Mechanism of BCL2L2-PABPN1 expression induced by sodium arsenite and its metabolites in 16HBE cells / 中国职业医学

@#Objective - - （BCL2L2）- （ ） To investigate the differential expression of the fusion gene BCL 2 like protein 2 poly A （PABPN1） （ ） binding protein nuclear 1 induced by sodium arsenite SA and its methylated metabolites in 16HBE cells and the Methods ） ， related mechanism. i The 16HBE cells exposed to SA at concentrations of 1.5 3.0 and 4.5 µmol/L were set as -， - - low medium and high dose arsenic exposure groups. The 16HBE cells exposed to 4.5 µmol/L monomethylarsonic acid （ ）， （ ） ， MMA dimethylarsonic acid DMA and SA were set as MMA group DMA group and SA group. The 16HBE cells without ， BCL2L2-PABPN1 toxic stimulation were set as control group. After the cells were cultured for 48 hours the expression of was - （ - ） ） （ ） detected by quantitative real time polymerase chain reaction qRT PCR . ii Two small interfering RNA siRNA silencing 基金项目：国家自然科学基金（ ）； 年云南省科技厅昆明医科大学应用基础研究联合专项面上项目 82160607 2021 （ ） 202101AY070001-054 作者简介：施雅（ —），女，在读大学本科生，主要从事劳动卫生与环境卫生学研究；尹锦瑶（ —），女，在读劳动卫生与环境卫 2001 1995 生学硕士研究生，主要从事劳动卫生与环境卫生学研究；施雅和尹锦瑶为共同第一作者 通讯作者：何越峰教授，博士研究生导师，- ： E mail heyuefeng@kmmu.edu.cn中国职业医学 年 月第 卷第 期 ， ， ， · · 2022 10 49 5 Chin Occup Med October 2022 Vol.49 No.5 523 BCL2L2-PABPN1， - fragments were designed and transfected into 16HBE cells to knockdown which were set as siRNA 1 group - - BCL2L2-PABPN1 and siRNA 2 group. Non transfected control group without knockdown of transfection was set up. After ， BCL2L2-PABPN1 - culturing for 48 hours the expression level of in the three groups of cells was detected by qRT PCR. The cell - survival rate and early apoptosis rate were detected by MTS method and JC 1 mitochondrial membrane potential detection ， （ ） ， method respectively. The apoptosis was detected by Hoechest33342/propidium iodide PI double staining and the expression - Results ） level of P53 signaling pathway related proteins was detected by Western blotting. i The relative expression of BCL2L2-PABPN1 （P ） BCL2L2- in 16HBE cells increased with the increasing SA doses <0.01 . The relative expression of PABPN1 - ， - - in high dose arsenic exposure was higher than that in control group low dose and medium dose arsenic exposure （ P ） BCL2L2-PABPN1 ， groups all <0.05 . The relative expression of in SA group was higher than those in control group MMA （ P ） BCL2L2-PABPN1 group and DMA group all <0.05 . The relative expression of showed no significant difference between ， （ P ） ） BCL2L2-PABPN1 control group MMA group and DMA group all >0.05 . ii The relative expression levels of and cell - - - （ P ） survival rate in siRNA 1 group and siRNA 2 group were lower than those in non transfected control group all <0.05 . ， （P ） However there was no significant difference in the early apoptosis rate among the three groups >0.05 . The results of - Hoechest33342/PI double staining showed that the number of nuclear shrinkage and early apoptotic cells in siRNA 1 group and - - ， - siRNA 2 group was higher than that in non transfected control group. The relative protein expression levels of P53 phospho ， - - ， - - （ P ） p53 BCL 2 associated death promoter P21 and cytochrome C in siRNA 1 group and siRNA 2 group were higher all <0.05 , - - P and the relative protein expression levels of P53 up regulated modulator of apoptosis were lower (all <0.05), when compared - Conclusion with the non transfected control group. SA may block the apoptosis of 16HBE cells by inducing the expression of BCL2L2-PABPN1 fusion gene . The mechanism may be related to the activation of P53 signaling pathway. The SA methylated BCL2L2-PABPN1 BCL2L2-PABPN1 - metabolites MMD and DMA had no effect on the expression of . may affect anti apoptosis BCL2L2 PABPN1 through affecting the synergistic effect of and genes.

