1.Comparative Study on the Effects of Qingchang Wenzhong Decoction (清肠温中方) on the Intestinal Mucus Barrier in Conventional and Pseudo-Germ-Free Model Mice of Acute Ulcerative Colitis
Mingxu GAO ; Leilei LIU ; Lijie LU ; Jiali WANG ; Juncong HU ; Yangzhe LIN ; Qirui LIU ; Yue DONG ; Luyue WANG ; Zeyu XUE ; Junxiang LI ; Lei SHI
Journal of Traditional Chinese Medicine 2026;67(14):1528-1537
ObjectiveTo explore the mechanisms of Qingchang Wenzhong Decoction (清肠温中方, QWD) in treating ulcerative colitis (UC). MethodsA total of 100 male C57BL/6J mice were assigned to a pseudo-germ-free group (n=45) and a conventional group (n=55) using a random number table. Pseudo-germ-free model was established by giving antibiotic treatment and subsequently subjected to acute UC induction. After the model was successfully established, the 45 pseudo-germ-free acute UC mice were randomly allocated to a pseudo-germ-free model group, a pseudo-germ-free QWD group, and a pseudo-germ-free mesalazine group, with 15 mice in each group. The 55 conventional mice were randomly divided into a normal control group (n=10), as well as a conventional model group, a conventional QWD group, and a conventional mesalazine group, with 15 mice in each group. Except for the normal control group, acute UC models were established in all other groups. Mice in the control group received no intervention. From day 32 of the experiment, mice in the pseudo-germ-free QWD group and the conventional QWD group were administered with QWD granules at a dose of 9.71 g/(kg·d) by gavage with deionized water, while those in the pseudo-germ-free mesalazine group and the conventional mesalazine group were given mesalazine enteric-coated tablets at a dose of 151.67 mg/(kg·d) by gavage. Mice in the remaining groups received an equal volume of deionized water once daily for 7 consecutive days. On day 25 of the experiment, six mice were randomly selected from both the pseudo-germ-free group and the conventional group to measure body weight, and fecal samples were collected for 16S rDNA sequencing to compare differences in gut microbiota, including operational taxonomic units (OTUs), Shannon index, Observed_species index, and phylogenetic diversity (PD whole tree). On day 39, body weight was measured in all groups, and the disease activity index (DAI) was evaluated. Colon length and colon wet weight were recorded. Histopathological changes in colonic tissues were assessed by hematoxylin and eosin (HE) staining, and histological injury scores were determined. In addition, the expression levels of mucopolysaccharides and mucin 2 (MUC2) in colonic tissues were measured. ResultsOn day 25, there was no significant difference in body weight between the pseudo-germ-free group and the conventional microbiota group (P>0.05). Compared to the conventional microbiota group, the pseudo-germ-free group showed significantly reduced OTUs, Shannon index, Observed_species index, and PD whole tree (P<0.01). On day 39, compared to the normal control group, mice in the conventional model group showed a significant decrease in the body weight and an increase in DAI, along with increased colon wet weight, shortened colon length, markedly elevated histopathological scores, and reduced expression of mucopolysaccharides and MUC2 (P<0.01). In contrast, no significant differences were observed in these parameters in the pseudo-germ-free model group (P>0.05). Compared to the conventional model group, the conventional QWD group showed significant improvement in all measured parameters, whereas the conventional mesalazine group showed reductions only in DAI scores and colon wet weight (P<0.01). No statistically significant differences were observed among the pseudo-germ-free model group, the pseudo-germ-free QWD group, and the pseudo-germ-free mesalazine group (P>0.05). ConclusionQWD may promote the reconstruction of the intestinal mucus barrier by regulating the intestinal flora, thereby exerting a therapeutic effect on acute UC.
