Objective To investigate the predictive value of serum homocysteine(HCY)and lipoprotein(a)[Lp(a)]in autologous arteriovenous fistula(AVF)failure in patients undergoing maintenance hemodialy-sis(MHD).Methods A total of 175 patients with MHD admitted to the hospital from January to December 2022 were selected as the observation group,among them,103 patients without AVF failure were selected as observation group 1,and 72 patients with AVF failure were selected as observation group 2.175 healthy indi-viduals who underwent physical examinations at the outpatient examination center of the hospital during the same period were selected as the control group.Enzyme-linked immunosorbent assay was used to detect the levels of HCY and Lp(a).Pearson method was used to analyze the correlation between HCY,Lp(a)with clin-ical indicators.Multivariate Logistic regression analysis was performed to analyze the influencing factors of AVF failure in MHD patients.Receiver operating characteristic(ROC)curve was plotted to analyze the pre-dictive value of serum HCY and Lp(a)for AVF failure in MHD patients.Results Compared with the control group,the serum levels of HCY and Lp(a)in observation group 1 were increased(P<0.05).Compared with the control group and observation group 1,the serum levels of HCY and Lp(a)in observation group 2 were in-creased(P<0.05).The serum levels of HCY and Lp(a)in observation group 2 were negatively correlated with ultrafiltration amount(P<0.05),and HCY and Lp(a)were positively correlated with parathyroid hor-mone(PTH),blood phosphorus(P),uric acid(UA)(P<0.05).Multivariate Logistic regression analysis showed that HCY,Lp(a),PTH,P,UA,diabetic nephropathy,duration of endovascular fistula use≥12 months,duration of puncture point compression≥30 min,and arteriovenous internal diameter<2 mm and vascular stenosis were the risk factors for AVF failure in MHD patients(P<0.05),and ultrafiltration a-mount was a protective factor for AVF failure in MHD patients(P<0.05).ROC curve results showed that the area under the curve of HCY and Lp(a)alone and in combination in the prediction of AVF failure in MHD patients were 0.822,0.820 and 0.908,respectively(Zcombination vs.HCY=3.144,Zcombination vs.Lp(a)=2.925,P<0.05).Conclusion The serum levels of HCY and Lp(a)in MHD patients with AVF failure are increased.They are the influencing factors of AVF failure in MHD patients,and the value of the combination of the two for pre-dicting AVF failure in MHD patients is relatively high.

OBJECTIVE：To establish a method for de termining stiripentol （STP）concentration in plasma and brain of rats ， and to compare the concentrations of STP and its self-nanoemulsifying drug delivery system （STP-SNEDDS）in plasma and brain. METHODS：Using xanthone as internal standard ，HPLC-fluorescence（HPLC-FLR）method was adopted. The determination was performed on Diamonsil C 18 column with mobile phase consisted of acetonitrile- 25 mmol/L KH 2PO4 solution [ 44 ∶ 56（V/V），pH 2.6] at a flow rate of 1.5 mL/min；the excitation and emission wavelengths were 210 nm and 400 nm，respectively；the column temperature was 30 ℃；the sample size was 10 μL. Totally 36 rats were randomly divided into 2 groups，with 18 rats in each group. They were given STP-SNEDDS and STP suspension （40 mg/kg，by STP ）intragastrically. Blood and brain tissue samples were collected at 0.5，1，2 h after administration （6 rats in each group at different time point ）. After the protein was precipitated by acetonitrile （brain tissue should be homogenized ），the concentrations of STP were determined by the above chromatographic conditions. RESULTS ：The linear ranges of STP concentration in plasma and brain tissue were 0.02-8.00 μg/mL（r were 0.999 6， 0.999 4，respectively）. The limits of quantitation were both 0.02 μg/mL. The inter-day and intra-day RSDs were all less than 5％. The extraction recovery and method recovery were all no less than 90％. Compared with STP suspension group ，the plasma concentration（except for 1 h after administration ）and cerebral concentration （except for 2 h after administration ）of STP in STP-SNEDDS group were all significantly increased （P＜0.05），showing significant linear relationship between them （for STP-SNEDDS）. CONCLUSIONS ：Established HPLC-FLR method presents high accuracy and precision ，and can be used for the distribution of STP and STP-SNEDDS in plasma and brain.The concentration of STP in plasma and brain tissue isincreased after STP is made into SNEDDS.

