Mechanism of Buzhong Yiqitang Intervening in Pyroptosis of AIT in NOD.H-2h4 Mice Based on NLRP3/Caspase-1/GSDMD Pathway / 中国实验方剂学杂志

ObjectiveTo explore the mechanism of Buzhong Yiqitang on pyroptosis in autoimmune thyroiditis （AIT） mice based on the NOD-like receptor hot protein domain related protein 3 （NLRP3）/cysteinyl aspartate specific proteinase-1（Caspase-1）/Gasdermin D （GSDMD） pathway. MethodSixty NOD.H-2h4 mice were divided into normal group， model group， low， medium， and high dose groups （4.10， 8.19， 16.38 g·kg-1）of Buzhong Yiqitang， and selenium yeast tablet group （0.26 mg·kg-1）， with 10 mice in each group. Except for the normal group， all other groups were given 0.05% NaI by gavage for eight weeks to establish a model and then received the drug treatment for eight weeks. The serum levels of thyroid peroxidase antibody （TPO-Ab） and thyroglobulin antibody （TgAb） were detected using enzyme-linked immunosorbent assay （ELISA） method. Hematoxylin-eosin （HE） staining was used to observe the pathological changes in mouse thyroid tissue. The immunohistochemical method was used to detect the protein expression of NLRP3， Caspase-1， interleukin-1β （IL-1β）， and interleukin-18 （IL-18）. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was used to detect the mRNA expression of NLRP3， Caspase-1， IL-1β， and IL-18. Western blot was used to detect the levels of pyroptosis-related proteins in thyroid tissue. ResultCompared with the normal group， the serum levels of TPO-Ab and TgAb in the model group were significantly increased （P<0.01）. Thyroid follicles either increased in a cubic shape or were damaged and atrophied， with a large number of lymphocytes infiltrating around the follicles. Compared with the model group， the levels of TPO-Ab and TgAb in other groups were significantly reduced （P<0.01）， and the morphology and structure of follicles were improved. The degree of lymphocyte infiltration was reduced. Among them， the medium dose group of Buzhong Yiqitang had the most significant reduction and improvement effect. Compared with the normal group， the positive products and mRNA expression of NLRP3， Caspase-1， IL-1β， and IL-18 proteins in the thyroid tissue of the model group significantly increased （P<0.01）， and the protein expression levels of NLRP3， cleaved Caspase-1， IL-1β， IL-18， and GSDMD-N were significantly increased （P<0.05， P<0.01）. Compared with the model group， the positive products and mRNA expression of NLRP3， Caspase-1， IL-1β， and IL-18 proteins in other groups were significantly reduced （P<0.05， P<0.01）， with the most significant reduction effect in the medium dose group of Buzhong Yiqitang. The protein expression levels of NLRP3， cleaved Caspase-1， IL-1β， IL-18， and GSDMD-N were significantly reduced （P<0.05， P<0.01）. ConclusionBuzhong Yiqitang can improve AIT， and its mechanism may be achieved by regulating the NLRP3/Caspase-1/GSDMD signaling pathway to inhibit pyroptosis.

Efficacy and safety of evolocumab in elderly patients with high-risk cardiovascular diseases / 中华老年心脑血管病杂志

Objective To explore the efficacy and safety of evolocumab in elderly patients with high-risk cardiovascular diseases.Methods A total of 153 patients with poor lipid control after conventional statin therapy who were hospitalized in the cardiologic departments in the First,Sec-ond,Sixth and Eighth Medical Centers of Chinese PLA General Hospital from November 2019 to November 2022 were included,and divided into non-elderly group(＜60 years old,46 cases),eld-erly group(60-74 years old,66 case)and very elderly group(≥75 years old,41 cases).They were all given evolocumab treatment according to guidelines.Another 50 over-75-year-old patients with high-risk cardiovascular diseases and poor lipid control who were hospitalized in the above cardiologic departments during the same period were treated with a statin drug combined with ezetimibe,and served as conventional treatment group(control group).The baseline clinical data and the blood indicators at 4th and 12th week after drug administration,and the occurrence of ad-verse drug reactions and major adverse cardiovascular events(MACE)within 12 weeks were com-pared among the groups.Results The levels of LDL-C and TC were significantly decreased in the three evolocumab treatment groups at 4 and 12 weeks after medication when compared with the baseline values(P＜0.05,P＜0.01),but there were no obvious differences in the 2 levels among the 3 groups at 12 weeks(P＞0.05).At the time point,no statistical difference was observed in the incidence of adverse events in the three groups(2.2％vs 3.0％vs 2.4％,P＞0.05).The levels of LDL-C and TC were decreased significantly in the very elderly group and the conventional treatment group at the 12th week when compared with the baseline levels(P＜0.05,P＜0.01),and the LDL-C level at the week was notably lower in the very elderly group than the convention-al treatment group(1.36±0.44 mmol/L vs 1.87±0.56 mmol/L,P＜0.01).But no difference was seen in the incidence of MACE between the 2 groups(12.2％vs 16.0％,P＞0.05),either in sur-vival rate between them(P=0.576).Conclusion For patients of all ages,evolocumab has good short-term efficacy in lipid control,and for those over 75 years old,the drug also shows good effi-cacy and sound safety.

