoagulation strategies during continuous blood purification (CBP) treatment,providing a reference for anticoagulation strategies in critically ill children undergoing CBP.Methods:A retrospective study was conducted,including children admitted to the PICU of Children's Hospital of Fudan University from January 2019 to December 2024.According to the anticoagulation methods used during CBP treatment,patients were divided into the sodium citrate group and the heparin group.CBP was performed using continuous venovenous hemofiltration or continuous venovenous hemodialysis filtration mode,with a blood flow rate of 3-5 mL/(kg·min),replacement fluid rate of 30-50 mL/(kg·h),and dialysis fluid rate of 20-30 mL/(kg·h).The filter lifespan,28-day all-cause mortality,total length of hospital stay,PICU stay duration,adverse events,and associated costs were compared between the two groups.Results:A total of 221 children were included (105 in the sodium citrate group and 116 in the heparin group),with a cumulative use of 666 filters (284 in the sodium citrate group and 382 in the heparin group).(1) There were no statistically significant differences in general data,including age,sex ratio,underlying diseases,the ratio and duration of invasive mechanical ventilation,vasopressor scores at baseline,and indications for CBP between the two groups (all P>0.05).(2) The filter lifespan was 20(14,32) hours for the sodium citrate group and 21(13,35) hours for the heparin group,with no statistically significant difference between the two groups ( P>0.05); the proportion of accidental downstroke was 2.8% and 6.5%,respectively,with a statistically significant difference ( P=0.029); among the 221 children,86 died,with 38 deaths (35.2%) in the sodium citrate group and 49 deaths (38.9%) in the heparin group,showing no statistically significant difference.(3) The sodium citrate group had a higher incidence of metabolic alkalosis,hypocalcemia,and sodium citrate accumulation (44.4% vs. 1.6%,32.7% vs 9.4%,7.7% vs 0,all P<0.01); the heparin group had a greater proportion of bleeding (6.0% vs. 2.9%) and was more likely to develop heparin-induced thrombocytopenia (10.2% vs. 0, P<0.01).(4) The total hospitalization costs for the sodium citrate group were significantly higher than for the heparin group (200 327 yuan vs. 152 077 yuan, P=0.05); costs related to the use of anticoagulants and monitoring indicators during CBP treatment were also higher in the sodium citrate group (2 479 yuan vs. 682 yuan, P<0.01). Conclusions:Sodium citrate is a safe and effective anticoagulation method for critically ill children undergoing CBP,which can reduce the risk of filter clotting compared to systemic heparin anticoagulation.

Objective To explore the effect of curcumin(CUR)on oxidative damage of keratinocytes induced by ultraviolet B(UVB)irradiation through Toll-like receptor 4(TLR4)/nuclear factor-kappa B(NF-κB)/nucleotide-binding oligomerization domain-containing protein 3(NLRP3)signaling pathway.Methods Human keratinocytes of HaCaT were cultured normally in vitro,and the keratinocyte oxidative damage model was established by the irradiation of 57 mJ/cm2 UVB.The cells with normal culture were as the control group,the cells treated after modeling were as the UVB group,the cells treated with 5 μmol/L CUR after modeling were as the CUR group,the cells treated with 100 μg/L TLR4 inhibitor of TAK-242 after modeling were as the TAK-242 group,and the cells treated with 5 μmol/L CUR and 100 nmol/L TLR4 activator of lipopolysaccharide(LPS)were as the CUR+LPS group.qRT-PCR was applied to detect the relative expression levels of TLR4,NF-κB,and NLRP3 mRNAs of cells in each group.CCK-8 was applied to detect the cell proliferation in each group.The relative content of reactive oxygen species(ROS),the viabilities of superoxide dismutase(SOD)and catalase(CAT),and the concentrations of glutathione(MDA)and glutathione(GSH)of cells in each group were detected by fluorescence assay according to the kit instruction.ELISA kit was used to detect the expression of inflammatory factors of tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β)of cells in each group.Flow cytometry was applied to detect the cell apoptosis in each group.Western blot was applied to detect the expression of proliferation related protein of proliferating cell nuclear antigen(PCNA),apoptosis related proteins[B-cell lymphoma-2(Bcl-2)and Bcl-2-associated X protein(Bax)],and TLR4/NF-κB/NLRP3 signaling pathway related proteins(TLR4,NF-κB and NLRP3)of cells in each group.Results Compared with the Control group,the cell survival rate,the expression levels of PCNA and Bcl-2 proteins,the viabilities of SOD and CAT,and the GSH concentration in the UVB group decreased,while the apoptosis rate,the level of Bax protein,the relative content of ROS,the concentration of MDA,the levels of TNF-α and IL-1β,and the mRNA and protein levels of TLR4,NF-κB and NLRP3 increased(P<0.05).Compared with the UVB group,the cell survival rate,the expression levels of PCNA and Bcl-2 proteins,the viabilities of SOD and CAT,and the GSH concentration in the TAK-242 group and CUR group increased,while the apoptosis rate,the level of Bax protein,the relative content of ROS,the concentration of MDA,the levels of TNF-α and IL-1β,and the mRNA and protein levels of TLR4,NF-κB and NLRP3 decreased(P<0.05).Compared with the CUR group,the cell survival rate,the expression levels of PCNA and Bcl-2 proteins,the viabilities of SOD and CAT,and the GSH concentration in the CUR+LPS group decreased,while the apoptosis rate,the level of Bax protein,the relative content of ROS,the concentration of MDA,the levels of TNF-α and IL-1β,and the mRNA and protein levels of TLR4,NF-κB and NLRP3 increased(P<0.05).Conclusion CUR can increase the antioxidant stress level of keratinocytes,alleviate inflammatory response,promote cell proliferation,and improve cell damage caused by UVB irradiation,which may be related to the inhibition of TLR4/NF-κB/NLRP3 signaling pathway.