2.Comparative Study on the Effects of Qingchang Wenzhong Decoction (清肠温中方) on the Intestinal Mucus Barrier in Conventional and Pseudo-Germ-Free Model Mice of Acute Ulcerative Colitis
Mingxu GAO ; Leilei LIU ; Lijie LU ; Jiali WANG ; Juncong HU ; Yangzhe LIN ; Qirui LIU ; Yue DONG ; Luyue WANG ; Zeyu XUE ; Junxiang LI ; Lei SHI
Journal of Traditional Chinese Medicine 2026;67(14):1528-1537
ObjectiveTo explore the mechanisms of Qingchang Wenzhong Decoction (清肠温中方, QWD) in treating ulcerative colitis (UC). MethodsA total of 100 male C57BL/6J mice were assigned to a pseudo-germ-free group (n=45) and a conventional group (n=55) using a random number table. Pseudo-germ-free model was established by giving antibiotic treatment and subsequently subjected to acute UC induction. After the model was successfully established, the 45 pseudo-germ-free acute UC mice were randomly allocated to a pseudo-germ-free model group, a pseudo-germ-free QWD group, and a pseudo-germ-free mesalazine group, with 15 mice in each group. The 55 conventional mice were randomly divided into a normal control group (n=10), as well as a conventional model group, a conventional QWD group, and a conventional mesalazine group, with 15 mice in each group. Except for the normal control group, acute UC models were established in all other groups. Mice in the control group received no intervention. From day 32 of the experiment, mice in the pseudo-germ-free QWD group and the conventional QWD group were administered with QWD granules at a dose of 9.71 g/(kg·d) by gavage with deionized water, while those in the pseudo-germ-free mesalazine group and the conventional mesalazine group were given mesalazine enteric-coated tablets at a dose of 151.67 mg/(kg·d) by gavage. Mice in the remaining groups received an equal volume of deionized water once daily for 7 consecutive days. On day 25 of the experiment, six mice were randomly selected from both the pseudo-germ-free group and the conventional group to measure body weight, and fecal samples were collected for 16S rDNA sequencing to compare differences in gut microbiota, including operational taxonomic units (OTUs), Shannon index, Observed_species index, and phylogenetic diversity (PD whole tree). On day 39, body weight was measured in all groups, and the disease activity index (DAI) was evaluated. Colon length and colon wet weight were recorded. Histopathological changes in colonic tissues were assessed by hematoxylin and eosin (HE) staining, and histological injury scores were determined. In addition, the expression levels of mucopolysaccharides and mucin 2 (MUC2) in colonic tissues were measured. ResultsOn day 25, there was no significant difference in body weight between the pseudo-germ-free group and the conventional microbiota group (P>0.05). Compared to the conventional microbiota group, the pseudo-germ-free group showed significantly reduced OTUs, Shannon index, Observed_species index, and PD whole tree (P<0.01). On day 39, compared to the normal control group, mice in the conventional model group showed a significant decrease in the body weight and an increase in DAI, along with increased colon wet weight, shortened colon length, markedly elevated histopathological scores, and reduced expression of mucopolysaccharides and MUC2 (P<0.01). In contrast, no significant differences were observed in these parameters in the pseudo-germ-free model group (P>0.05). Compared to the conventional model group, the conventional QWD group showed significant improvement in all measured parameters, whereas the conventional mesalazine group showed reductions only in DAI scores and colon wet weight (P<0.01). No statistically significant differences were observed among the pseudo-germ-free model group, the pseudo-germ-free QWD group, and the pseudo-germ-free mesalazine group (P>0.05). ConclusionQWD may promote the reconstruction of the intestinal mucus barrier by regulating the intestinal flora, thereby exerting a therapeutic effect on acute UC.
