1.Heterogenous acellular dermal matrix seeded with adipose derived stem cells for urethral reconstruction in a rabbit model
Bingwei HUANG ; Sen LI ; Xi YU ; Peng GE ; Jian LIN
Chinese Journal of Urology 2013;34(10):787-792
Objective To assess the feasibility of heterogeneous acellular dermal matrix(ADM)seeded with adipose derived stem cells(ADSC)for urethroplasty in a rabbit model.Methods ADSC were isolated from a rabbit and expanded in vitro,then identified by flow cytometry.We seeded ADSC onto the ADM and made it into tissue-engineered urethra.12 male rabbits were removed 1 cm urethra and divided into experiment group and control group.There were 6 rabbits in each group.Reconstructed urethra with tissueengineered urethra was used in experiment group,while unseeded ADM were used in control group.Urethrography was performed at 6 months after surgery.The animals were scarified at 3 and 6 months after surgery and the repaired urethra were harvested.H&E staining and immunohistochemical staining were performed with cytokeratin AE1/AE3 and smooth muscle desmin makers.Results The morphology of isolated ADSC was with long spindle cross-links,and had multicentral growth.Flow cytometry showed that the ADSC expressed CD166,CD105,CD90 and CD44,but not expressed CD45 and CD13.The cells could growth well on the ADM and showed good biocompatibility with it.All animals could void normally,urethrography showed there was no significant stenosis.3 months after surgery,the experiment group appeared regenerated smooth muscle but not in the control group,both groups did not regenerate urothelium.At 6 months urothelium and smooth muscle cells could be observed in the experiment group,but only the smooth muscle was evident in the control group.Conclusions By applying tissue engineering methods,we can seed the ADSC onto the heterogeneous ADM and make it into tissue-engineered urethra,which can help improve the reconstructive effect of urethra.
2.Recent memory consequences of drug-resistant medial temporal lobe epilepsy and MRI-positive hippocampns
Bingwei PENG ; Liwen WU ; Yan CHEN ; Qiong MENG ; Feng FENG ; Yan HUANG
Chinese Journal of Neurology 2008;41(1):37-40
Objective To assess the effects of MRI-positive and interictal epileptic charges(IEDs) dominance on the memory of the patients with drug-resistant medial temporal lobe epilepsy (MTLE). Methods Fifty right-handed patients(age ranging from 16 to 60 years old)diagnosed as drug-resistant MTLE in our hospital with normal intelligence between September 2006 and April 2007 were investigated. All patients were classified as left MRI-positive(MRI(+)),right MRI(+),MRI(-),bilateral MR[ (+)by high-quality MR[protocol.The EEG was defined as dominant IEDs if≥75%independent IEDs was confined to one temporal lobe in all EEG recordings.Clinical memory scale was administered as memory assessment of MTLE.ANOVA and non-parametric statistics were used to analyze the data in SPSS 12.0. Results The distribution of age,sex,education,occupation,living condition,course,seizure and treat among left MRI(+),right MRI(+),MRI(-),bilateral MRI(+)groups was similar.All scores in the patients with MTLE was significantly lower than normal(P<0.05).Right MRI(+)MTLE patients had deftcits in nonsense graphics recognition(9.42±7.46)compared to left MRI(+)and MRI(-) groups((16.26±4.43)and(18.26±5.49),F=4.281,P<0.05).Among MRI(-)patients,left IEDs,right IEDs and bilateral IEDs groups displayed not significantly different impairment in memory. Conclusion Right MRI(+)MTLE has more severe impairment in non-verbal memory,and nonsense graphics recognition can be used to detect the deficit.
3.Application value of 3D printing technology in the surgery of sphenoid ridge meningioma
LIU YUQING ; HE BINGWEI ; HUANG SHENGYUE ; YANG ZHIKUN ; ZHUANG JIANGHUI ; CHEN MINGWU ; CHEN SHOU ; LIAO ZHENGJIAN
Chinese Journal of Clinical Oncology 2017;44(22):1146-1150
Objective:To investigate the value of 3D printing technology in sphenoid ridge meningioma dissection. Methods:By using craniocerebral spiral enhanced CT scan DICOM images, the skull, vessels, and tumor were extracted, reconstructed, and assembled and integrated in the same coordinate system. Then, we constructed a 3D virtual model and a 3D-printed entity model, which was ap-plied for preoperation and postoperation. Results:Virtual models of the brains of five patients were reconstructed successfully and 3D entity models were produced. The models expressed the relationship among tumors, adjacent blood vessels, and the important posi-tion of the nerve tissue. Then, the models were applied to the reference before surgery planning and after surgery. Five cases were successfully performed. Conclusion:The use of the entity model of sphenoid ridge meningioma is important in optimizing operation plans, improving tumor resection, and reducing intraoperative bleeding.