Effects of Zhuyu Pill （茱萸丸） and its Ingredient on Aortic Plaque and the NLRP3/Asc/Caspase-1 Signaling Pathway in Aortic Tissue of Mice in Atherosclerosis Model / 中医杂志

ObjectiveTo investigate the possible mechanism of action and combination of medicinals of Zhuyu Pill （茱萸丸） in the treatment of atherosclerosis. MethodsThirteen normal C57BL/6J mice were used as blank group， and 40 ApoE-/- mice of the same strain were randomly divided into model group， Huanglian （Coptis chinensis Franch.） group， Wuzhuyu （Tetradium ruticarpum） group， and Zhuyu Pill group， with 10 mice in each group. Except the blank group， the other groups were fed with high-fat diet for modeling， and the total modeling cycle was 12 weeks. After modelling， Huanglian group was given Huanglian solution 143.34 mg/（kg·d）， Wuzhuyu group with Wuzhuyu solution 301.78 mg/（kg·d）， Zhuyu Pill group with 261.07 mg/（kg·d） of Zhuyu Pill solution， blank group and model group were given 0.3ml of normal saline. Mice in all groups were gavaged for 12 weeks. The levels of total cholesterol （TC）， triglyceride （TG）， low density lipoprotein cholesterol （LDL-C） and fasting blood glucose were measured by biochemical method； fasting insulin in serum was detected by enzyme-linked immunosorbent assay （ELISA） and insulin resistance index was calculated； HE staining was used to observe the pathological and morphological changes of aortic tissue， liver and epididymal fat in mice， and oil red O staining was used to observe the lipid deposition of aortic tissue in mice， and the percentage of positive area of aortic HE staining， the percentage of positive area of aortic oil red O staining， the non-alcoholic fatty liver activity score （NAS） score， and the cross-sectional area of epididymal fat were calculated； serum levels of interleukin 1β （IL-1β） and interleukin 18 （IL-18） were detected by ELISA； protein expression of nucleotide-binding oligomerisation structural domain-like receptor family pyridoxal structural domain protein 3 （NLRP3） in aortic tissues was detected by Western blot； and immunofluorescence was used to detect the levels of apoptosis-associated granulocyte-like protein （ASC）， which contains the CARD structural domain （ASC） and cysteine-containing aspartic protease 1 （Caspase-1） protein expression； and real-time fluorescence PCR was used to detect NLRP3， ASC， Caspase-1， IL-1β， and IL-18 mRNA expression in mouse aortic tissues. ResultsCompared with blank group， serum TC， TG and LDL-C were increased， insulin was decreased， fasting blood glucose and insulin resistance index were increased in model group （P＜0.01）. Aortic plaque area increased， liver lipid deposition increased， epididymal fat cells volume increased， liver NAS score increased， and epididymal fat cross-sectional area increased. Serum IL-1β and IL-18 increased， and the expression levels of NLRP3， ASC， Caspase-1 protein and mRNA in aortic tissue increased （P＜0.01）. Compared with model group， serum TC， TG and LDL-C in Huanglian group， Wuzhuyu group and Zhuyu Pill group decreased， fasting insulin increased， fasting blood glucose and insulin resistance index decreased （P＜0.01）； aortic plaque area decreased significantly， liver lipid deposition decreased， liver NAS score decreased， epididymal fat cell volume decreased， epididymal fat cross section area decreased， serum IL-1β and IL-18 were decreased， the expression levels of NLRP3， ASC， Caspase-1 protein and mRNA in aortic tissue were decreased （P＜0.01）， and the improvement of all indexes in the Zhuyu Pill group was better than that in Huanglian and Wuzhuyu groups （P＜0.01）. ConclusionZhuyu Pill has a good therapeutic effect on atherosclerosis in mice， and the combination of Huanglian and Wuzhuyu has a synergistic effect， the mechanism of which may be related to regulation of the NLRP3/ASC/Caspase-1 aortic signaling pathway， and thus reducing the vascular inflammatory response.

Salvianolic Acid F Regulates Bax/Caspase-3/GSDME Signaling Pathway to Inhibit Pyroptosis of HK-2 Cells / 中国实验方剂学杂志

ObjectiveTo investigate the mechanism of salvianolic acid F （Sal F） in repairing the high glucose-induced injury in human kidney-2 （HK-2） cells via the B-cell lymphoma-2 （Bcl-2）-associated X protein （Bax）/cysteinyl aspartate-specific proteinase 3 （Caspase-3）/gasdermin-E （GSDME） pathway. MethodThe cell counting kit-8 （CCK-8） was used to measure the relative viability of HK-2 cells exposed to high glucose and different concentrations （2.5， 5， 10， 20 μmol·L-1） of Sal F and the relative viability of HK-2 cells treated with Sal F for different time periods. The levels of lactate dehydrogenase （LDH） and interleukin-1β （IL-1β） in the supernatant of the cell culture were measured by the LDH assay kit and enzyme-linked immunosorbent assay （ELISA） kit， respectively. Flow cytometry combined with Annexin V-FITC/propidium iodide （PI） and Hoechst 33342/PI staining was employed to reveal the proportion of PI-positive HK-2 cells exposed to high glucose. Western blotting was employed to determine the protein levels of Bax， Bcl-2， cytochrome C， cysteinyl aspartate-specific proteinase （Caspase）-9， Caspase-3， and GSDME in the HK-2 cells exposed to high glucose and treated with Sal F. The 2，7-dichlorodihydrofluorescein diacetate fluorescence probe （DCFH-DA） and mitochondrial membrane potential assay kit （JC-1） were used to determine the production of reactive oxygen species （ROS） and the mitochondrial membrane potential in the HK-2 cells exposed to high glucose and treated with Sal F. ResultCompared with the blank group， the model group showed decreased cell viability （P<0.01）， elevated levels LDH and IL-1β， increased proportion of PI-positive cells （P<0.01）， up-regulated protein levels of Bax， cytochrome C， Caspase-9， Caspase-3， and GSDME （P<0.01）， down-regulated protein level of Bcl-2 （P<0.01）， decreased mitochondrial membrane potential， and excessive ROS accumulation. Compared with the model group， Sal F repaired the high glucose-induced injury in HK-2 cells （P<0.05）， lowered the levels of LDH and IL-1β （P<0.05， P<0.01）， and decreased the proportion of PI-positive cells （P<0.01）. In addition， Sal F down-regulated the protein levels of Bax， cytochrome C， Caspase-9， Caspase-3， and GSDME and up-regulated the protein level of Bcl-2 （P<0.05， P<0.01）， increased the mitochondrial membrane potential， and decreased the accumulation of ROS in HK-2 cells. ConclusionSal F can reduce the production of ROS， restore the balance of mitochondrial membrane potential， and inhibit pyroptosis via the Bax/Caspase-3/GSDME signaling pathway to repair the high glucose-induced injury in HK-2 cells.