Clinical application effects of two longitudes three transverses method in perforator location of thoracodorsal artery perforator flap and deep wound repair / 中华烧伤杂志

Objective: To explore the clinical application value of two longitudes three transverses method in the location of the perforator of thoracodorsal artery perforator and deep wound repair. Methods: The retrospectively observational study was conducted. From December 2018 to June 2020, 17 patients with deep wounds who were admitted to the Affiliated Hospital of Zunyi Medical University met the inclusion criteria and were included in this study, including 7 males and 10 females, aged 12 to 72 years. The wound areas of patients after debridement were 7 cm×3 cm to 11 cm×7 cm. Two longitudinal lines were located through the midpoint of the armpit, the posterior superior iliac spine, and the protruding point of the sacroiliac joint, and three transverse lines were located 5, 10, and 15 cm below the midpoint of the armpit between the two longitudinal lines, i.e. two longitudes three transverses method, resulting in two trapezoidal areas. And then the thoracodorsal artery perforators in two trapezoidal areas were explored by the portable Doppler blood flow detector. On this account, a single or lobulated free thoracodorsal artery perforator flap or flap that carrying partial latissimus dorsi muscle, with an area of 7 cm×4 cm to 12 cm×8 cm was designed and harvested to repair the wound. The donor sites were all closed by suturing directly. The number and location of thoracodorsal artery perforators, and the distance from the position where the first perforator (the perforator closest to the axillary apex) exits the muscle to the lateral border of the latissimus dorsi in preoperative localization and intraoperative exploration, the diameter of thoracodorsal artery perforator measured during operation, and the flap types were recorded. The survivals of flaps and appearances of donor sites were followed up. Results: The number and location of thoracodorsal artery perforators located before operation in each patient were consistent with the results of intraoperative exploration. A total of 42 perforators were found in two trapezoidal areas, with 2 or 3 perforators each patient. The perforators were all located in two trapezoid areas, and a stable perforator (the first perforator) was located and detected in the first trapezoidal area. There were averagely 1.47 perforators in the second trapezoidal area. The position where the first perforator exits the muscle was 2.1-3.1 cm away from the lateral border of the latissimus dorsi. The diameters of thoracodorsal artery perforators were 0.4-0.6 mm. In this group, 12 cases were repaired with single thoracodorsal artery perforator flap, 3 cases with lobulated thoracodorsal artery perforator flap, and 2 cases with thoracodorsal artery perforator flap carrying partial latissimus dorsi muscle. The patients were followed up for 6 to 16 months. All the 17 flaps survived with good elasticity, blood circulation, and soft texture. Only linear scar was left in the donor area. Conclusions: The two longitudes three transverses method is helpful to locate the perforator of thoracodorsal artery perforator flap. The method is simple and reliable. The thoracodorsal artery perforator flap designed and harvested based on this method has good clinical effects in repairing deep wound, with minimal donor site damage.

Effects of sodium arsenite and arsenic metabolites on the expression of linear and circular RNA of Nup107 in A549 cells / 中国职业医学

OBJECTIVE: To explore the effect of sodium arsenite and arsenic metabolites monomethylarsenic acid(MMA) and dimethylarsenic acid(DMA)on the expression of linear and circularRNAs of nucleoporin 107(Nup107) in human lung adenocarcinoma A549 cells. METHODS: i) The A549 cells in logarithmic phase were treated with 0, 30, 60, 90 μmol/L sodium arsenite for 48 hours. ii)The A549 cells in logarithmic phase were treated with 90 μmol/L sodium arsenite, MMA and DMA for 48 hours, the control group received no treatment. After culturing, the relative expression of linear RNA and circular RNA(circRNA) of Nup107 was detected by real-time quantitative polymerase chain reaction. RESULTS: i) The relative expression of linear RNA of Nup107 decreased and four circRNA isomers such as hsa＿circ＿0003599, hsa＿circ＿0027477, hsa＿circ＿0027478 and hsa＿circ＿0027479 increased with the increase of sodium arsenite dose(all P<0.01), showing a dose-effect relationship. In the 90 μmol/L sodium arsenite stimulated group, the relative expression of linear RNA of Lin-Nup107 decreased, and the four circRNA isomers increased compared with the control group in the A549 cells(all P<0.01). ii) The relative expression of Lin-Nup107 increased in the MMA and DMA stimulated groups and decreased in the sodium arsenite stimulated group compared with the control group in the A549 cells(all P<0.05). The relative expression of Lin-Nup107 decreased in the sodium arsenite stimulated group compared with the MMA and DMA stimulated groups in the A549 cells(all P<0.05). The relative expressions of hsa＿circ＿0003599, hsa＿circ＿0027478, hsa＿circ＿0027479 in the sodium arsenite stimulated group were higher than that in the MMA and DMA stimulated groups as well as the control group(all P<0.05).The relative expressions of hsa＿circ＿0027479 in the DMA stimulated group was lower than that in the control group(P<0.05). CONCLUSION: Sodium arsenite can induce the down-regulation of linear RNA and the up-regulation of circRNA of Nup107 in a dose-dependent manner. The metabolites of arsenic MMA and DMA can induce the overexpression of linear RNA of Nup107, however, they had no obvious effect on the circRNA of Nup107 in A549 cells.