4.Mechanism of heparin-binding protein increasing vascular permeability in early burn
Mingming SONG ; Lu LIU ; Xinxin QI ; Yunxi YANG ; Jiamin HUANG ; Ran SUN ; Bingwei SUN
Chinese Critical Care Medicine 2020;32(3):330-335
Objective:To investigate the effect of heparin-binding protein (HBP) on the damage of vascular permeability in early burn.Methods:① Clinical research: 12 patients with severe burns admitted to Suzhou Hospital of Nanjing Medical University from January 1st to August 30th in 2019 were enrolled. Eight patients with severe trauma admitted to the same hospital during the same period were also enrolled as controls to explain the specificity of burn injury. Whole blood samples were obtained within 0.5 hour after admission. The white blood cell count (WBC), absolute value and ratio of neutrophils, and serum HBP levels were measured. Serum samples of 12 patients with severe burn were collected within 9 days after admission, and enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of metabolism products of glycocalyx including syndecan-1 and hyaluronic acid (HA). The correlation between HBP and neutrophils ratio, syndecan-1 and HA were analyzed by linear correlation. ② Basic research: a 30% total body surface area (TBSA) Ⅲ° burn model of Sprague-Dawley (SD) rat aged 6-8 weeks was prepared. In low molecular weight heparin (LMWH) intervention group ( n = 5), 200 U/kg LMWH was injected subcutaneously immediately and every 2 hours after injury for 4 times in total; the burn group ( n = 5) was given the same amount of normal saline. No intervention was given to the normal control group ( n = 5). The peripheral venous blood was collected at 0, 2, 4, and 8 hours after injury, and the serum levels of HBP, syndecan-1 and HA were measured; the injury of glycocalyx on pulmonary vascular endothelial cells was observed under transmission electron microscope. Results:① Clinical research results: the WBC, neutrophils absolute value and ratio, and HBP levels were increased in 12 patients with severe burn and 8 patients with severe trauma. There was no significant difference in the WBC, absolute value and ratio of neutrophils between severe burn and severe trauma patients [WBC (×10 9/L): 14.5±6.1 vs. 10.8±3.6, the absolute value of neutrophils (×10 9/L): 12.0±5.9 vs. 9.0±4.0, the ratio of neutrophils: 0.81±0.10 vs. 0.79±0.14, all P > 0.05], but the HBP levels in the burn patients were significantly higher than those in the trauma patients (μg/L: 192.92±61.73 vs. 51.17±23.05, P < 0.01). Twelve patients with severe burns had a sharp increase in serum syndecan-1 and HA levels after burns, which continued to maintain high levels and peaked at the 9th day [syndecan-1 (μg/L): 16.02±0.39, HA (μg/L): 106.83±4.90]. The analysis showed that HBP was positively correlated with neutrophils ratio, syndecan-1 and HA in severe burn patients at the 1st day after admission ( r values were 0.805, 0.732 and 0.900, respectively, all P < 0.01). It indicated that the sharp increase of neutrophils after the burn released a lot of HBP, and the glycocalyx of the vascular endothelium was severely damaged. ② Basic research results: the levels of serum HBP, syndecan-1 and HA in the burn group were increased sharply as compared with the normal control group, and continued to increase with time, reaching a peak at 8 hours after burn. In the LMWH intervention group, the serum levels of HBP, syndecan-1 and HA were significantly lower than those in the burn group, and the difference was still statistically significant after 8 hours [HBP (μg/L): 6.47±0.25 vs. 12.48±0.08, syndecan-1 (μg/L): 19.06±1.48 vs. 25.92±3.34, HA (μg/L): 35.76±2.10 vs. 54.91±2.64, all P < 0.01]. The results of transmission electron microscopy showed that in the normal control group, the glycocalyx pulmonary vascular endothelial cells was continuous, evenly distributed and dense. The glycocalyx on pulmonary vascular endothelial cells of rats were significantly damaged and shed 2 hours after burn in the burn group, and no glycocalyx was observed at 8 hours. In the LMWH intervention group, the glycocalyx on pulmonary vascular endothelial cells was damaged and the phenomenon of shedding was significantly relieved, and the glycocalyx could be observed 8 hours after the injury. Conclusion:The massive exudation of body fluids and the significant increase of vascular permeability in patients in early burns may be related to the destruction of the glycocalyx on endothelial cells by HBP released from increased neutrophils.