The incidence of allergic diseases in China is increasing year by year, which has caused heavy public health burden to individuals and society. The detection of specific IgE (sIgE) is an important way to diagnose the etiology of allergic disease. Currently, the diagnosis of allergic rhinitis In vitro mainly focus on the specific IgE of crude extracts in clinical practice, while the detection of sIgE in allergen components is rarely carried out. Clinicians, especially non-allergists, do not have sufficient understanding about the importance of sIgE in allergen component detection. Knowing the related types and clinical significance of allergen components can improve the diagnostic level of allergic diseases. Allergen component detection can distinguish the major components of common allergens, identify cross-sensitization, predict the risk of anaphylaxis, guide allergen immunotherapy and develop precise dietary regimens, so as to provide accurate prevention and control recommendations for patients.

The incidence of allergic diseases in China is increasing year by year, which has caused heavy public health burden to individuals and society. The detection of specific IgE (sIgE) is an important way to diagnose the etiology of allergic disease. Currently, the diagnosis of allergic rhinitis In vitro mainly focus on the specific IgE of crude extracts in clinical practice, while the detection of sIgE in allergen components is rarely carried out. Clinicians, especially non-allergists, do not have sufficient understanding about the importance of sIgE in allergen component detection. Knowing the related types and clinical significance of allergen components can improve the diagnostic level of allergic diseases. Allergen component detection can distinguish the major components of common allergens, identify cross-sensitization, predict the risk of anaphylaxis, guide allergen immunotherapy and develop precise dietary regimens, so as to provide accurate prevention and control recommendations for patients.

To investigate the inhibitory effect of sildenafil on Caco-2 cell proliferation and its anti-inflammatory effect on menadione-induced NCM460 cell inflammation model, MTT assay was used to determine cell proliferation. Intracellular reactive oxygen species(ROS)and nitric oxide(NO)levels were detected by fluorescent probe. Western blot was used to detect the expression of eNOS/ERK/JNK pathway related proteins in Caco-2 cells and correlated inflammatory cytokines in NCM460 cells. The effect of sildenafil on the growth of two probiotics was determined by spectrophotometry. Results showed that sildenafil signi-ficantly inhibited the proliferation of Caco-2 cells and enhanced the expression levels of eNOS, p-eNOS, p-JNK1/2 and p-ERK1/2 proteins in Caco-2 cells; while after adding NG-nitro-L-arginine methyl ester(L-NAME), the expression levels of eNOS, p-eNOS, p-JNK1/2 and p-ERK1/2 proteins were significantly lower than those of the sildenafil group. Compared with the menadione group, sildenafil significantly reduced ROS levels in NCM460 cells and inhibited the expression levels of IL-6, IL-1β, p62, and TNF-α. Moreover, high concentrations of sildenafil had no obvious toxic effects on Lactobacillus casei and Lactobacillus rhamnosus. Thus, the results indicated that sildenafil could effectively inhibit the intestinal inflammatory response without affecting the balance of the intestinal flora, and prevent colorectal cancer by reducing the oxidative stress responses in the intestinal cells.