Study on Dose-effect Relationship and Mechanism of Protective Effects of Total Asiaticoside on Gastrointestinal Motility and Enteric Nervous System in Aged Functional Dyspepsia Model Rats / 中国药房

OBJECTIVE：To explo re the dose-effect relationship and mechanism of protective effects of total asiaticoside （TA） on gastrointestinal motility and enteric nervous system （ENS）in aged functional dyspepsia （FD）model rats. METHODS ：Aged male SD rats of 16 months old were randomly divided into blank control group ，model group ，TA low dose ，medium dose and high dose groups （15，30，60 mg/kg），with 8 rats in each group. FD model was established by tail-stimulation combined with irregular diet for 4 weeks. The next day after modeling ，administration groups were given relevant doses of TA solution intragastrically ； control group and model group were given constant volume of normal saline intragastrically ，once a day ，for consecutive 15 d. Gastric emptying rate and small intestinal propulsion rate of rats were examined. ELISA were used to detect serum contents of MTL and VIP. Immunofluorescence and immunohistochemistry were proposed to measure the expression of ENS marker （S100β and GDNF）in gastric antrum tissue. The protein expression of S 100β，GFAP，PGP9.5，GDNF，p-MEK and p-ERK 1/2 in gastric antrum tissue were measured by Western blotting assay. RESULTS ：Compared with blank control group ，gastric emptying rate and small intestinal propulsion rate ，serum MTL content and protein expression of PGP 9.5 in gastric antrum tissue of model and TA low，medium dose group were decreased significantly ，while serum VIP content ，protein expressions of S 100β，GFAP，GDNF， p-MEK and p-ERK 1/2 in gastric tissue were increased significantly （P＜0.05）. Compared with model group ，gastric emptying rate and small intestinal propulsion rate of TA groups were increased significantly （P＜0.05）；except for GFAP protein in TA low dose group（P＞0.05），the serum MTL content and the expression of PGP 9.5 protein in gastric antrum tissue of rats in TA groups were increased significantly ，while serum VIP content ，protein expression of S 100β，GFAP，GDNF，p-MEK and p-ERK 1/2 in gastric antrum tissue were decreased significantly （P＜0.05）. Some or most of the content of gastrointestinal motility indexes and related factor protein expression were significantly different among TA groups （P＜0.05），and the indexes in TA high dose group could recover to the levels which were not significantly different with blank control group （P＞0.05）. CONCLUSIONS ：TA can dose-dependently improve the gastrointestinal motility deficiency and ENS dysfunction in aged FD model rats ，especially in high dose（60 mg/kg）of TA group. Its mechanism may be related with promoting the release of endogenous MTL ，inhibiting the secretion of VIP ，expression of GDNF and the activation of downstream signaling pathway ，and promoting the repair of ENS and intestinal neurons.

Expression of IFN-γ and IL-10 in peripheral blood and intestinal mucosa of patients with diarrhea post-infectious irritable bowel syndrome / 重庆医学

Objective To investigate the expression change of cytokines in peripheral blood and intestinal mucosa in the patients with diarrhea post-infectious irritable bowel syndrome(PI-IBS) and its relation with clinical symptoms scores.Methods Thirty outpatients and inpatients with diarrhea PI-IBS(observation group) and contemporaneous 30 individuals undergoing physical examination(control group) in the Hainan Provincial People's Hospital from January to December 2013 were selected.The peripheral blood mononuclear cells(PBMC) were separated and cultured.Then the levels of IFN-y and IL-10 in peripheral blood and cell culture supernatant fluid were detected by ELISA.The colonic mucosal tissue was taken by coloscopy.Then colonic mucosal IFN-γ and IL-10 protein expression was detected by immunohistochemistry staining.Furthermore,the correlationship between the level change of IFN-γ and IL-10 with clinical symptom score was analyzed by using the Spearman correlation method.Results Peripheral blod IL-10 and IFN-γ levels had no statistical difference between the two groups(P＞0.05).Compared with the control group,in PBMC seperation and cuture,the IFN-γ level in the observation group was increased and IL-10 level was decreased,the difference was statistically signifieant(P＜0.01).The intestinal main symptom score in the observation group had the positive correlation with IFN-γ expression level of PBMC culture supernatant fluid and colonic mucosal IFN-γ expression level(r=0.45,0.94,P＜0.01),and had the negative correlation with IL-10 expression level(r=-0.52,-0.79,P＜0.01).Conclusion The unbalance of IFN-γ and IL-10 level could be involved in the pathogenesis of diarrhea PI-IBS,which can serve as the observation indicators of disease activity.