5.Neutrophils mediate T lymphocyte function in septic mice via the CD80/cytotoxic T lymphocyte antigen-4 signaling pathway
Ran SUN ; Jiamin HUANG ; Lu LIU ; Yunxi YANG ; Mingming SONG ; Yiming SHAO ; Linbin LI ; Bingwei SUN
Chinese Critical Care Medicine 2021;33(7):849-854
Objective:To investigate the effect of neutrophils on T lymphocyte function in septic mice and the role of CD80/cytotoxic T lymphocyte antigen-4 (CTLA-4) signaling pathway in this modulated effects.Methods:① In vivo experiment: 6-8 weeks old male C57BL/6 mice were divided into sham operation group (Sham group, n = 20), Sham+CTLA-4 antibody treatment group (Sham+aCTLA-4 group, n = 20), cecal ligation and perforation (CLP) induced sepsis model group (CLP group, n = 30) and CLP+CTLA-4 antibody treatment group (CLP+aCTLA-4 group, n = 30) according to the random number table. CLP was used to reproduce mouse sepsis model. The mice in the Sham group were treated identically but their cecums were neither punctured nor ligated. In CTLA-4 antibody treatment groups, 50 μg CTLA-4 antibody was injected intraperitoneally 6 hours and 24 hours after the operation. Forty-eight hours after operation, 6 mice in Sham group and Sham+aCTLA-4 group, 14 mice in CLP group and CLP+aCTLA-4 group were randomly selected to detect the expression of CD69 in spleen. At the same time, spleen, bone marrow and peripheral blood were collected, and the expression of CD80 on neutrophils was detected by flow cytometry. The expression of CTLA-4 on the surface of T lymphocytes in spleen was detected by immunofluorescence and flow cytometry. The remaining mice in each group were used to observe the 96-hour survival after operation.② In vitro experiment 1: neutrophils were extracted from bone marrow of healthy mice and stimulated with LPS (1 mg/L) for 4, 8 and 12 hours respectively. The control group was added with the same amount of phosphate buffered saline (PBS) at each time point, and the expression of CD80 was detected at each time point.③ In vitro experiment 2: splenic T lymphocytes of healthy mice were extracted and divided into PBS control group, LPS group (final concentration of LPS 1 mg/L), neutrophil group and neutrophil+LPS group. In the latter two groups, the co-culture model of neutrophils and T lymphocytes was established, and then the corresponding treatment was given to detect the expression of CTLA-4 on the surface of T lymphocytes. With the above four groups as controls, CTLA-4 antibody treatment groups (final concentration of CTLA-4 antibody 50 mg/L) were set up respectively. After 48 hours, the level of interleukin-2 (IL-2) in the cell supernatant was detected by enzyme linked immunosorbent assay (ELISA). Results:① Results of in vivo experiment: compared with Sham group, the expression of CD80 on neutrophils in spleen, bone marrow and peripheral blood was significantly up-regulated, while the expression of CTLA-4 on the surface of T lymphocytes was significantly increased [(9.98±0.84)% vs. (3.48±0.64)%, P < 0.05]. It suggested that neutrophils may affect T lymphocytes function through CD80/CTLA-4 pathway in sepsis. Compared with CLP group, CTLA-4 antibody could significantly improve the 96-hour cumulative survival rate of CLP mice (56.25% vs. 18.75%, P < 0.05), and increase the expression of CD69 on the surface of T lymphocytes. It suggested that CTLA-4 antibodies might increase T lymphocytes activation in sepsis and improve survival. ② Results of in vitro experiment: with the prolongation of LPS stimulation, the expression of CD80 on neutrophils gradually increased in time-dependent manner as compared with PBS control group [4 hours: (6.35±0.40)% vs. (3.41±0.40)%, 8 hours: (8.57±0.64)% vs. (3.09±0.27)%, 12 hours: (19.83±1.06)% vs. (5.16±0.36)%, all P < 0.05]. Compared with PBS control group, the expression of CTLA-4 on CD4 +/CD8 + T lymphocytes was not significantly affected by LPS stimulation alone, but CTLA-4 was increased after co-culture with neutrophils [CD4 +: (4.92±0.30)% vs. (3.33±0.25)%, CD8 +: (4.26±0.21)% vs. (2.53±0.66)%, both P < 0.05], and the increased trend of CTLA-4 was more obvious after co-culture with LPS-stimulated neutrophils [CD4 +: (6.34±0.50)% vs. (3.33±0.25)%, CD8 +: (6.21±0.41)% vs. (2.53±0.66)%, both P < 0.05]. In the PBS control group and LPS group, CTLA-4 antibody had no significant effect on IL-2 secretion of T lymphocytes. Compared with PBS control group, co-culture with neutrophils could inhibit the secretion of IL-2 by T lymphocytes (ng/L: 1 938.00±68.45 vs. 2 547.00±218.00, P < 0.05), and the inhibitory effect of neutrophils stimulated by LPS was more obvious (ng/L: 1 073.00±34.39 vs. 2 547.00±218.00, P < 0.05). CTLA-4 antibodies could partially restore IL-2 secretion. In conclusion, after promoting the expression of CTLA-4 on the surface of T lymphocytes, neutrophils might mediate the inhibition of T lymphocytes function by reducing the production of IL-2. Conclusions:Neutrophils mediate T lymphocytes dysfunction in sepsis, and the CD80/CTLA-4 pathway plays an important role. The CTLA-4 antibody improves survival and T lymphocytes function in sepsis mice, which may be a new method of immunotherapy for sepsis.