Objective:To investigate the effects of combination therapy with aqueous extract of corn silk(CS)and training on exercise capacity and glycolipid metabolism in mice with metabolic syndrome(MS).Methods:In this study,db/db mice were used as the animal model of MS.The mice were administered aqueous extract of CS via gavage and subjected to different intensities of training for 12 weeks(3 months).The specific experimental design was as follows:24 db/db mice were randomly divided into four groups on average:negative control group(NC),aqueous extract of CS group(CS),aqueous extract of CS+moderate-intensity training group(CS+MT),and CS aqueous extract of CS+high-intensity training group(CS+HT).The maximum running speed,forelimb grip strength,body weight and fasting blood glucose of mice were measured before and after treatment.After the intervention,oral glucose tolerance test(OGTT)and insulin tolerance test(ITT)were conducted to assess glucose metabolism,while serum triglyceride(TG),total cholesterol(TC),high-density lipoprotein cholesterol(HDL-C),and low-density lipoprotein cholesterol(LDL-C)levels were measured to evaluate lipid metabolism.Results:After 3 months of intervention,there were significant differences in the maximum running speed and forelimb grip strength among the four groups(P＜0.05).The maximum running speed and forelimb grip strength of CS group,CS+MT group and CS+HT group were higher than those of NC group(P＜0.05).The CS+MT group exhibited higher forelimb grip strength,and the CS+HT group showed higher maximum running speed and forelimb grip strength compared to the CS group(P＜0.05),while no significant difference was found between the CS+MT and CS+HT groups(P＞0.05).Significant differences in body weight were observed among the four groups after 3 months of intervention(P＜0.05).Specifically,the CS+MT and CS+HT groups exhibited significantly lower body weight compared to both the NC and CS groups(P＜0.05),with the CS+MT group having the lowest body weight(P＜0.05).Fasting blood glucose levels also differed significantly among the groups after 2 and 3 months of intervention(P＜0.05).The CS,CS+MT,and CS+HT groups had lower fasting blood glucose levels compared to the NC group(P＜0.05),with the CS+MT and CS+HT groups showing the lowest levels(P＜0.05).No significant difference was found between the CS+MT and CS+HT groups(P＞0.05).After 3 months of intervention,significant differences in the area under the curve(AUC)of OGTT and ITT were observed among the four groups(P＜0.05).The AUC of OGTT and ITT were significantly lower in the CS,CS+MT,and CS+HT groups compared to the NC group(P＜0.05).The CS+MT and CS+HT groups exhibited the lowest AUC values for both OGTT and ITT(P＜0.05),with the CS+MT group showing the lowest AUC for OGTT(P＜0.05).Significant differences in serum lipid levels were observed among the four groups after 3 months of intervention(P＜0.05).TG,TC,and LDL-C levels were significantly lower,while HDL-C levels were higher in the CS,CS+MT,and CS+HT groups compared to the NC group(P＜0.05).The CS+MT group had the lowest TG levels and the highest HDL-C levels compared to the CS+HT group(P＜0.05),with no significant differences in TC and LDL-C levels between these two groups(P＞0.05).Conclusion:Aqueous extract of CS combined with different intensity training can significantly improve the exercise capacity and glycolipid metabolism of MS mice and reduce body weight,especially CS combined with MT treatment is more effective in improving lipid metabolism.In addition,when combined with HT,aqueous extract of CS can also play an auxiliary role in reducing the side effects of high-intensity exercise and improving the therapeutic effect.