Objective:To observe the efficacy and safety of sirolimus combined with calcineurin inhibitor (CNI) in the treatment of glucocorticoid resistant/dependent extensive chronic graft-versus-host disease (cGVHD) .Methods:A total of 27 patients with steroid-resistant/steroid-dependent extensive cGVHD from November 2015 to January 2019 were enrolled and given sirolimus capsules combined with cyclosporine or tacrolimus to observe the clinical efficacy and adverse events.Results:The median duration of medication was 14.2 months and the mean duration was 16.7 months. The median follow-up time was 20.1 months (12.9-46.1 months) . Following the 6-month follow-up, 3 cases achieved complete response (CR) and 12 cases partial response (PR) . The overall response rate (ORR) was 55.6% ; for progression-free survival (PFS) , PFS-6 reached 88.9% (24/27) , and for overall survival (OS) , OS-6 was 100% . At the 1-year follow-up, there were 5 cases of CR and 11 cases of PR, ORR was 59.3% , PFS-12 reached 62.9% (17/27) , and OS-12 was 100% . The subgroup analysis found that the program was more effective for cGVHD in male donors and the target organ analysis had an advantage in the treatment of oral cavity, skin, and liver rejection. Adverse events were observed: hyperlipidemia 11.1% , oral ulcer 7.4% , fungal infection 11.1% , liver injury 3.7% , renal insufficiency 0, and no new CMV and EB viremia.Conclusion:Sirolimus combined with calcineurin inhibitors is effective in treating steroid-resistant/steroid-dependent extensive cGVHD, especially because adverse reactions (renal toxicity, CMV, EBV infection) are low in number, which is suitable for long-term treatment of cGVHD.

Objective: Chronic graft-versus-host disease (cGVHD) is a major long-term complication after allogeneic hematopoietic stem cell transplantation (allo-HSCT) . It is important to study the changes of serum biomarkers expression in patients for early diagnosis and treatment. Methods: The expression levels of five serum protein markers (IL-1b, IL-16, CXCL9, CCL19, CCL17) in patients with or without cGVHD after allo-HSCT were detected by liquid suspension microarray. Results: Compared with the control group without cGVHD, the expression levels of CXCL9 and CCL17 in serum of patients with cGVHD were significantly increased (P<0.05) . CCL17 was correlated with the severity of cGVHD (P<0.001) . CXCL9 was significantly increased in the serum of patients with skin lesion (P<0.01) , and CCL17 was significantly expressed in cGVHD patients with liver as the target organ (P<0.01) . Conclusion The combination of CXCL9 and CCL17 can be used as serum biomarkers of cGVHD, which has certain reference value in assisting the diagnosis and evaluation of cGVHD severity.