Osmoregulatory Appraisal of Some Osmoprotectants on Hydrolytic Activities of Some Enzymes on Seeds of Two Water Stressed Cultivars of Sorghum bicolor (Moench)

Enzymes play significant roles in metabolic processes of seeds. Therefore, this study evaluated osmoregulatory potential of some osmoprotectants on activities of some hydrolytic enzymes in the seeds of two cultivars (SOSAT.C-88 and CV. LCIC 9702) of sorghum bicolor. Matured seeds of the two cultivars were harvested and prepared for alpha, beta, total amylase and proteinase activities assay. The osmoprotectants produced significant variations on the enzymes at 10 and 14 days (DA) of 8 weeks after treatments (WAT). Seeds of well-watered SOSAT.C-88 produced higher alpha (2.10 IU/ml), beta (1.70 IU/ml) and total amylase activities (3.30 IU/ml) at 14 days (DA). Higher alpha (2.01 IU/ml and total amylase activities (2.61 IU/ml) were recorded in the seeds of CV. LCIC 9702 well-watered at 14 days DA 8WAT. Furthermore, total amylase activities (3.87 IU/ml) were recorded in the seeds produced by CV. LCIC 9702 well-watered at 14 days DA. Significant increase was noticed in beta (1.14 IU/ml) and alpha amylase (1.58 IU/ml) in the seeds of CV. LCIC 9702 treated with mycorrhiza. CV. LCIC 9702 well watered produced highest proteinase activities (1.57 U/ml) while least of the parameters were recorded in SOSAT.C-88 and CV. LCIC 9702 droughted. In conclusion, the osmoprotectants had regulatory effects on the activities of hydrolytic enzymes therefore the use of the osmoprotectants in farming should be encouraged.

Effect of Modified Huangqi Guizhi Wuwutang on Apoptosis-related Bax and Caspase-12 of Sciatic Nerve Cells in Diabetes Rats / 中国实验方剂学杂志

ObjectiveTo investigate the effect of modified Huangqi Guizhi Wuwutang （MHGW） on the protein and mRNA expression of B-cell lymphoma-2-associated X protein （Bax） and cysteinyl aspartate specific proteinase-12 （Caspase-12） related to the apoptosis of sciatic nerve cells in diabetes rats to explore the mechanism of MHGW in the treatment of peripheral neuropathy in diabetes. MethodAnimal experiments were conducted. A diabetes model was induced in sixty male sprague-dawley （SD） rats by feeding on a high-sugar and high-fat diet combined with streptozotocin （STZ） intraperitoneal injection. Rats with random blood glucose levels ≥ 16.7 mmol·L-1 for three consecutive days were considered to have successfully developed diabetes. Forty-eight rats that successfully developed diabetes were randomly divided into a model group， an α-lipoic acid group （0.026 8 g·kg-1·d-1）， a high-dose MHGW group （2.5 g·kg-1·d-1）， and a low-dose MHGW group （1.25 g·kg-1·d-1）， with 12 rats in each group. Another 10 rats were assigned to the normal group. Body weight and random blood glucose levels of the rats were monitored. At the end of a 16-week intervention period， the sciatic nerve conduction velocity of the rats was measured using the Key point electromyography collection system. The protein and mRNA expression of Bax and Caspase-12 in the sciatic nerve cells was detected by Western blot analysis and real-time quantitative polymerase chain reaction （Real-time PCR）， respectively. ResultCompared with the normal group， the model group showed a significant decrease in body weight （P<0.01） and a significant increase in random blood glucose levels （P<0.01）. After a 16-week intervention， compared with the model group， the high-dose MHGW group exhibited a significant increase in body weight （P<0.05）， while there were no statistically significant differences in body weight changes among the other treatment groups. Random blood glucose levels significantly decreased in all treatment groups （P<0.01）. After 16 weeks of intervention， compared with the normal group， the model group had significantly reduced motor and sensory nerve conduction velocities （P<0.01）. Compared with the model group， all treatment groups showed significant increases in motor and sensory nerve conduction velocities （P<0.05， P<0.01）. The expression of Bax and Caspase-12 proteins in the sciatic nerve cells was significantly elevated in the model group compared with that in the normal group （P<0.01）. In contrast， all treatment groups showed significant reductions in the expression of Bax and Caspase-12 proteins in the sciatic nerve cells as compared with that in the model group （P<0.01）. The expression of Bax and Caspase-12 mRNA in the sciatic nerve cells significantly increased in the model group compared with that in the normal group （P<0.01）. Compared with the model group， the α-lipoic acid group and the high-dose MHGW group showed significant reductions in the expression of Bax mRNA in the sciatic nerve cells （P<0.05， P<0.01）， while the low-dose MHGW group showed a decreasing trend in the expression of Bax mRNA. The expression of Caspase-12 mRNA in the sciatic nerve cells significantly decreased in all treatment groups （P<0.01）. ConclusionMHGW may improve and repair sciatic nerve damage in diabetes rats by inhibiting sciatic nerve cell apoptosis.