Aim To compare the observation results of atomic force microscopy(AFM)and scanning electron microscopy(SEM),and to summarize the main problems and solutions of AFM in observing biological macromolecules,using the observa-tion subjects of protein samples purified by our research group.Methods The protein samples were diluted to 15 nmol·L-1 with PBS,fixed on glass slides,silicon wafers,and mica sheets,dried,and made into solid-phase observation samples.SEM sam-ples were plated with platinum before observation.The surface structures of proteins were observed using AFM and SEM,sample heights were calculated,and differences in results were com-pared.Results Protein samples with positive charges tended to shift to the right during observation due to the repulsion of the AFM probe;mica sheets could effectively eliminate the positive charge of proteins to avoid sample movement;PBS provided a stable environment for protein samples,but the crystallization of PBS salts interfered with probe operation and imaging clarity;SEM samples needed to be plated with platinum before observa-tion and could not achieve the precision of AFM.Conclusions Both AFM and SEM can directly observe protein structures in vitro,with AFM providing higher precision results;when protein sample stability permits,ultrapure water is preferred as the sol-vent carrier,and volatile liquids such as ethanol can also serve as solvent carriers.The application of AFM offers a new approach for pharmacological studies on interactions between biological macromolecules.

Acupuncture and moxibustion therapy is a kind of treatment and health care method with original advantages of China.With the rapid development of epigenetics and systems biology technology,non-coding RNA(ncRNA)related research has made continuous breakthroughs in the field of acupuncture and moxibustion.This article collected the basic research literature on acupuncture and moxibustion related to ncRNA,and reviewed the research subsystems related to microRNA(miRNA),long chain non coding RNA(lncRNA)and circular RNA(circRNA).NcRNAs are widely involved in the growth,development and reproduction of the organism,as well as in the occurrence and development of various diseases,which fits with the multi-layer,multi-pathway and multi-target action network of acupuncture and moxibustion therapy.Taking ncRNAs as the breakthrough point to explore the mechanism of acupuncture and moxibustion in depth is not only conducive to promoting the exploration of new targets of acupuncture and moxibustion effect,but also can reveal the epigenetic regulation axis of acupuncture and moxibustion effect molecules,and provide ideas and methods for clinical diagnosis and treatment of diseases and evaluation of efficacy.

Objective:To investigate the effect of signals mediated by activated CD28 in promoting survival of multiple myeloma(MM)cells and metabolic fitness and its possible mechanism.Methods:The expression of CD28 on 4 MM cell lines(XG2,XG1,RPMI 8226 and U266)was determined by flow cytometry.Two cell lines with the highest or lowest CD28 expression were selected.The proliferation,cell cycle,migration and apoptosis of MM cells in vitro were determined in medium containing high glucose concentration or CD28 agonist monoclonal antibody with different bioassays.shRNA interference assay was used to knock down the expression of CD28 on U266 cells.Then,the effect of activated CD28 on glucose uptake rate and drug resistance in MM cells were analyzed using fluorescent glucose analogues(2-NBDG).The expression of Glut1/4,HkII and Fasn was determined with real time quantitative PCR.Results:Flow cytometry analysis showed that all the four tested MM cell lines expressed CD28 and U266 cells had the highest positive rate.The results of in vitro experiment showed that CD28 activation could significantly up-regulate the expression of Glut4 and HkII,promote MM cell metabolic remodeling,enhance 2-NBDG/glucose uptake,increase energy metabolism,thereby elevating cell proliferation and migration abilities,leading to an increase in the number of cells in S-and G2-phases.Meanwhile,activated CD28 subsequently up-regulated resistance of MM cells to bortezomib or dexamethasone.Conclusion:MM cells express high levels of CD28 abnormally,and activation of CD28 can promote up-regulation of glucose uptake in MM cells,thereby promoting cell proliferation and enhancing drug resistance.