3.The Influence of Additional Trunk Load and Different Running Speeds on Six Degree of Freedom Kinematics of the Knee Joint
Xiaofan HUANG ; Juncong YANG ; Ye LUO ; Zhuman LI ; Mengling HU ; Shaobai WANG
Journal of Medical Biomechanics 2025;40(3):677-683
Objective By exploring changes of six degrees of freedom(6DOF)kinematics of the knee joint during extra weight bearing of the body trunk,the influence of extra weight on knee joint movement patterns was studied.Methods A total of 24 healthy subjects were recruited to walk/run on a treadmill at four speeds under two states:self-weight and wearing a 16 kg vest,and gait analysis was conducted.A three-dimensional(3D)portable knee kinematics analysis system based on infrared stereophotography was used to capture 6DOF movement trajectory data of the tibia relative to the femur.Results Compared to the self-weight state,when additional trunk weight was added,the knee external rotation angle was reduced at 3.6 km/h speed(1.4°-2.1°)and 5.4 km/h speed(2.2°-2.7°);the knee internal rotation angle was reduced at 10.8 km/h speed(2.1°-4.2°);the knee flexion angles was increased significantly at the speed of 3.6 km/h(1.5°-1.8°),9 km/h(1.6°-3.3°)和 10.8 km/h(1.9°-3.1°);the knee adduction angle increased at 5.4 km/h speed(0.5°-0.6°),and decreased at 10.8 km/h speed(0.9°-1.3°).At 10.8 km/h speed,the distal knee displacement(0.2-0.4 mm)was increased,and the lateral knee displacement(0.1-0.2 mm),and anterior knee displacement(0.2-0.3 mm)were significantly reduced.Conclusions The 6DOF kinematics of human knee is significantly affected by the extra trunk weight.Performance is also different at lower and higher speeds.It is suggested that there may exist a hidden injury in military training,and this study provides a kinematic basis for the occurrence of sports injury.
4.The Influence of Additional Trunk Load and Different Running Speeds on Six Degree of Freedom Kinematics of the Knee Joint
Xiaofan HUANG ; Juncong YANG ; Ye LUO ; Zhuman LI ; Mengling HU ; Shaobai WANG
Journal of Medical Biomechanics 2025;40(3):677-683
Objective By exploring changes of six degrees of freedom(6DOF)kinematics of the knee joint during extra weight bearing of the body trunk,the influence of extra weight on knee joint movement patterns was studied.Methods A total of 24 healthy subjects were recruited to walk/run on a treadmill at four speeds under two states:self-weight and wearing a 16 kg vest,and gait analysis was conducted.A three-dimensional(3D)portable knee kinematics analysis system based on infrared stereophotography was used to capture 6DOF movement trajectory data of the tibia relative to the femur.Results Compared to the self-weight state,when additional trunk weight was added,the knee external rotation angle was reduced at 3.6 km/h speed(1.4°-2.1°)and 5.4 km/h speed(2.2°-2.7°);the knee internal rotation angle was reduced at 10.8 km/h speed(2.1°-4.2°);the knee flexion angles was increased significantly at the speed of 3.6 km/h(1.5°-1.8°),9 km/h(1.6°-3.3°)和 10.8 km/h(1.9°-3.1°);the knee adduction angle increased at 5.4 km/h speed(0.5°-0.6°),and decreased at 10.8 km/h speed(0.9°-1.3°).At 10.8 km/h speed,the distal knee displacement(0.2-0.4 mm)was increased,and the lateral knee displacement(0.1-0.2 mm),and anterior knee displacement(0.2-0.3 mm)were significantly reduced.Conclusions The 6DOF kinematics of human knee is significantly affected by the extra trunk weight.Performance is also different at lower and higher speeds.It is suggested that there may exist a hidden injury in military training,and this study provides a kinematic basis for the occurrence of sports injury.
5.Generation and immunogenicity evaluation of Senecavirus A virus-like particles.