Clinical Features and Expression of Cytokine in Post-infectious Irritable Bowel Syndrome Patients / 中山大学学报(医学科学版)

[Objective]To study the clinical features and expressions of IL-17A,IFN-γ,and IL-10 in serum and intestinal mucosa of patients with post-infectious irritable bowel syndrome and non post-infectious irritable bowel syndrome.[Methods]44 diar?rhea-predominate IBS patients(21 with PI-IBS,23 with NPI-IBS)and 10 healthy controls were recruited in this study. Investigation questionnaires of GSRS,SAS,SDS were carried out to evaluate the gastrointestinal function,anxiety status and depression status of IBS patients. The expressions of IL-17A,IFN-γ,and IL-10 in intestinal mucosa and serum were measured by immunohistochemis?try(IHC)and enzyme-linked immunosorbent assay(ELISA).[Results]The SDS scores of NPI-IBS patients were higher than those of controls(P 0.05).[Conclusion]PI-IBS and NPI-IBS patients existed various anxiety and depression. The levels of IL-17A and IFN-γ increased and level of IL-10 decreased in PI-IBS and NPI-IBS group. But the clinical symptoms and changes of cytokines of PI-IBS patients were more significant. There may exist other pathogenesis in PI-IBS but not in NPI-IBS.

Changes of Peripheral Blood Marrow-Derived Suppressor Cell Level after Chemotherapy Induction Remission by VDLP Regimen and Their Relationship with Immune System in B-ALL Children / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the changes of peripheral blood marrow-derived suppressor cell level after chemotherapy induction remission by regimen consisting of vincristine, daunorubicin, L-asparaginase and prednisone (VDLP) and to analyze their relationship with immume system in B-ALL children.</p><p><b>METHODS</b>Thirty B-ALL children after induction remission by VDLP regimen from August 2015 to August 2016 were selected as B-ALL group and 30 normal healthy children were selected as control group. The peripheral blood in 2 groups was collected and detected by flow cytometry, then the ratios of CD30cells and CD33HLA-DRmarrow-derived suppressor cells, CD14CD33HLA-DRmarrow-derived suppressor cells and CD15CD33HLA-DRmarrow-derived suppressor cells were calculated, and their changes after induction remission by VDLP regimen and the relationship with immune system were analyzed.</p><p><b>RESULTS</b>After treatment the ratio of CD33cells in peripheral blood of B-ALL group and control group was not significantly different (P> 0.05), moreover, the ratio of CD33cells in B-ALL group was significantly higher than that before treatment (P<0.05), while the ratios of CD33HLA-DRmarrow-derived suppressor cells, CD14CD33HLA-DRmarrow-derived suppressor cells and CD15CD33HLA-DRmarrow-derived suppressor cells in B-ALL group were significantly lower than those in control group (all P<0.05), but the ratios of these cells in B-ALL group were higher than those before treatment, and yet there was no statistical significance (P>0.05).</p><p><b>CONCLUSION</b>The ratios of marrow-derived suppressor cells in peripheral blood of B-ALL children in complete remission after treatment with VDLP regimen are higher than those before treatment, but are significantly lower than normal value, which may be related with non-complese recovery of immune system in B-ALL children after treatment.</p>

Study on phenotype and function of intestinal gamma delta T lymphocytes in mice with post-infectious irritable bowel syndrome / 中国免疫学杂志

Objective:To investigate the phenotype and function of the intestinal γδT lymphocytes in post-infectious irritable bowel syndrome mouse model. Methods:The mouse model for post-infectious irritable bowel syndrome was established by the infection with trichinella spiralis. The intestinal inflammation,abdominal withdrawal reflex( AWR) and colon transportation test were observed. 2 and 8 weeks later,the animals were sacrificed and the lymphocytes in the intestinal lymph nodes and spleen were collected,from which the γδT lymphocytes were isolated and purified by monoclonal antibody-immuno-microbeads method. The functions of the purified γδT lymphocytes were evaluated,including proliferation by 3 HTdR;CD69,CD62L molecule staining by flow cytometry. Furthermore,the con-centration of cytokine IL-17 and IFN-γ in the supernatant of the cultured γδT lymphocytes were detected by ELISA. Results: At 2nd weeks after infection,significant intestinal inflammation was observed,with increasingγδT lymphocytes,proliferating and activating with increasing production of IL-17. At 8th weeks after infection, the intestinal inflammation disappeared, whereas the number of γδT lymphocytes remained increasing,also with proliferating and activating with increasing production of IL-17. Meanwhile,the mice show higher AWR score and Bristol score. Conclusion: γδT lymphocytes could participate in the pathogenesis of PI-IBS via their proliferation,activation and production of IL-17.