Preliminary study of the role of IL-18/IL-18BP in mediating cytotoxic ability of NK-92MI cells against endothelial cells from GTKO porcine models / 器官移植

Objective To evaluate the role and potential mechanism of interleukin (IL)-18/IL-18 binding protein (BP) in mediating the killing effect of natural killer (NK)-92MI cells upon endothelial cells from α-1, 3- galactosyltransferase gene-knockout (GTKO) porcine models. Methods NK-92MI cells were divided into the NK, NK+IL-18, NK+GTKO, IL-18+NK+GTKO and IL-18+IL-18BP+NK+GTKO groups. The messenger ribonucleic acid (mRNA) levels of inflammation-related genes in NK-92MI cells were detected by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). The killing effect of NK-92MI cells on endothelial cells from GTKO porcine models was evaluated by lactate dehydrogenase (LDH) assay. The apoptosis of endothelial cells from GTKO porcine models was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay. The expression levels of proteins with killing effect and apoptosis-related proteins were determined by Western blot. Results Compared with the NK, NK+IL-18 and NK+GTKO groups, the expression levels of interferon (IFN)-γ, tumor necrosis factor (TNF)-α, IL-8, IL-3, IL-6 and granulocyte-macrophage colony stimulating factor (GM-CSF) mRNA were up-regulated in NK-92MI cells in the IL-18+NK+GTKO group, and the differences were statistically significant (all P < 0.05). Compared with the IL-18+NK+GTKO group, the expression levels of IFN-γ, TNF-α, IL-8, IL-3, IL-6 and GM-CSF mRNA were down-regulated in NK-92MI cells in the IL-18+IL-18BP+NK+GTKO group, and the differences were statistically significant (all P < 0.05). Compared with the NK+GTKO group, the expression levels of perforin, granzyme B and IFN-γ proteins in NK-92MI cells were up-regulated, the killing rate of NK-92MI cells against endothelial cells from GTKO porcine models was enhanced, the apoptosis rate of endothelial cells from GTKO porcine models was increased, and the ratios of B cell lymphoma-2 (Bcl-2)-associated X protein (Bax)/Bcl-2 and cleaved Caspase-3/Caspase-3 in endothelial cells from GTKO porcine models were elevated in the IL-18+NK+GTKO group, and the differences were statistically significant (all P < 0.05). Compared with the IL-18+NK+GTKO group, the expression levels of perforin, granzyme B and IFN-γ proteins were down-regulated, the killing rate of NK-92MI cells against endothelial cells from GTKO porcine models was decreased, the apoptosis rate of endothelial cells from GTKO porcine models was decreased, and the ratios of Bax/Bcl-2 and cleaved Caspase-3/Caspase-3 in endothelial cells from GTKO porcine models were declined in the IL-18+IL-18BP+NK+GTKO group, and the differences were statistically significant (all P < 0.05). Conclusions IL-18BP may block the expression of inflammation-related genes in NK-92MI cells induced by IL-18 and the killing effect of NK-92MI cells on endothelial cells from GTKO porcine models.

Relationship between interleukin-1β and silicosis development / 中国基层医药

Silicosis is a diffuse pulmonary fibrosis disease caused by occupational exposure to silica, which is one of the occupational diseases with high incidence in developing countries. Up to now, there is no definite drug to relieve or reverse the lung injury caused by silicosis, so it is very important to prevent, diagnose and treat pulmonary fibrosis as soon as possible. Studies have shown that a chronic inflammatory environment contributes to pulmonary fibrosis to a certain extent. Interleukin-1β is a cytokine that increases the number of inflammatory factors in the microenvironment in the immune response and plays a key role in inflammatory reaction. Therefore, the release of interleukin-1β is of great significance in the pathogenesis of silicosis. This paper aims to systematically expound the development course of silicosis, the signal pathway of interleukin-1β production, and the relationship between them.

Relationship between serum CASP2 expression and NBNA score and prognosis in children with hypoxic-ischemic encephalopathy / 国际检验医学杂志

Objective To study the correlation between serum cysteine aspartate proteinase 2(CASP2)level and neonatal behavioral neurological assessment(NBNA)score in children with hypoxic-ischemic en-cephalopathy(HIE),and the value of CASP2 in evaluating prognosis.Methods A total of 114 neonates with HIE in this hospital from January 2018 to April 2022 were selected as HIE group.According to the severity of the disease,the neonates were divided into mild group(40 cases),moderate group(52 cases)and severe group(22 cases).According to the prognosis after 6 months of treatment,114 neonates with HIE were divided into poor prognosis group(26 cases)and good prognosis group(88 cases).At the same time,106 healthy neonates born in the hospital were selected as the control group.The NBNA score was recorded,and the serum CASP2 level was detected by enzyme-linked immunosorbent assay.Pearson correlation was used to analyze the corre-lation between serum CASP2 level and NBNA score in HIE children.Receiver operating characteristic(ROC)curve was used to analyze the predictive value of NBNA score and serum CASP2 level for the prognosis of ne-onates with HIE.Results The level of serum CASP2 in HIE group was higher than that in control group,and the NBNA score was lower than that in control group(P<0.05).The level of serum CASP2 in the severe group was higher than that of the moderate group and mild group,and the NBNA score was lower than that of moderate group and mild group(P<0.05).The level of serum CASP2 in the moderate group was higher than that of the mild group,and the NBNA score was lower than that of the mild group(P<0.05).The NBNA score was negatively correlated with the serum level of CASP2 in the children with HIE(r=-0.608,P<0.05).The poor prognosis group had a significantly higher serum level of CASP2 and a significantly lower NBNA score than the good prognosis group(P<0.05).The serum level of CASP2 and NBNA score separate and joint area under the curve of the poor children with HIE prognosis were 0.875(95%CI 0.797-0.952),0.748(95%CI 0.639-0.857),0.927(95%CI 0.873-0.981).Conclusion The level of serum CASP2 increases with the severity of HIE,and is closely related to the NBNA score,which has an important prognostic value.