Chunping WU ; Yi RU ; Hong TIAN ; Kun MA ; Rongzeng HAO ; Yajun LI ; Juncong LUO ; Zhengwang SHI ; Huanan LIU ; Zhi ZUO ; Haixue ZHENG
Chinese Journal of Biotechnology 2021;37(9):3211-3220
To develop Senecavirus A (SVA) virus-like particles (VLPs), a recombinant prokaryotic expression plasmid pET28a-SVA-VP031 was constructed to co-express SVA structural proteins VP0, VP3 and VP1, according to the genomic sequence of the field isolate CH-FJ-2017 after the recombinant proteins were expressed in E .coli system, and purified by Ni+ ion chromatographic method. The SVA VLPs self-assemble with a high yield in vitro buffer. A typical VLPs with an average diameter of 25-30 nm which is similar to native virions by using TEM detection. Animals immunized by SVA VLPs shown that the VLPs induced high titers neutralizing antibodies in Guinea pigs. This study indicated that the VLPs produced with co-expressing SVA structural proteins VP0, VP3 and VP1 in prokaryotic system is a promising candidate and laid an important foundation for the development of a novel SVA VLPs vaccine.
Animals
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Antibodies, Neutralizing
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Escherichia coli/genetics*
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Genomics
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Guinea Pigs
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Picornaviridae/genetics*
6.Short-term outcomes of Da Vinci robotic surgical system and laparoscopy in pancreaticoduodenectomy:a Meta analysis
Juncong ZHANG ; Jiaping HU ; Taiyuan LI ; Chen XU
Chinese Journal of Digestive Surgery 2017;16(8):839-843
Objective To systematically evaluate the short-term outcomes of Da Vinci robotic surgical system and laparoscopy in pancreaticoduodenectomy.Methods Literatures were researched using PubMed,Embase,Medline,VIP database,Chinese Journal Fulltext Database from January,2013 to October,2016 with the key words including laparoscopic,robotic,Da Vinci,pancreaticoduodenectomy,腹腔锐,达芬奇机器人,胰十二指肠切除术”.The cohort studies about comparison of short-term outcomes of Da Vinci robotic surgical system and laparoscopy in pancreaticoduodenectomy were received and enrolled.Patients undergoing pancreaticoduodenectomy using Da Vinci robotic surgical system and laparoscopy were respectively allocated into the Da Vinci group and laparoscopy group.Two reviewers independently screened literatures,extracted data and assessed the risk of bias.Count data were described as odd ratio (OR) and 95% confidence interval (CI).Measurement data were represented as weighted mean difference (WMD) and 95%CI.The heterogeneity of the studies was analyzed using the I2 test.Results Five retrospective cohort studies were enrolled in the Meta analysis,and the total sample size was 364 patients,including 159 in the Da Vinci group and 205 in the laparoscopy group.The results of Meta analysis showed that there were statistically significant differences in the rate,of conversion to open surgery,spleen-preserving rate,operation time and duration of postoperative hospital stay between Da Vinci group and laparoscopy group (OR=0.18,3.80,WMD =-37.54,-4.47,95%CI:0.05-0.60,2.01-7.18,-47.46--27.62,-6.70--2.24,P<0.05).There were no statistically significant difference in the incidence of pancreatic fistula and overall incidence of complications between Da Vinci group and laparoscopy group (OR=0.95,0.55,95%CI:0.59-1.54,0.29-1.03,P>0.05).Conclusions Da Vinci robotic surgical system and laparoscopy are safe and feasible in pancreaticoduodenectomy,with the same incidence of postoperative complications.Compared with laparoscopy,Da Vinci robotic surgical system can not only reduce the rate of conversion to open surgery,operation time and duration of postoperative hospital stay,but also increase spleen-preserving rate,meanwhile,it does not increase the incidence of postoperative complications.