Preinduced intestinal HSP70 improves visceral hypersensitivity and abnormal intestinal motility in PI-IBS mouse model

OBJECTIVE@#To investigate the impact of the preinduced intestinal heat shock protein 70 (HSP70) on the visceral hypersensitivity and abnormal intestinal motility in a post-infectious irritable bowel syndrome (PI-IBS) mouse model.@*METHODS@#Eighty-four female C57BL/6 mice were randomly assigned to four groups: control group (n = 21) and induction + PI-IBS group (n = 21), PI-IBS group (n = 21) and induction group (n = 21). The mice in PI-IBS group were infected in vivo with Trichinella spiralis by oral administration. The visceral hypersensitivity and intestinal motility were evaluated respectively with abdominal withdrawal reflex and colon transportation test. The intestinal HSP70 protein and mRNA level was measured by Western blot and real-time PCR. Meanwhile, the intestinal proinflammatory cytokines IL-10 and TNF-α level was detected by ELISA.@*RESULTS@#Compared with their counterparts in PI-IBS group, the animals in the Induction + PI-IBS group show significantly increased intestinal level of HSP70 and obviously ameliorative clinical figures, including abdominal withdrawal reflex score, intestine transportation time and Bristol scores (P < 0.05). Meanwhile, the intestinal post-inflammatory cytokines remarkably changed, including increased IL-10 level and decreased TNF-α level (P < 0.05).@*CONCLUSIONS@#Intestinal HSP70 may play a potential protective role through improving the imbalance between the intestinal post-inflammatory and anti-inflammatory cytokines in PI-IBS.

Preinduced intestinal HSP70 improves visceral hypersensitivity and abnormal intestinal motility in PI-IBS mouse model

Objective: To investigate the impact of the preinduced intestinal heat shock protein 70 (HSP70) on the visceral hypersensitivity and abnormal intestinal motility in a post-infectious irritable bowel syndrome (PI-IBS) mouse model. Methods: Eighty-four female C57BL/6 mice were randomly assigned to four groups: control group (n = 21) and induction + PI-IBS group (n = 21), PI-IBS group (n = 21) and induction group (n = 21). The mice in PI-IBS group were infected in vivo with Trichinella spiralis by oral administration. The visceral hypersensitivity and intestinal motility were evaluated respectively with abdominal withdrawal reflex and colon transportation test. The intestinal HSP70 protein and mRNA level was measured by Western blot and real-time PCR. Meanwhile, the intestinal proinflammatory cytokines IL-10 and TNF-α level was detected by ELISA. Results: Compared with their counterparts in PI-IBS group, the animals in the Induction + PI-IBS group show significantly increased intestinal level of HSP70 and obviously ameliorative clinical figures, including abdominal withdrawal reflex score, intestine transportation time and Bristol scores (P < 0.05). Meanwhile, the intestinal post-inflammatory cytokines remarkably changed, including increased IL-10 level and decreased TNF-α level (P < 0.05). Conclusions: Intestinal HSP70 may play a potential protective role through improving the imbalance between the intestinal post-inflammatory and anti-inflammatory cytokines in PI-IBS.

Available value of semi-quantitative scoring system for contrast-enhanced ultrasound quantitative analysis's color images in the differential diagnosis of breast nodules / 中华超声影像学杂志

Objective To assess the feasibility of semi-quantitative scoring system for contrast-enhanced ultrasound (CEUS)quantitative analysis's color images in the differential diagnosis of breast nodules.Methods Totally 244 BI-RADS 4 breast solid lesions received CEUS before core needle biopsy or surgical resection were included.A semi-quantitative scoring system for color images of CEUS quantitative analysis were built.The scores were given as follows:1 )Color type and its distribution (0 to 4);2)Color scope (0 to 1 );3)Color margin (0 to 1 );4)Color shape (0 to1 ).The total score for each lesion would be from 0 to 7.And the differenital value between benign and malignant lesions were assessed.Results The total semi-quantitative scores of 102 malignant tumors (5.1 ±1 .7)was significant higher than that of benign lesions (3.34±0.7)(P < 0.05 ).In 102 malignant lesions,the total scores of 81 lesions (79.41 %)were more than 4 points,and in 142 benign lesions,the total scores of 89 lesions (62.67%)were less than 4 points.Depending on the Wilcox rank sum test (Mann-Whitney)analysis,the distribution of total scores between benign and malignant lesions was significant different (P <0.000 1).Total score 4 was selected as the best cutoff,the area under ROC curve was 0.749,on which the sensitivity,specificity and accuracy were 79.4%,62.7% and 69.67%,respectively.Conclusions The semi-quantitative scoring system of CEUS quantitative analysis color images showed good sensitivity but not satisfied specificity and accuracy in differential diagnosis between malignant and benign breast lesions.

