1.Hepatocyte growth factor/c-Met signaling pathway decreases doxorubicin sensitivity in hepatocellular carcinoma in vitro
Dhruba KADEL ; Yu ZHANG ; Haoran SUN ; Qiongzhu DONG ; Lunxiu QIN
Cancer Research and Clinic 2017;29(5):289-293,299
Objective To explore the effect of hepatocyte growth factor (HGF)/c-Met signaling in doxorubicin (DOX) treatment of hepatocellular carcinoma (HCC). Methods Different biologic and genetic characteristics human HCC cell lines, Huh7, HepG2, MHCC97-L and MHCC97H were used in this experiment. Variation in c-Met mRNA expression level among different HCC cell lines was analyzed by RT-PCR. Western blot analysis was performed to detect c-Met and p-Met expression levels in these cell lines. CCK-8 experiment was carried to analyze the DOX sensitivity in various cell lines. t test and repeated measure analysis of variance were used for statistical analysis. Results Both c-Met and p-Met were overexpressed in MHCC97-L and MHCC97-H cell lines and these cell lines were resistant to DOX compared to Huh7 and HepG2. However, treatment of HGF in Huh7 and HepG2 cells activated c-Met signaling pathway and decreased the sensitivity of these two cell lines to DOX [inhibition rate: Huh7 (34.848 ±5.370) vs. (66.409±5.792)%, HepG2 (34.351±3.305) %vs. (62.308±5.453) %, both P=0.002]. Whereas administration of c-Met inhibitor in MHCC97-L and MHCC97-H cell lines significantly increased the sensitivity to DOX [inhibition rate: MHCC97-L (73.106 ±3.472) % vs. (13.636 ±4.097) %; MHCC97-H (64.444 ±4.006) % vs. (6.296 ±2.796) %, both P< 0.001]. Conclusion HGF/c-Met signaling pathway is related the treatment efficacy of DOX in HCC.
2.Isolation, culture and identification of hepatocellular carcinoma-associated fibroblast and its effect on the biological function of hepatocellular carcinoma cells
Haoran SUN ; Yu ZHANG ; Dhruba KADEL ; Qiongzhu DONG ; Lunxiu QIN
Cancer Research and Clinic 2017;29(8):505-509
Objective To establish an in vitro isolation and culture system for hepatocellular carcinoma (HCC)-associated fibroblast (CAF) and identify based on its specific markers of CAF. Methods CAF was isolated from the tumor tissue of the HCC patients after hepatectomy, by digesting with collagenase enzyme, centrifugation and resuspension. The morphology of CAF was observed by electron microscopy. Immunofluorescence staining was performed for detecting α-SMA and fibronectin as well as other specific markers such as AFP on the surface of HCC cells and CD31 on the surface of vascular endothelial cells. On the basis of the study, the function work between CAF and HCC cell line Huh7, HepG2 was detected under co-culture system. Meanwhile, CCK-8 was used to observe the effect of CAF on the proliferation of Huh7 and HepG2 cells, and Transwell assay was used to analyze CAF effect on the invasion of Huh7 and HepG2 cells. Results Compared with the other cells, the morphological analysis showed that CAF was more elongated or spindle-shaped. Moreover, the cell size and the nucleus were larger than normal epithelial cells. Immunofluorescence staining revealed that the CAF surface specific markers including α-SMA and fibronectin were positive, and mainly were the cell membrane staining. The proliferation and invasion of Huh7 and HepG2 were significantly increased by CAF. The results show that the increasing percentage of cells in 24 hours between blank group and the experimental group was (63 ± 4) %, (78 ± 5) % and (69 ± 5) %, (81 ± 3) %respectively after co-culture with CAF, the difference was statistically significant (P<0.05). In transwell model, the number of cells in the blank group and the experimental group was (59.4 ± 3.1), (162.9 ± 3.9) and (104.8 ± 2.6), (166.4 ± 4.2), and the difference was also statistically significant (P< 0.05). Conclusion The isolated CAF from HCC enhances the ability of tumor's proliferation and invasion.