7.Effects of hippocampal-sparing intensity-modulated radiotherapy on dose distribution of target volume and organs at risk in locally advanced nasopharyngeal carcinoma
Wendong GU ; Qilin LI ; Ye TIAN ; Juncong MO ; Honglei PEI
Chinese Journal of Radiation Oncology 2017;26(1):6-11
Objective To investigate the effects of hippocampal?sparing intensity?modulated radiotherapy ( IMRT) on dose distribution of target volume and organs at risk ( OARs) in locally advanced nasopharyngeal carcinoma. Methods A retrospective dosimetric analysis was performed among 11 patients with locally advanced nasopharyngeal carcinoma. The MONACO ? v5. 10 Treatment Planning System was used to design three treatment plans:routine volumetric modulated arc therapy ( VMAT ) , hippocampal?sparing VMAT, and nine fixed?fields IMRT. The D98%, D50%, D2%, Dmean , conformity index ( CI ) , and homogeneity index (HI) of planning target volume (PTV) and PTVnx as well as dose distribution of the hippocampus and OARs were evaluated. Using single factor analysis of variance,two group comparative was LSD or paired t?test. Results For the above three plans,the D2% values of PTVnx were ,7 513,and 7 462 cGy,respectively (P=0. 016);the D98% values of PTV were 5837,5812,and 5914 cGy,respectively (P=0. 029);the average D2% values of PTV were 7 399,7 380,and 7 333 cGy,respectively ( P=0. 047);the HI values of PTV were 0. 239,0. 241,and 0. 220,respectively (P=0. 016);the V10 values of the brain stem were 97. 2%,88. 1%,and 90. 3%,respectively ( P=0. 001);the V20 values of the brain stem were 74. 2%, 62. 3%,and 67. 1%,respectively ( P=0. 032);the V30 values of the brain stem were 50. 9%,35. 8%,and 45. 5%, respectively ( P= 0. 020 );the V40 values of brain stem were 24. 4%, 14. 4%, and 23. 3%, respectively ( P=0. 018);the Dmean values of hippocampus were 1 518,899,and 896 cGy,respectively ( P=0. 000);the D40% values of hippocampus were 1 379,642,and 639 cGy,respectively ( P=0. 000);the V10 values of the hippocampus were 54. 1%,25. 1%,and 3. 8%,respectively ( P=0. 000);the V20 values of the hippocampus were 26. 2%, 12. 6%, and 12. 0%, respectively ( P=0. 001 ) . Conclusions Hippocampal?sparing VMAT and nine fixed?fields IMRT can significantly reduce the dose to the hippocampus without affecting dose distribution of target volume and OARs. VMAT may be superior to IMRT because VMAT can simultaneously reduce the dose to the brain stem.
8.The effect of high mobility group box-1 protein in central nerve system on septic brain injury
Chao REN ; Xiouhua LI ; Bilei XU ; Juncong LI ; Yao WU ; Ning DONG ; Zhongqiu LU ; Yongming YAO
Chinese Journal of Emergency Medicine 2016;25(4):433-438
Objective To investigate changes in high mobility group box-1 protein ( HMGB1 ) level in brain tissues with severe sepsis, and the relationship between HMGB1 and septic brain injury.Methods Forty wild C57BL/6 mice were randomly ( random number) divided into 4 groups: sham group, sepsis group, cerebroventricular injection control group, and sepsis with BoxA ( HMGB1 inhibitor) cerebroventricular injection group.Septic model was reproduced by cecal ligation and puncture, and the cerebroventricular catheterization model was established by motorized mice brain stereotaxic instruments.After septic challenge, 1 μg BoxA was injected into the ventricle of brain via cerebroventricular catheter immediately.Mice were sacrificed and brains were harvested at 24 h after sepsis, and hippocampus tissue was separated immediately.Expressions of brain HMGB1 and caspase-3 changed in apoptotic neurons and brain injury were determined by brain tissue immunofluorescence, Western blotting, TUNEL and HE staining respectively.One-way analysis of variance ( ANOVA) for analyzing inter-group differences, student t test for comparing difference between two groups . Results (1) HMGB1 expression in hippocampus was significantly enhanced in the septic group compared to the sham group [ (22.74 ±9.29) vs.4.57 ±2.18, P<0.01].(2) Compared to the sham group, neuronal apoptosis [ (35 ±9.17) vs.(1.67 ±1.53) , P<0.01) and caspase-3 expressions [ (16.79 ±8.17) vs.( 3.39 ±2.09), P<0.05] were significantly increased in hippocampus with aggravated brain injury in the septic group.(3) Cerebroventricular injection of BoxA significantly inhibited HMGB1 in hippocampus [ (2.66 ± 2.06) vs.( 22.74 ±9.29), P<0.01];(4) Cerebroventricular injection of BoxA obviously alleviated acute brain injury, and decreased neuronal apoptosis [ ( 12 ±4.36 ) vs.( 35 ±9.17 ) , P <0.01 ] as well as caspase-3 activity [ (4.10 ±2.11) vs.(16.80 ±8.17), P<0.05].Conclusions The elevated expression of brain HMGB1 is closely related to pathogenesis and development of septic brain injury, and treatment with antagonist towards brain HMGB1 can markedly attenuate acute brain injury following severe sepsis.