The effect of Ephedra aqueous extract on lung injury of rats with Streptococcus pneumoniae pneumonia via regulating ROS/NLRP3/Caspase-1 pathway / 免疫学杂志

This study was performed to evaluate the role and mechanism of Ephedra aqueous extract in alleviating lung injury of rats with Streptococcus pneumoniae pneumonia via regulating reactive oxygen species(ROS)/NOD-like receptor protein 3(NLRP3)/caspase-1 pathway.Male SD rats were randomly divided into control group,model group,Ephedra aqueous extract(100 mg/kg,200 mg/kg,400 mg/kg)groups,solvent control group(110 ml/kg DMSO),model+solvent group,high-dose Ephedra aqueous extract+solvent group,and high-dose Ephedra aqueous extract+trimethylamine N-oxide(TMAO,110 mg/kg)group.Pneumonia model was established by instilling Streptococcus pneumoniae solution into the trachea.After 24 hours of modeling,each group was given medication by gavage for 10 consecutive days.24 hours after the last gastric lavage,pathological changes in lung tissue and the contents of interleukin(IL)-1β,IL-6,tumor necrosis factor-α(TNF-α)in serum and bronchoalveolar lavage fluid(BALF)were detected;and the contents of malondialdehyde(MDA),superoxide dismutase(SOD),the activity of ROS and the expression levels of NLRP3,Caspase-1,GSDMD-N in lung tissue were detected.The model group rats demonstrated pathological changes in lung injury including thickening of alveolar septa and infiltration of inflammatory cells,as well as the contents of IL-1β,IL-6,TNF-α in BALF and serum,the content of MDA,the activity of ROS,the expression levels of NLRP3,Caspase-1 and GSDMD-N in lung tissue were higher than those of the control group,while the SOD content in lung tissue was lower than that of the control group(P<0.05).The lung injury of the low,medium,and high dose Ephedra aqueous extract groups were reduced,and the contents of IL-1β,IL-6,TNF-α in BALF and serum,the content of MDA,the activity of ROS,the expression levels of NLRP3,Caspase-1 and GSDMD-N in lung tissue were lower than that of the model group,while the SOD content in lung tissue was higher than that of the model group(P<0.05).The lung injury of rats in the high-dose ephedra aqueous extract+TMAO group worsened,the contents of IL-1β,IL-6,TNF-α in BALF and serum,the content of MDA,the activity of ROS,the expression levels of NLRP3,Caspase-1 and GSDMD-N in lung tissue were higher than those of the high-dose Ephedra aqueous extract+solvent group,while the SOD content in lung tissue was lower than that of the high-dose Ephedra aqueous extract+solvent group(P<0.05).In conclusion,Ephedra aqueous extract alleviates the lung injury of rats with Streptococcus pneumoniae pneumonia via inhibiting the inflammatory response,oxidative stress and pyroptosis mediated by ROS/NLRP3/Caspase-1 pathway.

Effects of different doses of sivelestat sodium on perioperative acute lung injury in patients undergoing acute Stanford type A aortic dissection surgery / 中华麻醉学杂志

Objective:To evaluate the effects of different doses of sivelestat sodium on perioperative acute lung injury (ALI) in the patients undergoing emergency surgery for acute Stanford type A aortic dissection (AAAD).Methods:A total of 120 patients of both sexes, aged 30-64 yr, with body mass index of 18.5-24.9 kg/m 2, of American Society of Anesthesiologists Physical Status classification Ⅲ or Ⅳ, scheduled for emergency AAAD surgery, were divided into 3 groups using a random number table method: low-dose sivelestat sodium group (SL group), medium-dose sivelestat sodium group (SM group)and high-dose sivelestat sodium group (SH group), with 40 patients in each group. Sivelestat sodium 4.8, 6.0 and 7.2 mg/kg were intravenously infused starting from 10 min before anesthesia until 24 h after surgery in SL, SM and SH groups, respectively. Blood samples from the radial artery were collected for blood gas analysis after anesthesia induction and before skin incision (T 1), immediately after the end of surgery (T 2), at 24 h after surgery (T 3), and 72 h after surgery (T 4), the alveolar-arterial oxygen tension difference (PA-aDO 2), oxygenation index (OI)and respiratory index (RI) were calculated. The duration of postoperative mechanical ventilation, length of stay in the intensive care unit (ICU) and length of postoperative hospital stay were recorded. Central venous blood samples were collected at T 1-T 4 to measure serum concentrations of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6)and IL-8. Peripheral venous blood samples were collected on preoperative day 1 and postoperative days 1 and 3 to measure white blood cell (WBC) count, neutrophil (NEUT) count, neutrophil percentage (NEUT%), and C-reactive protein (CRP) concentration. The occurrence of postoperative pulmonary complications (PPCs)and 90-day all-cause mortality were recorded. Results:Compared with the baseline at T 1, PA-aDO 2 and RI were significantly increased, OI was decreased, and the serum concentrations of TNF-α, IL-6 and IL-8 were increased at T 2-T 4 in all the three groups ( P<0.05). WBC, NEUT, NEUT% and concentrations of CRP were significantly higher on postoperative days 1 and 3 than on 1 day before surgery in the three groups ( P<0.05). Compared with SL and SM groups, PA-aDO 2 and RI were significantly decreased, OI was increased, and the serum concentrations of TNF-α, IL-6 and IL-8 were decreased, the WBC count, NEUT count, NEUT% and concentrations of CRP were decreased, the incidence of postoperative hypercapnia, hypoxemia, emerging lung rales and bronchospasm was decreased, and the duration of postoperative mechanical ventilation and length of intensive care unit stay were shortened( P<0.05), and no significant change was found in the postoperative length of hospital stay and 90-day all-cause mortality rate in SH group ( P>0.05). Conclusions:Sivelestat sodium 7.2 mg/kg can significantly inhibit the inflammatory responses, alleviate perioperative ALI, and improve early prognosis in the patients undergoing AAAD surgery.