3.Efficacy Comparison among Three Kinds of Surgical Operation for Type 2 Diabetes Mellitus in Rats
Jia CHEN ; Xiao WANG ; Yongbin ZHANG ; Xin GAO ; Chuanlan SANG ; Yuan CHEN ; Haoran DONG ; Chongbo CAO
World Science and Technology-Modernization of Traditional Chinese Medicine 2013;(4):764-769
This study was aimed to compare the efficacy of gastric banding (GB), Roux en-Y gastric bypass (RYGBP) and biliopancreatic diversion (BPD) in the treatment of rats with type 2 diabetes mellitus (T2DM). Ani-mal models of T2DM were induced by streptozotocin (STZ) injection and high-sugar-fat diets. A total of 70 T2DM rats were randomly allocated into the GB group (G group, n = 20), RYGBP group (R group, n = 20), BPD group ( B group , n = 20 ) , and the sham operation group ( S group , n = 10 ) . The fasting blood glucose ( BG ) , triglyceride ( TG ) , total cholesterol ( TC ) and insulin ( INS ) content were determined before and 1 , 2 , 3 , 4 , 8 , 16 weeks after operation. The insulin sensitivity index (ISI) was calculated. The mortality and complications were ob-served in each group. The results showed that the fasting weight of the GB group, RYGBP group and BPD group were (324.4 ± 22.5) g, (338.9 ± 17.5) g, (333.3 ± 28.4) g, respectively. The BG content was (12.44 ± 1.29) mmol/L, (9.70 ± 0.81) mmol/L, (11.93 ± 2.39) mmol/L, respectively. The TC content was (2.32 ± 0.45) mmol/L, (2.22 ± 0.79) mmol/L, (2.13 ± 0.31) mmol/L, respectively. The TG content was (1.38 ± 0.32) mmol/L, (1.16± 0.41) mmol/L, (1.23 ± 0.35) mmol/L, respectively. The ISI were (-6.38 ± 0.29), (-6.67 ± 0.24), (-6.65 ±0.23), respectively. And the INS content of the RYGBP group were (69.43 ± 18.73) mU/L. There were signifi-cant differences between before and after operation on the 16th week ( P < 0 . 05 , P < 0 . 01 ) . The mortality rate was 5% in the GB group, 20% in the RYGBP group, and 35% in the BPD group. It was concluded that the GB, RYGBP and BPD are effective in reducing blood glucose and blood lipids in the treatment of rat with T2DM. The treatment effect is obvious in the improvement of insulin resistance ( IR ) .
4.Establishment of a rabbit model of heart failure by aortic regurgitation
Xiao WANG ; Haoran DONG ; Xihuan MA ; Ludi FU ; Chuanlan SANG ; Wang LIANG
Acta Laboratorium Animalis Scientia Sinica 2015;(2):127-131
Objective To establish a New Zealand rabbit model of heart failure by aortic regurgitation.Methods Adapting catheterization-induced aortic regurgitation to establish a volume overloat rabbit model of heart failure.The SBP, LVSP, LVDP, LV+dp/dt and LV ±dp/dt were observed before and after modeling.The successful criteria of heart failure:the LV ±dp/dtmax was decreased more than 40%and the LVDP increased more than 40%, or the LV ±dp/dtmax fell down to less than 40%and the DBP should be decrease more than 40%.Evaluating the model by observing the coat color, mental status, physical activity, calculating the feed consumption index, weight gain index, heart rate, respiration frequency and other indicators.The activity of serum SOD and MDA concentration were assayed to determine the antioxidant capacity of the model animals.Enzyme-linked immunosorbent assay kit was used to detect the serum cAMP and cGMP con-centration.Gene chip technology was used to analyze the difference of gene expression.Results After modeling, the he-modynamic index of SBP, DBP and LVSP were significantly decreased, LVDP was significantly decreased, LVDP was sig-nificantly increased and the LV+dp/dt and LV ±dp/dt were significantly decreased.Compared with the normal control group, the model animals showed coat withered, less movement, less eating, unresponsiveness, listlessness, and reduced grab resistance after modeling.The respiratory rate of the model group was significantly increased, and this trend was in-creased over time.The serum SOD activity was lower, MDA concentration was higher, cAMP concentration was lower, and cGMP concentration was higher in the model group.665 differentially expressed genes were detected.Compared with the human gene sequences, 16 characteristic genes were obtained.In these 16 genes, which were closely related to heart func-tion, were mainly related to ion channels, muscle contraction, and signal transduction function.Conclusions This repor-ted method to establish rabbit model of heart failure by using aortic regurgitation is reliable.The aortic regurgitation increa-ses cardiac preload, than leads to an increase of the left ventricular end-diastolic volume, and finally results in left ventric-ular hypertrophy and heart failure.The results of myocardial tissue gene chip test show that there are some changes in gene expression of the model rabbits.