9.Effects of ulinastatin on immune function of patients with severe burn injury.
Juncong LI ; Chao HU ; Hongming YANG ; Yongming YAO
Chinese Journal of Burns 2016;32(6):345-350
OBJECTIVETo primarily evaluate the effects of ulinastatin on immune function of patients with severe burn injury.
METHODSForty patients with severe burn admitted to our ward from March 2013 to October 2015, conforming to the study criteria, were divided into conventional treatment group (CT, n=20) and ulinastatin treatment group (UT, n=20) according to the random number table and patient's consent. After admission, patients in group CT received antishock treatment, antibiotic treatment, debridement, skin grafting, and nutrition support, etc. On the basis of the above-mentioned treatment, patients in group UT received intravenous drip of ulinastatin from first day after admission twice a day, with a dosage of 8×10(5) U every time, for 7 days in addition. Peripheral venous blood samples were collected from patients in groups CT and UT on post treatment day (PTD) 1, 3, 5 and 7, respectively. Twenty healthy volunteer were selected as health control group (HC), and peripheral venous blood samples were collected on the first day of the study. Percentage of CD4(+) CD25(+) regulatory T lymphocytes (Tregs) was determined by flow cytometer. The proliferative activity of T lymphocytes was detected by microplate reader (denoted as absorbance value). Content of interleukin 2 (IL-2) in culture supernatant of T lymphocytes, and content of IL-4 and γ interferon (IFN-γ) in serum were detected by enzyme-linked immunosorbent assay. Expression of human leukocyte antigen-DR (HLA-DR) on CD14(+) monocytes was determined by flow cytometer. Data were processed with analysis of variance for repeated measurement, chi-square test, and LSD-t test.