Propofol attenuates hydrogenperoxide-induced apoptosis in human umbilical vein endothelial cells via multiple signaling pathways / 대한마취과학회지

BACKGROUND: Propofol has been reported to protect vascular endothelial cells against oxidative stress. In this study we investigated its effect on hydrogen peroxide (H2O2)-induced apoptosis of human umbilical vein endothelial cells (HUVECs) and examined the possible signaling pathways. METHODS: HUVECs were pretreated with propofol (1, 5, 25, and 50 microM) for 30 min and then co-incubated with 0.4 mM H2O2 for 4 h. Cell viability was assessed using a Cell Counting Kit-8. Cell apoptosis was analyzed using flow cytometry with annexin V/propidium iodide staining, and evaluated by quantifying caspase-3, Bax, and Bcl-2 expression levels. The expression levels of p38 mitogen activated protein kinase (MAPK), phosphorylated (p)-p38 MAPK, cJun-N-terminal kinases (JNK), phosphorylated (p)-JNK, Akt and phosphorylated Akt [(p)-Akt] (Ser473) were measured by western blotting. RESULTS: H2O2 treatment induced the activation of caspase-3, downregulated Bcl-2 expression, and up-regulated Bax expression, all of which were dose-dependently attenuated by propofol pretreatment. Furthermore, propofol significantly ameliorated H2O2-induced phosphorylation of p38 MAPK, JNK, and Akt in HUVECs. CONCLUSIONS: Propofol can protect HUVECs against H2O2-induced apoptosis via a mechanism that may involve p38 MAPK, JNK, and Akt signaling pathways.

HPLC method for the simultaneous determination of four compounds in rat plasma after intravenous administration of Portulaca oleracea L. extract

The objective of the present study was to develop a simple and selective HPLC method for the simultaneous determination of hesperidin (HP), caffeic acid (CA), ferulic acid (FA) and p-coumaric acid (p-CA) in rat plasma after intravenous administration of Portulaca oleracea L. extract (POE). With the hyperoside as the internal standard, the sample pretreatment procedure involved simple single-step extraction with methanol of 0.2 mL plasma. The mobile phase consisted of methanol-acetonitrile-tetrahydrofuran-0.5% glacial acetic acid (5:3:18:74, v/v/v/v). The calibration curves were linear over the range of 0.1-25 µg mL-1, 0.1-25 µg mL-1, 0.1-25 µg mL-1and 0.015-3 µg mL-1 for HP, CA, FA and p-CA, respectively. The method developed was suitable for the pharmacokinetic study of HP, CA, FA and p-CA in rats after intravenous administration of POE.

O objetivo do estudo foi desenvolver um método simples e específico de HPLC para a determinação simultânea de hesperidina (HP), ácido caféico (CA), ácido ferúlico (FA) e ácido p-cumárico (p-CA) em plasma de rato após a administração intravenosa de extrato Portulaca oleracea L. (POE) empregando hyperosídeo como padrão interno de referência. Metanol foi empregado para os analitos em plasma (0,2 mL). A fase móvel isocrática foi composta por metanol-acetonitrila-tetraidrofurano-0,5% ácido acético glacial (5:3:18:74, v/v/v/v). Curvas de calibração foram lineares na faixa de concentração de 0,1-25 µg mL-1, 0,1-25 µg mL-1, 0,1-25 µg mL-1 e 0,015-3 µg mL-1 para HP, CA, FA e p-CA, respectivamente. O método desenvolvido foi adequado para estudo farmacocinético de HP, CA, FA e p-CA em ratos após a administração intravenosa de POE.

Effects of skeletal muscle denervation on potency of rocuronium.

Background: Rocuronium is an alternative to succinylcholine for rapid tracheal intubation after major thermal injury and other forms of critical illness that cause denervation changes in skeletal muscle. Rocuronium may decrease the potencies of non-depolarizing muscle relaxants. Objectives: Examine whether potency of rocuronium changed during the first month after denervation, and investigate the effects of skeletal muscle denervation on potency of rocuronium. Methods: The denervation mouse model was developed to create denervated individual cells from the flexor digitorum brevis of the hindfoot. The skeletal muscle cells were examined at day 0 in the innervated control and days 1, 4, 7, 14, 21, and 28 in the denervation group. Nicotinic acetylcholine receptors in the cells were activated with 30 M acetylcholine, alone or in combination with various concentrations of rocuronium. Currents were recorded with a whole-cell patch-clamp technique. Results: Rocuronium reversibly inhibited acetylcholine-activated currents in a dose-dependent fashion at different times after denervation. The inhibition concentration for the half-maximal responses of rocuronium increased 1.2- (p >0.05), 1.8-, 2.8-, 2.3-, 2.1-, and 1.9-fold (p<0.01) at day 1, 4, 7, 14, 21, and 28 after denervation, respectively, compared to that at day 0 after denervation. Conclusion: Rocuronium dose required to achieve satisfactory clinical effects changed at different durations after skeletal muscle denervation.