5.Effects of medicated serum of the Zhuangguqiangjin tablets on the proliferation and osteogenic differentiation of bone marrow stromal stem cells of aged rats
Zhifeng ZHANG ; Jia CHEN ; Weihong YE ; Pei LI ; Haoran DONG ; Zhongyuan HONG ; Zhixiong HE
Chinese Journal of Primary Medicine and Pharmacy 2013;20(12):1765-1768,1921
Objective To study the effects of serum of the Zhuartgguqiang jin tablets on the proliferation and osteogenic differentiation of bone marrow stromal stem cells (BMSCs) of aged rats.Methods The BMSCs were obtained from male SD rats of 18 months.The cell surface markers CD34,CD31,CD29 were detected by flow cytometry.The proliferation of BMSCs treated by different concentration of medicated serum (2.5%,5%,10%) at 24,48,72hours was detected by CCK-8 method.The BMSCs were divided into 4 groups,which were osteogenic induction group,the Chinese medicated osteogenic induction group,Chinese medicine group,the blank group.After the BMSCs were cultured for 7 days,alkaline phosphatase (ALP) vitality was detected.After the BMSCs were cultured for 14 days,the BMSCs were stained by alizarin red,the mRNA of ALP and osteocalcin(OC) was detected.Results The expressionof CD29 was positive,and the expression of CD31 and CD34 were negative.2.5%,5% of the medicated serum could promote cell proliferation.The ALP vitality of osteogenic induction group[202.76 ± 15.44(U/gprot)],the Chinese medicated osteogenic induction group [240.48 ± 18.55 (U/gprot)],Chinese medicine group [178.87 ± 17.29 (U/gprot)]were significantly higher than that of blank group [111.24 ± 20.71 (U/gprot)] (t =22.50,7.985,3.535,all P <0.01).The ALP activity of Chinese medicated osteogenic induction group was higher than that of osteogenic induction group (t =3.103,P < 0.05).The ALP gene relative OD value of osteogenic induction group (0.40 ± 0.20) and Chinese medicated osteogenic induction group (0.60 ± 0.06) were higher than the blank group (0.09 ± 0.03) (t =2.372,9.547,all P <0.01).The ALP gene expression of Chinese medicated osteogenic induction group was obviously higher than that of the osteogenic induction group(P <0.05).The OC gene relative OD value of osteogenic induction group(0.58 ± 0.09) and Chinese medicated osteogenic induction group(0.76 ± 0.11) were higher than the blank group(0.41 ± 0.02) (P < 0.05,P < 0.01).The OC gene expression of Chinese medicated osteogenic inductiongroup was obviously higher than that of the osteogenic induction group(t =2.673,P < 0.05).Conclusion Medicated serum of Zhuangguqiangjin tablets can promote the proliferation and osteogenic differentiation of BMSCs of aged rats,which may be the mechanism of prevention senile osteoporosis.