6. Mechanism of delayed apoptosis of peripheral blood neutrophils in severely burned patients
Xinxin QI ; Yunxi YANG ; Jiamin HUANG ; Ran SUN ; Lu LIU ; Bingwei SUN
Chinese Critical Care Medicine 2019;31(10):1208-1211
Objective:
To observe the changes of peripheral blood neutrophil apoptosis rate and the effect of p38 mitogen-activated protein kinase (p38MAPK) inhibitor on peripheral blood neutrophil apoptosis in severely burned patients.
Methods:
Severe burn patients [burn area ≥ 30% total body surface area (TBSA), deep Ⅱ to Ⅲ degrees, burn index ≥ 20%, age > 20 years old] admitted to the department of burn and plastic surgery of Affiliated Suzhou Hospital of Nanjing Medical University from January to May in 2019 and health examination volunteers during the same period were enrolled. The peripheral blood neutrophils were isolated by Ficoll gradient centrifugation. The neutrophils of severely burned patients were divided into burn group and p38MAPK inhibitor SB203580 group (after 30 minutes incubation at room temperature and 24 hours incubation in incubator); the neutrophils of healthy volunteers were used as control group. The apoptosis of neutrophils was detected by flow cytometry; the level of reactive oxygen species (ROS) in neutrophils was detected by fluorescence probe; the expression of p38MAPK protein and its phosphorylation (p-p38MAPK) level were detected by Western Blot.
Results:
Compared with the control group, the apoptosis rate of neutrophils in burn group was significantly decreased [(37.42±3.14)% vs. (50.20±9.87)%,
7.Analysis of the diagnosis and treatment of 68 cases of iatrogenic ureteral stricture
Peng ZHANG ; Bingwei HUANG ; Guicao YIN ; Shengqi ZHENG ; Zhe LI ; Shuo QIAO ; Xuesong LI
Journal of Modern Urology 2023;28(7):613-618
【Objective】 To investigate the common etiology, characteristics and treatment of iatrogenic ureteral stricture. 【Methods】 The clinical data of 226 patients with ureteral stricture repaired during May 2019 and Mar. 2022 were retrospectively analyzed, including 68 cases of iatrogenic ureteral stricture. According to the etiology, the patients were divided into urinary group and non-urinary group. 【Results】 There were 42 females and 26 males, aged 25 to 67 (average 49.0±10.4) years. Upper ureteral stricture was detected in 24 (35.3%) cases, who received oral mucosal repair of the ureter. Middle ureteral stricture was detected in 12 (17.6%) cases, who underwent ileal ureterography. Lower ureteral stricture was observed in 24 (35.3%) cases, who were treated with vesical wall flap ureteroplasty. Full-length stricture was observed in 8 (11.8%) cases,who were treated with ileal ureterography. There were significant differences in age, gender, stenosis side, stenosis location and length, surgical methods and types between patients in the urinary group and non-urinary group (P<0.05). During the follow-up of 8 to 20 (average 12.3±5.6) months, the symptoms and renal function of all patients improved, and no recurrence occurred. 【Conclusion】 Invasive endourological surgery is the most common cause of iatrogenic ureteral stenosis. Different treatment strategies should be adopted according to patients’ condition, time of diagnosis and location and length of ureteral injury.
8. Effect of autophagy on expression of neutrophil programmed death ligand-1 in mice with sepsis
Jiamin HUANG ; Ran SUN ; Xinxin QI ; Lu LIU ; Yunxi YANG ; Bingwei SUN
Chinese Critical Care Medicine 2019;31(9):1091-1096
Objective:
To investigate the effect and mechanism of autophagy on the expression of neutrophil programmed death ligand-1 (PD-L1) in mice with sepsis.
Methods:
①