Mechanism of Shenqi Yiliu Prescription Combined with Cisplatin on H22 Liver Cancer-bearing Mice Based on NLRP3/Caspase-1/GSDMD Pyroptosis Pathway / 中国实验方剂学杂志

ObjectiveTo explore the anti-tumor effect and mechanism of Shenqi Yiliu prescription in the intervention of pyroptosis. MethodTen male BALB/c mice were randomly selected and assigned to the blank group. The remaining 40 mice underwent the induction of the liver cancer xenograft model. After 5 days of modeling， 40 surviving mice were randomly divided into model group， cisplatin group ［2.5×10-3 g·kg-1·（3 d）-1］， Shenqi Yiliu prescription group （27 g·kg-1·d-1）， and a combination group （Shenqi Yiliu prescription group + cisplatin）. The mice in the blank group and the model group were treated with an equal volume of normal saline for 10 days. The general conditions of mice in each group were observed. After the intervention， the tumor weight of the mice was weighed and the tumor inhibition rate was calculated. Hematoxylin-eosin （HE） staining was used to observe the pathological changes in tumor tissues. The levels of mouse liver function indicators， including alanine aminotransferase （ALT） and aspartate aminotransferase （AST） were detected. The TdT-mediated dUTP-biotin nick end labeling （TUNEL） assay was used to detect DNA damage in mouse tumor tissue cells. Immunohistochemistry （IHC）， immunofluorescence （IF）， and Western blot were used to detect the protein expression levels of NOD-like receptor protein 3 （NLRP3）， cysteinyl aspartate-specific protease-1 （Caspase-1）， and gasdermin D （GSDMD） in tumor tissues. The levels of interleukin-1β （IL-1β） and interleukin-18 （IL-18） in tumor tissues were detected by enzyme-linked immunosorbent assay （ELISA）. ResultCompared with the mice in the blank group， those in the model group were in a poor mental state， sleepy， and lazy， and their fur color was dull， with increased levels of serum ALT and AST in liver function tests （P<0.01）. Compared with the model group， the groups with drug intervention showed improved mental state， inhibited tumor growth to varying degrees， and decreased tumor weight， and the tumor inhibition rate in the combination group was the highest （P<0.01）. HE staining showed that the pathological and morphological lesions of the tumor tissues in the model group were significant， while those in all groups with drug intervention were improved to a certain extent. The karyolysis and nuclear rupture in the Shenqi Yiliu prescription group and the combination group were more significant. In the liver function test， the serum ALT and AST levels of mice in the Shenqi Yiliu prescription group and the combination group decreased （P<0.01）， and the inflammatory factors IL-1β and IL-18 in each group with drug intervention decreased （P<0.05， P<0.01）. Among them， the declining trend of IL-1β and IL-18 in the Shenqi Yiliu prescription group was the most significant （P<0.01）. TUNEL staining showed that the positive TUNEL staining in each group with drug intervention decreased after intervention （P<0.05， P<0.01）， especially the cisplatin group and Shenqi Yiliu prescription group （P<0.01）. Western blot， IHC， and IF found that the protein expression levels of NLRP3， Caspase-1， and GSDMD in each group with drug intervention decreased （P<0.05， P<0.01）. Compared with the mice in the cisplatin group， those in the Shenqi Yiliu prescription group and the combination group had better mental state and regular tumor morphology， and the tumor weight of the mice in the combination group decreased （P<0.05）. The levels of ALT and AST in the Shenqi Yiliu prescription group decreased （P<0.05）， and the levels of IL-1β and IL-18 in the Shenqi Yiliu prescription group and the combination group decreased （P<0.05， P<0.01）， especially in the combination group （P<0.01）. The results of IHC showed that the expression of GSDMD protein in the tumor tissues of mice in the combination group was reduced （P<0.01）. IF detection showed that the expression of NLRP3 in the tumor tissues of the Shenqi Yiliu prescription group was reduced （P<0.01）. The results of Western blot showed that the expression level of NLRP3 protein in the Shenqi Yiliu prescription group and the combination group decreased （P<0.01）， and the expression level of Caspase-1 protein in the combination group decreased （P<0.01）. The decrease in GSDMD protein expression was not significant， and the difference was not statistically significant. ConclusionShenqi Yiliu prescription combined with cisplatin has an obvious anti-tumor effect， which may be achieved by down-regulating the NLRP3/Caspase-1/GSDMD inflammatory pyroptosis pathway to inhibit cell pyroptosis， and relieve the inflammatory response in mice with liver cancer.

Research progress on islet protection and functional maintenance / 器官移植

Islet transplantation is a promising treatment of diabetes mellitus and its complications. Nevertheless, dysfunction post-transplantation, rejection and shortage of donors are the bottleneck issues in the field of islet transplantation. Optimizing the preservation method of pancreas plays a positive role in obtaining a sufficient quantity of effective islets and maintaining their functions. During the culture stage, anti-rejection and anti-apoptosis treatment of islets, including mesenchymal stem cell (MSC), MSC-derived exosomes, anti-apoptosis drugs and gene modification, may become major approaches for islet protection and functional maintenance in clinical islet transplantation. Use of anti-instant blood-mediated inflammatory reaction (IBMIR) drugs after islet transplantation also plays a critical role in protecting islet function. In this article, the whole process from islet preparation to islet transplantation was illustrated, and relevant strategies of islet protection and functional maintenance were reviewed, aiming to provide reference for improving the quality of donors to compensate for the shortage of absolute quantity of donors and elevating the efficiency of islet transplantation.