6.The effects of nourishing lung and kidney formulas on inflammatory response of alveolar epithelial cells stimulated by monocytes conditioned medium
Yanqin QIN ; Haoran DONG ; Jiansheng LI ; Yulong CHEN ; Xiaoning MAO ; Mingming WU
Chinese Journal of Integrated Traditional and Western Medicine in Intensive and Critical Care 2017;24(1):63-67
Objective To observe the effects of nourishing lung and kidney formulas on inflammatory response of alveolar epithelial cells stimulated by monocytes conditioned medium and study the anti-inflammatory mechanism of the formulas for the treatment of chronic obstructive pulmonary diseases (COPD).Methods The reproduction of inflammation models of A549 cells were stimulated by monocyte THP-1 cell strain conditioned medium.A549 cells were randomly divided into blank control group (20% blank rabbits serum),model group (20% blank rabbits serum+25.0% THP-1 cell conditioned medium),nuclear transcription factor activator protein-1 (AP-1) pathway inhibitor T-5224group (20% blank rabbits serum+25.0% THP-1 cells conditioned medium+100 μmol/L T-5224),nourishing lung and kidney group (20% rabbits serum with nourishing lung and kidney formulas+25.0% THP-1 cells conditioned medium).The contents of interleukins (IL-6,IL-8),tumor necrosis factor-α (TNF-αα),matrix metalloprotein-9 (MMP-9) in cell culture supernatant were detected with enzyme linked immunosorbent assay (ELISA),the supernatant content of malondialdehyde (MDA) was detected with thibabituric acid (TBA) method,the total activity of superoxide dismutase (T-SOD) was detected with hydroxylamine method,and the activity of AP-1 pathway was detected with electrophoretic mobility shift assay (EMSA) method.Results Compared with the blank control group,the A549 cell proliferation were significantly increased at 24 hours,48 hours stimulation by 25.0% cell conditioned medium (A value:24 hours was 0.41 ± 0.02 vs.0.37 ± 0.04,48 hours was 1.30 ± 0.09 vs.1.15 ± 0.19).Compared with the blank control group,the contents of IL-6,IL-8,TNF-αα,MMP-9,MDA,AP-1 expression were significantly increased in model group [IL-6 (ng/L):35.00±3.63 vs.23.15±1.72,IL-8 (ng/L):273.09± 164.36 vs.231.45±33.90,TNF-α(ng/L):51.61 ± 9.51 vs.28.87 ± 3.34,MMP-9 (ng/L):442.85 ± 78.86 vs.235.60 ± 14.62,MDA (μmol/L):6.90 ± 0.11 vs.6.01 ± 0.12,AP-1 expression (A value):2.260 ± 0.062 vs.1.000 ± 0.000],MDA/T-SOD ratio was increased (4.43 ± 0.05vs.3.96 ± 0.06).Compared with model group,the levels of IL-8 (ng/L:100.29 ± 17.03),TNF-α (ng/L:25.13 ± 0.46),AP-1 expression (A value:1.38 ± 0.02),and the MDA/T-SOD ratio (4.23 ± 0.23) in T-5224 group,and MMP-9 (ng/L:195.44±9.80),MDA (μmol/L:5.86±0.30),MDA/T-SOD ratio (3.56±0.41),AP-1 expression (A value:0.76 ± 0.01) in nourishing lung and kidney group were all reduced significantly (all P < 0.05).Conclusion Nourishing lung and kidney formulas can suppress the inflammatory response through regulating the alveolar epithelial cells AP-1 signaling pathways.