RESULTS(1) Compared with that of volunteer in group HC, the percentage of CD4(+) CD25(+) Tregs of patients in group CT was significantly increased from PTD 1 to 7 (with t values from 13.303 to 26.043, P values below 0.01). Compared with that in group CT, the percentage of CD4(+) CD25(+) Tregs of patients in group UT was significantly decreased on PTD 5 and 7 (with t values respectively 8.317 and 15.071, P values below 0.01). (2) The proliferative activity of T lymphocytes of patients in group CT on PTD 1, 3, 5, and 7 was respectively 0.71±0.11, 0.61±0.15, 0.54±0.12, and 0.67±0.17, which was significantly lower than that in group HC (1.21±0.22, with t values from 8.686 to 11.957, P values below 0.01). The proliferative activity of T lymphocytes of patients in group UT on PTD 3, 5, and 7 were respectively 0.81±0.11, 0.85±0.14, and 1.08±0.13, which was significantly higher than that in group CT (with t values from 4.808 to 8.568, P values below 0.01). (3) Compared with those of volunteer in group HC, content of IL-2 in culture supernatant of T lymphocytes of patients in group CT was significantly decreased from PTD 1 to 7 (with t values from 8.073 to 9.288, P values below 0.01), content of IL-4 in serum of patients in group CT was significantly increased from PTD 1 to 7 (with t values from 18.926 to 41.451, P values below 0.01), and content of IFN-γ in serum of patients in group CT was significantly decreased from PTD 1 to 7 (with t values from 4.543 to 27.659, P values below 0.01). Compared with those in group CT, content of IL-2 in culture supernatant of T lymphocytes of patients in group UT was significantly increased from PTD 3 to 7 (with t values from 6.507 to 8.869, P values below 0.01), content of IL-4 in serum of patients in group UT was significantly decreased from PTD 3 to 7 (with t values from 6.922 to 8.843, P values below 0.01), and content of IFN-γ in serum of patients in group UT was significantly increased on PTD 5 and 7 (with t values respectively 5.369 and 13.521, P values below 0.01). (4) The percentages of CD14(+) monocytes with positive expression of HLA-DR of patients in group CT on PTD 1, 3, 5, and 7 were respectively (28±6)%, (25±7)%, (25±7)%, and (39±10)%, which were significantly lower than the percentage of volunteer in group HC [(87±8)%, with t values from 16.323 to 25.645, P values below 0.01]. The percentages of CD14(+) monocytes with positive expression of HLA-DR of patients in group UT on PTD 3, 5, and 7 were respectively (40±6)%, (42±9)%, and (49±10)%, which were significantly higher than those in group CT (with t values from 3.071 to 7.324, P values below 0.01).
CONCLUSIONSOn the basis of CT, additional ulinastatin intervention can decrease CD4(+) CD25(+) Tregs percentage, improve the immune function of T lymphocytes and T helper cells, and increase expression of HLA-DR on CD14(+) monocytes of patients with severe burn injury, thus improve the immune function of patients.
Burns ; drug therapy ; immunology ; Cells, Cultured ; Debridement ; Enzyme-Linked Immunosorbent Assay ; Glycoproteins ; therapeutic use ; Humans ; Interferon-gamma ; blood ; Interleukin-2 ; metabolism ; Interleukin-4 ; blood ; Monocytes ; immunology ; Skin Transplantation ; T-Lymphocytes, Regulatory ; immunology
10.Effects of ulinastatin on immune function of spleen in severely burned rats and its mechanism.
Juncong LI ; Chao HU ; Yongming YAO ; Hongming YANG
Chinese Journal of Burns 2016;32(5):266-271
OBJECTIVETo observe the effects of ulinastatin on immune function of splenic CD4(+) T lymphocytes and CD4(+) CD25(+) regulatory T lymphocytes (Tregs) and content of high mobility group box 1 (HMGB1) in peripheral blood of severely burned rats, and to analyze the possible mechanisms.
METHODSNinety-six male SD rats were divided into sham injury group, burn group, and ulinastatin group according to the random number table, with 32 rats in each group. Rats in sham injury group were sham injured on the back by immersing in 37 ℃ warm water for 12 s. Rats in burn group and ulinastatin group were inflicted with 30% total body surface area full-thickness scald (hereinafter referred to as burn) on the back by immersing in 94 ℃ hot water for 12 s. Immediately after injury, rats in each group were intraperitoneally injected with saline (40 mL/kg), meanwhile rats in ulinastatin group were intraperitoneally injected with ulinastatin (4×10(4) U/kg), once per 12 h, till post injury hour 72. Eight rats of each group were respectively selected on post injury day (PID) 1, 3, 5, and 7 to collect abdominal aortic blood samples. Serum content of HMGB1 was detected by enzyme-linked immunosorbent assay (ELISA). And then, rats of the 3 groups were sacrificed immediately to collect spleens and separate CD4(+) CD25(+) Tregs and CD4(+) T lymphocytes. Flow cytometer was used to detect positive expression rates of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) and forkhead-winged helix transcription factor p3 (Foxp3) in CD4(+) CD25(+) Tregs. Content of IL-10 in culture supernatant of CD4(+) CD25(+) Tregs, and content of interleukin 2 (IL-2), IL-4, and γ interferon (IFN-γ) in culture supernatant of CD4(+) T lymphocytes was detected by ELISA. The proliferative activity of CD4(+) T lymphocytes was determined by microplate reader. The sample number of above-mentioned experiments was 8 at each time point in each group. Data were processed with analysis of variance of factorial design and LSD test.