Acupuncture and moxibustion therapy is a kind of treatment and health care method with original advantages of China.With the rapid development of epigenetics and systems biology technology,non-coding RNA(ncRNA)related research has made continuous breakthroughs in the field of acupuncture and moxibustion.This article collected the basic research literature on acupuncture and moxibustion related to ncRNA,and reviewed the research subsystems related to microRNA(miRNA),long chain non coding RNA(lncRNA)and circular RNA(circRNA).NcRNAs are widely involved in the growth,development and reproduction of the organism,as well as in the occurrence and development of various diseases,which fits with the multi-layer,multi-pathway and multi-target action network of acupuncture and moxibustion therapy.Taking ncRNAs as the breakthrough point to explore the mechanism of acupuncture and moxibustion in depth is not only conducive to promoting the exploration of new targets of acupuncture and moxibustion effect,but also can reveal the epigenetic regulation axis of acupuncture and moxibustion effect molecules,and provide ideas and methods for clinical diagnosis and treatment of diseases and evaluation of efficacy.

AIM To explore the clinical effects of Huayu Ditan Linao Decoction combined with Suhexiang Pills on elderly patients with cerebral hemorrhage in convalescent stage.METHODS One hundred and ten patients were randomly assigned into control group(55 cases)for 1-month intervention of both conventional treatment and rehabilitation training,and observation group(55 cases)for 1-month intervention of Huayu Ditan Linao Decoction,Suhexiang Pills,conventional treatment and rehabilitation training.The changes in clinical effects,TCM syndrome score,NIHSS score,FMA score,BI index,cerebrovascular function indices(dynamic resistance,average flow velocity,peripheral resistance,average flow rate),serum inflammatory indices(IL-6,hs-CPR,TNF-α)and serum oxidative stress indices(SOD,MDA)were detected.RESULTS The observation group demonstrated higher total effective rate than the control group(P＜0.05).After the treatment,the two groups displayed decreased TCM syndrome score,NIHSS score,dynamic resistance,peripheral resistance,serum inflammatory indices,MDA(P＜0.05),and increased FMA score,BI index,average flow velocity,average flow rate,SOD(P＜0.05),especially for the observation group(P＜0.05).CONCLUSION For the elderly patients with cerebral hemorrhage in convalescent stage,Huayu Ditan Linao Decoction combined with Suhexiang Pills can enhance clinical effects,improve limb functions and nerve functions,regulate serum inflammatory and oxidative stress indice,and promote prognosis.

Objective:To explore the effectiveness and safety of different anticoagulation strategies during continuous blood purification (CBP) treatment,providing a reference for anticoagulation strategies in critically ill children undergoing CBP.Methods:A retrospective study was conducted,including children admitted to the PICU of Children's Hospital of Fudan University from January 2019 to December 2024.According to the anticoagulation methods used during CBP treatment,patients were divided into the sodium citrate group and the heparin group.CBP was performed using continuous venovenous hemofiltration or continuous venovenous hemodialysis filtration mode,with a blood flow rate of 3-5 mL/(kg·min),replacement fluid rate of 30-50 mL/(kg·h),and dialysis fluid rate of 20-30 mL/(kg·h).The filter lifespan,28-day all-cause mortality,total length of hospital stay,PICU stay duration,adverse events,and associated costs were compared between the two groups.Results:A total of 221 children were included (105 in the sodium citrate group and 116 in the heparin group),with a cumulative use of 666 filters (284 in the sodium citrate group and 382 in the heparin group).(1) There were no statistically significant differences in general data,including age,sex ratio,underlying diseases,the ratio and duration of invasive mechanical ventilation,vasopressor scores at baseline,and indications for CBP between the two groups (all P>0.05).(2) The filter lifespan was 20(14,32) hours for the sodium citrate group and 21(13,35) hours for the heparin group,with no statistically significant difference between the two groups ( P>0.05); the proportion of accidental downstroke was 2.8% and 6.5%,respectively,with a statistically significant difference ( P=0.029); among the 221 children,86 died,with 38 deaths (35.2%) in the sodium citrate group and 49 deaths (38.9%) in the heparin group,showing no statistically significant difference.(3) The sodium citrate group had a higher incidence of metabolic alkalosis,hypocalcemia,and sodium citrate accumulation (44.4% vs. 1.6%,32.7% vs 9.4%,7.7% vs 0,all P<0.01); the heparin group had a greater proportion of bleeding (6.0% vs. 2.9%) and was more likely to develop heparin-induced thrombocytopenia (10.2% vs. 0, P<0.01).(4) The total hospitalization costs for the sodium citrate group were significantly higher than for the heparin group (200 327 yuan vs. 152 077 yuan, P=0.05); costs related to the use of anticoagulants and monitoring indicators during CBP treatment were also higher in the sodium citrate group (2 479 yuan vs. 682 yuan, P<0.01). Conclusions:Sodium citrate is a safe and effective anticoagulation method for critically ill children undergoing CBP,which can reduce the risk of filter clotting compared to systemic heparin anticoagulation.