7.Comparison of the hemostatic effect of total knee arthroplasty in intravenous and intra-articular injection of tranexamic acid
Haoran WANG ; Jinbo DONG ; Jianghua LI ; Jian YANG ; Weiguo ZHU ; Jianhua SUN ; Bin HE
Chongqing Medicine 2016;45(30):4249-4251
Objective To find a more suitable approach for the application of tranexamic acid(TXA)on total knee arthroplas‐ty (TKA) .Methods Totally 60 patients who met the inclusion criteria from January 2014 to August 2015 in the First Affiliated Hospital of Shihezi University were selected and divided into two groups according to the different route of administration .Group A (n=30) was intravenously injected with 100 mL TXA ,and group B(n=30) was locally injected with 100 mL TXA .Three hours drainage tubes occlusion were carried out after operation in the two groups .The intraoperative and postoperative dominant blood loss ,hidden blood loss indexes and the amount of total blood loss were recorded ,and coagulation indexes and D‐2 polymer were reg‐ularly monitored ,the incidence of thrombosis and postoperative adverse events were also observed .Results The amount of total blood loss in group B[(895 .41 ± 239 .02)mL] was lower than that in group A[(1 020 .89 ± 210 .83)mL] ,and the difference was statistically significant (P<0 .05);the total drainage volume in group B[(294 .33 ± 54 .25) mL] was lower than that in group A [(373 .33 ± 48 .02)mL] ,and the difference was statistically significant (P<0 .05) .After operation ,there was no significant differ‐ence between the two groups in coagulation indexes ,D‐2 polymer and the amount of hidden blood loss (P>0 .05) .No blood trans‐fusion ,symptomatic deep venous thrombosis and fatal pulmonary embolism occurred in the two groups .Conclusion The hemostatic effect of local application of TXA is better than that of intravenous injection in patients′initial TKA .
8.Pedicle subtraction osteotomy for repairing thoracolumbar degenerative kyphosis:correlation of spino-pelvic parameters
Haoran LI ; Qing CUI ; Zhanyin DONG ; Jianhua ZHANG ; Haiqing LI ; Ling ZHAO
Chinese Journal of Tissue Engineering Research 2017;21(3):345-349
BACKGROUND:Thoracolumbar degenerative kyphosis could experience severe lumbar back pain due to sagittal plane imbalance, thereby affecting quality of life. Thus, it is very important to reconstruct spino-pelvic profile in these patients. OBJECTIVE:To explore the relationships between life quality and spino-pelvic parameters fol owing pedicle subtraction osteotomy for thoracolumbar degenerative kyphosis and the clinical significance. METHODS:Between May 2010 and October 2014, 59 patients with thoracolumbar degenerative kyphosis undergoing L2 pedicle subtraction osteotomy in Cangzhou Hospital of Integrated Traditional and Western Medicine were retrospectively reviewed. Anteroposterior and lateral X-ray films of al patients during standing were photographed before and after surgery. The pre-and post-operative thoracic kyphosis, lumbar lordosis, sagittal imbalance, T1 pelvic angle, pelvic incidence, sacral slope and pelvic tilt were measured in al patients. The patients’ quality of life was evaluated using SF-36 preoperatively and postoperatively. RESULTS AND CONCLUSION:(1) Significant differences were observed in the improvement of thoracic kyphosis, lumbar lordosis, pelvic tilt, sacral slope and sagittal imbalance (P<0.01). With respect to SF-36, postoperative SF-36 score was higher than preoperative postoperative SF-36 score (P<0.01). (2) The alteration of lumbar lordosis showed significant correlation with the change of pelvic tilt, sacral slope and sagittal imbalance. The change of pelvic tilt exhibited cardinal correlation with the change of sacral slope, body pain and general health. The improvement of sagittal imbalance significantly correlated with the improvement of lumbar lordosis, body pain and general health. The improvement of T1 pelvic angle significantly correlated with the improvement of lumbar lordosis, sagittal imbalance, body pain and general health. (3) Pedicle subtraction osteotomy can effectively restore spino-pelvic sagittal profile, improve the life quality and relieve pain for the patients with thoracolumbar degenerative kyphosis.