Enzymatic Digestion Method Coupled with Artificial Intelligence Techniques in Forensic Drowning Diatom Detection / 中山大学学报(医学科学版)

ObjectiveArtificial intelligence （AI） full smear automated diatom detection technology can perform forensic pathology drowning diatom detection more quickly and efficiently than human experts.However， this technique was only used in conjunction with the strong acid digestion method， which has a low extraction rate of diatoms. In this study， we propose to use the more efficient proteinase K tissue digestion method （hereinafter referred to as enzyme digestion method） as a diatom extraction method to investigate the generalization ability and feasibility of this technique in other diatom extraction methods. MethodsLung tissues from 6 drowned cadavers were collected for proteinase K ablation and made into smears， and the smears were digitized using the digital image matrix cutting method and a diatom and background database was established accordingly.The data set was divided into training set， validation set and test set in the ratio of 3：1：1， and the convolutional neural network （CNN） models were trained， internally validated， and externally tested on the basis of ImageNet pre-training. ResultsThe results showed that the accuracy rate of the external test of the best model was 97.65 %， and the area where the model features were extracted was the area where the diatoms were located. The best CNN model in practice had a precision of more than 80 % for diatom detection of drowned corpses. ConclusionIt is shown that the AI automated diatom detection technique based on CNN model and enzymatic digestion method in combination can efficiently identify diatoms and can be used as an auxiliary method for diatom detection in drowning identification.

The role of enzymes in the angiogenic activity of Hancornia speciosa latex

The Hancornia speciosa latex has shown angiogenic activity. Angiogenesis plays a major role in wound healing, and materials that stimulate this process could be used to develop drugs. This study aimed to explain the role of proteins in the H. speciosa serum fraction latex in angiogenesis. Hence, this material was treated with proteinase K and the proteins were inactivated. After protein inactivation, angiogenic activity was assessed with the chick chorioallantoic membrane assay. The result showed that the proteins in the serum fraction are responsible for angiogenic activity. Then, the total protein content in the serum fraction and its enzymatic activity were investigated. The low protein content observed in the H. speciosa serum fraction latex suggests that this biomaterial could be used to develop new drugs with a hypoallergenic response. Despite the low protein content, there was a significant enzymatic activity of at least three enzymes in the serum fraction latex: ß-1,3 glucanase, ß-glucosidase, and proteases. These enzymes seem to influence the healing process, assisting debridement, extracellular matrix remodeling, and collagen deposition, and decreasing the chances of contamination by microorganisms. In conclusion, the enzymes in the H. speciosa serum latex are associated with the angiogenic activity of this biomaterial and may be used to assist the wound healing process.

Research process on pathogenesis of gastroesophageal reflux disease / 中国医师杂志

Gastroesophageal reflux disease (GERD) is a gastrointestinal motility disorder that results from the reflux of stomach contents into the esophagus or oral cavity, causing symptoms or complications. The typical symptoms of GERD are heartburn and regurgitation of gastric contents into the oropharynx. Heartburn is the sensation of burning or discomfort behind the sternum. Heartburn may radiate into the neck, is typically worse after meals or when in a reclining position, and may be eased by antacids. Regurgitation is the backflow of gastric contents into the mouth or hypopharynx. Epigastric pain can also be a symptom of GERD. Extraesophageal symptoms of GERD include dental erosions, laryngitis, cough, and asthma. In recent years, great progress has been made in understanding the molecular basis of GERD, suggesting that its pathogenesis is more complex and multifactorial. In this paper, the molecular pathogenesis was taken as the starting point, including the mechanism of genes in the pathogenesis and development of GERD, the mechanism of NF-κB pathway in the pathogenesis and development of GERD, the role of proteinase-activated receptor-2 in the pathogenesis of GERD, the association between abnormal serotonin pathway and GERD, and the relationship between reactive oxygen species and GERD, to summarize the pathogenesis of gastroesophageal reflux disease.

Mechanism of Tingli Dazao Xiefeitang on Cell Pyrolysis and Apoptosis Induced by Caspase-1 in A549 Cells / 中国实验方剂学杂志

ObjectiveTo explore the mechanism of coking death and apoptosis of A549 cells induced by Tingli Dazao Xiefeitang. MethodA549 cells were randomized into blank group， traditional Chinese medicine（TCM） low， medium， and high concentration groups， which were treated with 20， 40， 60 mg·L-1 Tingli Dazao Xiefeitang， and TCM low， medium， and high concentration groups， respectively， and blank group was treated with equal volume culture medium. After 48 h of treatment， cell migration was detected by scratch assay and cell apoptosis was detected by flow cytometry. The relative expression levels of cysteine aspartate protease-1（Caspase-1）， NOD-like receptor protein 3 （NLRP3）， dermoderin D （GSDMD）， Survivin protein and nuclear transcription factor -κB （NF-κB） pathway proteins were detected by Western blot. The levels of intracellular reactive oxygen species （ROS） were determined by DCFH-DA fluorescence probe， and the contents of tumor necrosis factor -β （TNF-β） and interleukin-1β （IL-1β） in supernatant were determined by enzyme-linked immunosorbent assay （ELISA）. ResultCompared with blank group， the scratch healing rate， apoptosis rate， relative expression of Survivin protein， Caspase-1， GSDMD， NLRP3， ROS and NF-κB phosphorylation levels were significantly increased in low， medium and high concentration groups. The contents of TNF-β and IL-1β in supernatant were significantly increased （P<0.05）. Compared with the low concentration group， the scratch healing rate， apoptosis rate， Survivin protein relative expression， Caspase-1， GSDMD， NLRP3 relative expression， ROS and NF-κB phosphorylation levels were significantly increased in the medium and high concentration groups. The contents of TNF-β and IL-1β in supernatant were significantly increased （P<0.05）. Compared with the TCM group， the scratch healing rate， apoptosis rate， Survivin protein relative expression， Caspase-1， GSDMD， NLRP3 relative expression， ROS and NF-κB phosphorylation levels were significantly increased in the high concentration group. The contents of TNF-β and IL-1β in supernatant were significantly increased （P<0.05）. ConclusionTingli Dazao Xiefeitang can improve NLRP3 protein expression， inhibit Survivin protein expression and promote apoptosis of A549 cells. At the same time， it can activate NF-κB pathway and ROS system， up-regulate the expression of Caspase-1 and GSDMD， mediate scortosis of A549 cells.