Colonic expression of Runx3 protein and TGF-β(1) and their correlation in patients with irritable bowel syndrome

OBJECTIVE@#To investigate the role of Runx3 protein and TGF-β(1) in the pathogenesis of irritable bowel syndrome (IBS), as well as the correlation of these two proteins.@*METHODS@#Colonic tissue was collected from patients with IBS and normal persons. The colonic expression of Runx3 protein and TGF-β(1) was detected with immunohistochemistry method. Semi-quantitative analysis was used to evaluate the staining degree of these two proteins.@*RESULTS@#Compared with their counterparts, patients with IBS did not show any changes in the colonic expression of Runx3 protein and TGF-β(1) (P>0.05). Interestingly, there was a significant correlation between Runx3 protein and TGF-β(1) in patients with IBS(P<0.05).@*CONCLUSIONS@#The role of Runx3 protein and TGF-β(1) in the pathogenesis of IBS remains to be further studied.

Study on the expression of Runx3 and TGF-β₁ protein in the colonic tissue from rats with irritable bowel syndrome

OBJECTIVE@#To investigate the expression of Runx3 and TGF-β(1) protein in the colon from rats with irritable bowel syndrome (IBS).@*METHODS@#Rat model for IBS was established by intracolonic instillation with acetic acid and restraint stress methods, which was confirmed by determinating the visceral sensitivity of the animals, including abdominal withdrawal reflex (AWR) score and the electronic behavior of the abdomen wall. The rats were randomly assigned into three groups: IBS(1) group (restraint stress, n = 25); IBS(2) group (both instillation with acetic acid and restraint stress, n = 25) and Control group (n=16). The colonic tissue samples were collected for histological study and the expression of Runx3 and TGF-β(1) proteins were detected by immunohistochemistry. Meanwhile, the relationship of these two proteins was calculated.@*RESULTS@#Visceral hypersensitivity (AWR and abdominal electrical activity) was significantly enhanced in IBS(1) and IBS(2) groups than other groups. The colon tissue in all groups did not show any signs of inflammation. Furthermore, the expression of Runx3 and TGF-β(1) protein in the colon from all groups show no significant difference (P>0.05), with no remarkable relevancy between each other (P>0.05).@*CONCLUSIONS@#The rat model for IBS was successfully established. We did not find any significant changes in the expression of Runx3 and TGF-β(1) protein in the colon tissue from IBS rats, suggesting that the quantitative changes may be not the way by which Runx3 and TGF-β(1) protein play their roles in IBS. The accurate roles of Runx3 and TGF-β(1) proteins in the pathogenesis of IBS remains to be further studied.

The role of endotoxin in the pathogenesis of gastric mucosal damage in cirrhotic rats with portal hypertensive gastropathy

OBJECTIVE@#To study the role and the mechanism of endotoxin in the pathogenesis of gastric mucosa during portal vein hypertension gastrography (PHG) in the rats with cirrhosis.@*METHODS@#Rat model for PHG was established by injection of tetrachloride. The animals were injected with endotoxin i.p. at 3 mg/kg and endotoxin antagonist BPI21 i.v. at 2.0 mg/kg. The plasma level of endotoxin as well as the gastric mucosal level of tumor necrosis factor alpha (TNF-α) was measured with azobenzene and ELISA respectively. Furthermore, the pathological changes of the gastric mucosa were studied with HE stainning.@*RESULTS@#In rats with PHG, increased endotoxin and TNF-α as well as the gastric pathological lesion were observed. Injection of endotoxin remarkably increased plasma level of endotoxin as well as the gastric mucosal level of tumor necrosis TNF-α and induced more serious gastric lesion. Animals injected with endotoxin antagonist BPI21 showed improved gastric mucosal lesion, accompanied by the declining TNF-α level.@*CONCLUSIONS@#Our results suggestes that endotoxin may play a pathogenetic role in PHG by inducing the expression of TNF-α.

Everolimus vs.Rapamycin for Treating Diabetic Nephropathy in Diabetic Mouse Model / 华中科技大学学报（医学）（英德文版）

In order to evaluate the effectiveness of everolimus vs.rapamycin in the treatment of diabetic nephropathy,8-week old diabetic (db/db) mice received everolimus (2 mg/kg every day) or rapamycin (2 mg/kg every day) for 4 weeks or 12 weeks respectively.Blood and 24-h urine samples were collected for biochemical tests.One kidney from each mouse was homogenized for protein analysis and the other was removed for histological analysis.The expression levels of transforming growth factor-β1 (TGF-βl)and phospho-p70s6k were detected by using ELISA and Western blot,respectively in the renal tissue as well as in mesengial cell culture samples.Everolimus was significantly more effective than rapamycin in improving indexes of renal function and glomerular hypertrophy,and in decreasing accumulation and expansion of the extracellular matrix.However,everolimus inhibited TGF-β1 secretion and p70s6k phosphorylation induced by high glucose in vitro less efficiently than rapamycin at the same dose.Everolimus was more effective than rapamycin in preventing diabetic nephropathy in vivo,which may be contributed to the fact that everolimus has better bioavailability and a higher oral absorptionrate.