RESULTS(1) Compared with that in sham injury group, serum content of HMGB1 of rats in burn group was significantly increased from PID 1 to 7 (with P values below 0.01). Compared with that in burn group, serum content of HMGB1 of rats in ulinastatin group was significantly decreased from PID 1 to 7 (with P values below 0.01). (2) Compared with those in sham injury group, the positive expression rates of CTLA-4 and Foxp3 in CD4(+) CD25(+) Tregs and content of IL-10 in culture supernatant of CD4(+) CD25(+) Tregs of rats in burn group were significantly increased from PID 1 to 7 (with P values below 0.01), peaking on PID 3 [(65±10)%, (76±10)%, and (28.2±4.4) pg/mL respectively]. These 3 indexes of rats in sham injury group on PID 3 were (45±7)%, (46±7)%, and (11.2±2.3) pg/mL respectively. Compared with those in burn group, the positive expression rates of CTLA-4 and Foxp3 in CD4(+) CD25(+) Tregs and content of IL-10 in culture supernatant of CD4(+) CD25(+) Tregs of rats in ulinastatin group were significantly decreased from PID 1 to 7 (P<0.05 or P<0.01), reaching the nadir on PID 7 [(43±6)%], PID 1 [(50±8)%], and PID 7 [(12.4±3.4) pg/mL] respectively. These 3 indexes of rats in burn group on PID 7, 1, and 7 were (58±8)%, (71±9)%, and (19.7±2.8) pg/mL respectively. (3) Compared with those in sham injury group, the content of IL-2 and IFN-γ in culture supernatant of CD4(+) T lymphocytes of rats was significantly decreased, while the content of IL-4 in culture supernatant of CD4(+) T lymphocytes of rats was significantly increased in burn group from PID 1 to 7, with P values below 0.01. Compared with that in burn group, the content of IL-2 and IFN-γ in culture supernatant of CD4(+) T lymphocytes of rats was significantly increased, while the content of IL-4 in culture supernatant of CD4(+) T lymphocytes of rats was significantly decreased in ulinastatin group from PID 1 to 7, P<0.05 or P<0.01. (4) Compared with that in sham injury group, the proliferative activity of CD4(+) T lymphocytes of rats in burn group was significantly decreased from PID 1 to 7 (with P values below 0.01). Compared with that in burn group, the proliferative activity of CD4(+) T lymphocytes of rats in ulinastatin group was significantly increased from PID 1 to 7 (with P values below 0.01).
CONCLUSIONSUlinastatin can weaken the immunosuppressive function mediated by splenic CD4(+) CD25(+) Tregs in severely burned rats, and improve proliferative function and secretory function of splenic CD4(+) T lymphocytes, which may be attributed to the inhibiting effect of ulinastatin on the release of HMGB1 in large amount.
Animals ; Burns ; drug therapy ; CTLA-4 Antigen ; metabolism ; Enzyme-Linked Immunosorbent Assay ; Flow Cytometry ; Forkhead Transcription Factors ; metabolism ; Glycoproteins ; pharmacology ; HMGB1 Protein ; blood ; Interferon-gamma ; metabolism ; Interleukin-10 ; metabolism ; Interleukin-2 ; metabolism ; Interleukin-4 ; metabolism ; Male ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Spleen ; drug effects ; T-Lymphocytes, Regulatory ; cytology ; drug effects

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