OBJECTIVE： To study the dose-time-effect relationship of Tibetan medicine Rannasangpei in cerebral ischemic- reperfusion injury model rats with intragastric administration. METHODS： The rats were randomly divided into sham operation group （normal saline， 10 mL/kg）， model control group （normal saline， 10 mL/kg）， positive control group （nimodipine， 30      mg/kg）， Rannasangpei different dose groups （0.52， 1.04， 2.08， 4.17， 8.33， 16.67， 33.34， 66.68， 133.36， 266.72 and 533.44    mg/kg）， with 18 rats in each group. Each group was given relevant medicine intragastrically once； 25 min after intragastric administration， cerebral ischemic-reperfusion injury model was established with suture-occluded method in those groups except for sham operation group. 24， 48， 72 h after cerebral ischemia， neuroethology of rats were graded in each group. The rate of cerebral infraction was detected to evaluate the optimal effective time， the optimal dose （Dmax） and maximal effect （the rate of minimum cerebral infraction， Emax） of Ratnasampil at different periods of cerebral ischemia. Dose-time-effect relationship of Rannasangpei dose with the rate of cerebral infraction was fitted with Thermo Kinetica 5.1 software. The area under curve （AUClast） and retention dose （MRTlast） of dose-effect curve were calculated， and detect the levels of SOD and MDA. RESULTS： Compared with sham operation group， the neurobehavior of model group was significantly abnormal （P＜0.05 or P＜0.01）， and the rate of cerebral infarction was significantly increased （P＜0.01）； the level of SOD was decreased significantly （P＜0.01， 48 h）， and the level MDA was increased significantly （P＜0.05， 48 h）. Compared with model control group， there was no significant change in neurobehavioral abnormalities in the nimodipine group （P＞0.05）， and the rate of cerebral infraction was decreased significantly （P＜0.01， 24， 48 h）. The level of SOD in rats were increased significantly （P＜0.01， 48 h）， while the level MDA decreased significantly （P＜0.05， 48 h）. Rannasangpei 2.08-33.34 mg/kg could significantly improved neurobehavioral abnormalities （P＜0.05， 24 h）； 24 h after cerebral ischemia， the rate of cerebral infraction was decreased significantly in Rannasangpei 4.17-133.36 mg/kg group （the lowest is 33.34 mg/kg group， P＜0.05 or P＜0.01）. The level of SOD in rats were increased significantly in 33.34-533.44 mg/kg groups （P＜0.05）. the level MDA was decreased significantly in 0.52-2.08， 8.33， 33.34， 266.72 and 533.44 mg/kg groups （P＜0.05）. Dmax was 33.34 mg/kg， Emax was 3.02％， AUClast was 5 141.76 mg/kg and MRTlast was 329.161 mg/kg. 48 h after cerebral ischemia， the rate of cerebral infraction was decreased significantly in Rannasangpei 2.08-133.36 mg/kg groups （the lowest is 66.68 mg/kg group， P＜0.05 or P＜0.01）， while the level of SOD was increased significantly in 1.04-533.44（except for 4.17）mg/kg groups （P＜0.05）. The level of MDA was decreased significantly in 16.67-66.68， 533.44 mg/kg groups （P＜0.05）， Dmax was 66.68 mg/kg， Emax was 2.13％， AUClast was    5 219.36 mg/kg and MRTlast was 340.521 mg/kg. 72 h after cerebral ischemia， the rate of cerebral infraction and the level of MDA had no significant decreased in Rannasangpei groups （P＞0.05）， and the levels of SOD had no significant increase （except for 0.52 mg/kg group， P＞0.05）. CONCLUSIONS： The optimal effective time of Rannasangpei for the treatment of cerebral ischemia-reperfusion injury in rats is 48 h， and the Dmax is 66.68 mg/kg. The improvement mechanism may be related to increase the level of SOD and decrease the level of MDA.

In order to study the molecular mechanism and physiological significance of the interaction between PGRN and Rev-erbβ, the PGRN gene in HEK293 (Rev-erbβ-/-) marked as C3-6 cell lines was knocked out by CRISPR/Cas9 system to generate the Rev-erbβ and PGRN double genes knockout HEK293 cell lines. First, four sgRNAs were designed for PGRN gene, and PGRN sgRNA2 and sgRNA3 with the higher activity were used to construct the Lentiviral vector, pLenti/CMV-Loxp-Cas9-sgRNA2-U6-sgRNA3-U6-Loxp-EF1α-Puro. Then, the lentivirus vector carrying Cas9 and double PGRN sgRNA were used to infect HEK293 C3-6 cells. Through drug screening, cloning and sequencing, we obtained the monoclonal HEK293 (Rev-erbβ-/-; PGRN-/-) marked as C3-6/23 cell lines. Using qRT-PCR and Western blotting, we detected PGRN mRNA and protein expression in C3-6/23 cell lines. Finally, genetic complementation was used to study the effect of PGRN-mediated Rev-erbβ on the regulation of the target gene promoter transcriptional activity in the C3-6/23 cell lines. In HEK293 C3-6/23 cell lines, the two DNA chains of PGRN gene were both deletion mutagenesis, and the expression mRNA and protein of PGRN did not reach the detection level. At the same time, the interaction between PGRN and Rev-erbβ enhanced the regulation of Rev-erbβ on the transcription of target gene promoter in the cell lines. Using CRISPR/Cas9 system, we successfully constructed the double knockout HEK293 (Rev-erbβ-/-; PGRN-/-) monoclonal cell lines. The study found that PGRN could affect Rev-erbβ on the regulation of target gene promoter transcription in the C3-6/23 cell lines; however, the mechanism of PGRN involvement in mediating Rev-erbβ in transcriptional regulation remains to be further studied.