Immune changes in type 1 diabetes animal model after syngeneic bone marrow transplantation / 中国实验血液学杂志

Syngeneic bone marrow transplantation (syn-BMT), as a novel therapy for type 1 diabetes (T1D), has been used more and more widely. This study was aimed to detect the changes of peripheral CD4(+) T lymphocytes, CD8(+) T lymphocytes, CD4(+)/CD8(+) T lymphocytes and NK cells before and after T1D mice were treated with syn-BMT, and to investigate the effects of these cells in T1D and the effects of syn-BMT-inducing immunotolerance. T1D mouse model was established by multiple low dose streptozotocin injection, the syn-BMT was performed on 10 day after the onset of diabetes. The T1D model mice were divided into group of diabetic mice treated with syn-BMT and group of diabetic control mice (DC), 6 normal C57BL/6J mice were regarded as normal control group (NC). On 30 day after syn-BMT, peripheral proportion of CD4(+) T lymphocytes, CD8(+) T lymphocytes, CD4(+)/CD8(+) T lymphocytes and NK cells were detected by flow cytometry. These cells of normal control mice (NC), diabetes control mice (DC) and diabetes mice treated by syn-BMT were also detected. Blood glucose level in three groups was assayed during the whole observation period. The results showed that syn-BMT could reduce blood glucose level of T1D mice to near normal (p > 0.05). Hematopoietic reconstitution happened in a month. The proportion of peripheral CD4(+) T lymphocytes, CD4(+)/CD8(+) T lymphocytes, NK cells all increased in new-onset diabetic mice (p < 0.01), while the proportion of peripheral CD8(+) T lymphocytes decreased (p < 0.01). On 30 day after T1D mice were treated with syn-BMT, the proportion of peripheral CD4(+) T lymphocytes was significantly lower than that in DC mice (p < 0.01), but still higher than NC (p < 0.05). The proportion of CD8(+) T lymphocytes was higher than that in DC and NC mice (p < 0.01). The ratio of CD4(+)/CD8(+) T lymphocytes and proportion of NK cells were both obviously lower than that in DC and NC mice (p < 0.01). It is concluded that the syn-BMT can reverse hyperglycemia and immune disorder in diabetic mice. On early period of diabetes onset, the proportions of CD4(+) T lymphocytes and NK cells, the ratio of CD4(+)/CD8(+) T lymphocytes increase, while proportion of CD8(+) T lymphocytes decreases in peripheral blood which mye be associated with onset of diabetes.

Cryopreservation of mouse embryos in ethylene glycol-based solutions: a search for the optimal and simple protocols / 生物工程学报

Although ethylene glycol (EG) has been widely used for embryo cryopreservation in domestic animals, few attempts were made to use this molecule to freeze mouse and human embryos. In the few studies that used EG for slow-freezing of mouse and human embryos, complicated protocols for human embryos were used, and the protocols need to be simplified. Besides, freezing mouse morula with EG as a cryoprotectant has not been reported. In this paper, we studied the effects of embryo stages, EG concentration, duration and procedure of equilibration, sucrose supplementation and EG removal after thawing on the development of thawed mouse embryos, using the simple freezing and thawing procedures for bovine embryos. The blastulation and hatching rates (81.92% +/- 2.24% and 68.56% +/- 2.43%, respectively) of the thawed late compact morulae were significantly (P < 0.05) higher than those of embryos frozen-thawed at other stages. When mouse late compact morulae were frozen with different concentrations of EG, the highest rates of blastocyst formation and hatching were obtained with 1.8mol/L EG. The blastulation rate was significantly higher when late morulae were equilibrated in 1.8 mol/L EG for 10 min prior to freezing than when they were equilibrated for 30 min, and the hatching rate of embryos exposed to EG for 10 min was significantly higher than that of embryos exposed for 20 and 30 min. Both rates of blastocyst formation and hatching obtained with two-step equilibration were higher (P < 0.05) than with one-step equilibration in 1.8 mol/L EG. Addition of sucrose to the EG-based solution had no beneficial effects. On the contrary, an increased sucrose level (0.4 mol/L) in the solution impaired the development of the frozen-thawed embryos. In contrast, addition of 0.1 mol/L sucrose to the propylene glycol (PG)-based solution significantly improved the development of the frozen-thawed embryos. Elimination of the cryoprotectant after thawing did not improve the development of the thawed embryos. The cell numbers were less (P < 0.05) in blastocysts developed from the thawed morulae than in the in vivo derived ones. In summary, embryo stage, EG concentration, duration and procedure of equilibration and sucrose supplementation had marked effects on development of the thawed mouse embryos, and a protocol for cryopreservation of mouse embryos is recommended in which the late morulae are frozen in 1.8 mol/L EG using the simple freezing and thawing procedures of bovine embryos after a two-step equilibration and the embryos can be cultured or transferred without EG removal after thawing.