9.Screening and target gene mapping of differentially expressed microRNAs in the degenerative intervertebral disc of degenerative scoliosis patients
Haoran LI ; Qing CUI ; Zhanyin DONG ; Jianhua ZHANG ; Haiqing LI ; Ling ZHAO
Chinese Journal of Tissue Engineering Research 2015;(46):7396-7400
BACKGROUND:MicroRNAs (miRNAs) play an important role in many diseases. To analyze the miRNA expression profile in degenerative scoliosis patients is helpful for classifying its pathogenesis. OBJECTIVE: To compare miRNAs expression profile in the intervertebral disc tissue between degenerative scoliosis patients and healthy controls, and to investigate its role in the pathogenesis of degenerative scoliosis. METHODS:Degenerative nucleus pulposus tissues from 48 patients with degenerative scoliosis (male 36, female 12; 58-69 years old) and normal nucleus pulposus tissues from 36 patients with lumbar fractures were harvested to isolate, culture and identify nucleus pulposus cels folowed by total RNA extract. Differentialy expressed miRNAs were screened by microRNA microarray analysis and validated by real-Time qPCR. Target genes of highly expressed microRNAs were predicted by analyzing information from MicroCosm v5, TargetScan 5.1 and microRNA.org databases. Biological signal pathways associated with the target genes were analyzed, and qPCR was used to validate the screening results. RESULTS AND CONCLUSION:Nineteen differentialy expressed miRNAs were identified. The miR-98 was highly expressed in degenerative nucleus pulposus tissue, and the fold change was 6.368. Predicted miR-98 target gene was interleukin-10, which was involved in JAK-STAT signaling pathway and located in upstream of this pathway. In degenerative nucleus pulposus cels of degenerative scoliosis patients, miR-98 was highly expressed, and the corresponding target gene was interleukin-10. These results indicate that JAK-STAT signaling pathway may play an important role in the pathogenesis of degenerative scoliosis.
10.The influence of application-audit system for doctoral enrollment on scientific research ability and employment development of medical graduate students
Mozhi WANG ; Haoran DONG ; Chong WANG ; Yingying XU
Chinese Journal of Medical Education Research 2022;21(9):1131-1138
Objective:To investigate and analyze the differences of PhD student scientific research ability, employment and development among types of postgraduate students that admitted by various of doctoral admissions under the perspective of medical students to assess and optimize the existing applications-audit system (AAS).Methods:A self-compiled questionnaire was used to investigate the opinions of medical students at different grades and types on the AAS related issues. Excel and SPSS were used to perform frequency statistics, chi-square test and independent sample t-test. Results:The PhD enrolled from AAS tended to complete the paper-publication at first and second year (38.71%, 12/31; 35.48%, 11/31), while unified enrollment (UE) tended to complete in the second year (33.33%, 10/30) and seven-year degree (SYD) in the third year (40.54%, 15/37) ( P=0.021). In terms of the value of the materials required in the application stage to the doctoral research, 74.19%(23/31) of AAS students and 48.65%(18/37) of SYD students believed that scientific research results were the most helpful to the doctoral research process, while UE believed that English helped the most (26.67%, 8/30), with significant differences among groups ( P=0.002). In terms of fair recognition of enrollment in the application system, 71.53% (304/425) of graduate students believed that AAS for doctoral enrollment was fair. The institution ( P=0.001) and type of master's degree ( P=0.001) significantly affected graduate students' recognition of the fairness of AAS. Conclusion:The degree and speed of dissertation completion of the AAS PhD are prior, reflecting the advantages of AAS in the cultivation of medical doctors' scientific academic ability and achievements. The AAS plays a significant role in enhancing the reserve of high-level scientific researchers in China, and has a high recognition and influence in medical students. In the future, the application examination system will be optimized to create the most suitable way for doctoral enrollment in China.