Correlation between serum cystatin C level and clinicopathological features of IgA nephropathy / 中华检验医学杂志

Objective:To investigate the correlation between serum cystatin C (CysC) and clinical and pathological features of IgA nephropathy.Methods:Four hundred and twenty-one cases of primary IgA nephropathy diagnosed by renal biopsy in Longhua Hospital Affiliated to Shanghai University of Traditional Chinese Medicine from January 2010 to January 2021 were retrospectively analyzed. According to the serum CysC level at the time of renal biopsy, the patients were divided into high serum CysC group and normal serum CysC group, and the clinical data and pathological indices of the patients were compared. Spearman correlation analysis was used to analyze the correlation between estimated glomerular filtration rate (eGFR) and serum CysC. The clinicopathological factors related to the serum CysC level were analyzed by multiple linear regression. The area under the receiver operator characteristic curve (AUC) was used to evaluate the ability of serum CysC level to predict related pathological injury.Results:The age, prevalence of hypertension, serum creatinine, urea and uric acid levels of high serum CysC group were significantly higher than those of normal serum CysC group, while the eGFR level was significantly lower than that of normal serum CysC group ( P<0.05). Spearman correlation analysis showed that serum CysC was negatively correlated with eGFR ( r=-0.744, P<0.001). In terms of pathological injury, the degree of renal tubular atrophy and renal interstitial fibrosis (T) and renal arteriole wall thickening (A) in high serum CysC group were more serious than those in normal serum CysC group ( P<0.05). Multiple linear regression analysis showed that the prevalence of hypertension, serum creatinine, urea, uric acid, T and A were correlated with serum CysC levels (standard regression coefficient β=0.048, 0.299, 0.260, 0.134, 0.195, 0.068, respectively, P<0.05). After adding serum CysC on the basis of clinical features, the prediction efficiency of renal tubular atrophy and renal interstitial fibrosis was higher (AUC were 0.829 [95% CI 0.787-0.870], 0.847 [95% CI 0.808-0.886], P<0.05). Conclusions:Patients with older age, hypertension, poor renal function and severe pathological damage are more likely to have elevated serum CysC levels. Serum CysC was related to the prevalence of hypertension, creatinine, urea, uric acid, T and A. Combined with serum CysC level can effectively improve the ability prediction of T.

Research progress in the relationship between deubiquitinase OTUB1 and urological cancer / 国际生物医学工程杂志

Ovarian tumor-associated protease B1(OTUB1) is a member of the deubiquitinase family. Its highly specific recognition and cleavage function of polyubiquitinated chains has attracted widespread attention, and it can regulate a variety of important signaling pathways, such as the epithelial-mesenchymal transition (EMT) pathway, the MAPK signaling pathway, and the p53-related signaling pathway. In recent years, it has gradually become a new direction of oncology research. More and more studies have proved that OTUB1 is closely related to various tumors such as hepatocellular carcinoma, colorectal cancer, and breast cancer. It regulates the occurrence, development, and prognosis of tumors. OTUB1 could be a potential treatment for tumors. Urinary tract tumors mainly include prostate cancer, bladder cancer, and renal cell carcinoma. In this review, the research progress on the correlation between OTUB1 and urological tumors was reviewed, including its important role in the occurrence and development of urological tumors and the possibility of treating urological tumors with OTUB1.

Effect of HMGB1/Caspase-1/GSDMD signaling axis-mediated hepatocyte pyroptosis on liver ischemia-reperfusion injury / 器官移植

Objective To evaluate the effect of high mobility group box 1 (HMGB1)/ cysteinyl aspartate specific proteinase (Caspase)-1/Gasdermin D (GSDMD) signaling axis-mediated hepatocyte pyroptosis on liver ischemia-reperfusion injury (IRI). Methods C57BL/6 mice were randomly divided into the sham operation group (Sham group), IRI 2 h group, IRI 6 h group, IRI 12 h group, glycyrrhizic acid (GA)+Sham group and GA+IRI 12 h group (n=8 in each group). AML12 cells were evenly divided into the Sham group, IRI 12 h group, GA+Sham group and GA+IRI 12 h group. The serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), interleukin (IL)-1β and IL-6 in each group were detected by enzyme-linked immune absorbent assay(ELISA). The messenger ribonucleic acid (mRNA) levels of IL-1β and IL-6 were detected by reverse transcription polymerase chain reaction(RT-PCR). The pathological score of liver ischemia and cell apoptosis were compared among all groups. The expression level of HMGB1 in the liver tissues of each group was determined by immunohistochemistry. The expression levels of HMGB1, Caspase-1 and GSDMD proteins in the mouse liver tissues and AML12 cells were measured by Western blot. Results Compared with the Sham group, the serum levels of ALT, AST, IL-1β and IL-6 and the relative expression levels of IL-1β and IL-6 mRNA in the liver tissues were all significantly up-regulated after IRI in each group (all P < 0.05), and showed significant time-dependent pattern along with the prolongation of reperfusion time. Compared with the Sham group, the pathological score of hepatic ischemia and the apoptosis rate of hepatocytes were significantly increased after IRI in each group (all P < 0.05). Immunohistochemical results showed that the expression level of HMGB1 in the liver tissues was significantly up-regulated after IRI, which showed an increasing trend along with the prolongation within the period of 2-12 h. Western blot showed that compared with the Sham group, the relative expression levels of HMGB1, Caspase-1 and GSDMD proteins in vivo and in vitro were up-regulated in the IRI 12 h group. The relative expression level of HMGB1 protein was significantly up-regulated, whereas those of Caspase-1 and GSDMD proteins were significantly down-regulated in the GA+IRI 12 h group compared with those in the IRI 12 h group (all P < 0.05). Conclusions Hepatocytes probably activate the Caspase-1/GSDMD signaling pathway by releasing HMGB1, thereby triggering hepatocyte pyroptosis and leading to liver IRI. Inhibition of extracellular release of HMGB1 by GA may mitigate liver IRI.