Objective To study the nuclear protein association of high-mobility group box-1 (HMGB1) and histone deacetylase 1 (HDAC1),and the effect of interaction on radiosensitivity in human breast cancer cells.Methods The protein-protein interaction was determined by immunoprecipitationWestern blot and glutathione-S-transferase capture assays.Cell growth was examined by MTT (methyl thiazolyl tetrazolium)assay and clonogenic assay.Histone deacetylase activity was analyzed by histone deacetylase assay.Results A significant increase of HMGB1 protein and radiosensitivity was observed in MDA-MB-231 and MDA-MB-468 cells transfected with a pCMV-Tag2B expression vector carrying with a full-length of HMGB1 cDNA.HMGB1 binding to HDAC1 was demonstrated as GST (glutathione Stransferase)-pull down and immunoprecipitation Western blot assay,and the association was elevated by irradiation.An LXCXE motif was required for the HMGB1-HADC1 interaction and HMGB1 radiosensitization.A significant difference of IC50 value was observed,for example,1.8 and 2.2 Gy (wtHMGB1 transfectants,P ＜ 0.05),3.6 and 3.8 Gy (HMGB1/C103F transfectants,P ＞ 0.05),both compared with 3.9 and 4.1 Gy (pCMV-Tag2B transfectants) in MDA-MB-231 and MDA-MB-468 cells,respectively.A specific HDAC1 inhibitor trichostatin A markedly reduced the HMGB1-mediated radiosensitivity,0.5 Gy in the presence of trichostatin A versus 1.8 Gy in absence of trichostatin A in MDA-MB-231 transfectants,1.2 Gy (with trichostatin A) versus 2.2 Gy (without trichostatin A) in MDA-MB-468 transfectants,P ＜ 0.05.Histone deacetylase activity was also detected in immunoprecipitates prepared from these cells with antibodies to HMGB1,and this activity was abolished by the histone trichostatin A.Conclusions These results suggest a previous unanticipated role for HDAC1 in modification of HMGB1-mediated radiosensitivity by its direct interaction with HMGB1.

Objective To discuss the degenerative factors, the spinal segment distribution, and the mechanism in hyperextension injury of cervical spine. Method Eighty-nine patients with hyperextension injury of cervical spine were retrospectively analyzed by observing the degenerativelesion, the spinal cord segment with high signal in T2WI, and the location of facial trauma. Results Fifty-eight cases showed the disc hemiation which was the most common lesion, followed by 8 cases showing the calcification of the posterior longitudinal ligament. Besides, 7 cases presented the developmental stenosis of spinal canal, and also, 6 cases showed disc hemiation combined with the yellow ligament hypertrophy. The intervertebral level of the spinal cord with high signal in T2WI were distributed as follows:4 cases were at C2/3, of which onesuffered the forehead trauma; 12 cases were at C3/ 4, of which 10 had the forehead trauma, and one had the zygomatic trauma; 12 cases were at C4/5, of which 5 had the forehead trauma, one had both the zygomatic and the forehead trauma, and one had both the forehead and with the lower jaw trauma; 11 cases were at CS/6,of which 3 had the forehead trauma, 3 had the zygomatic trauma, and 2 had the lower jaw trauma. The location of the spinal cord with single high signal in T2WI did not correspond with the intervertebral disc level in 4 cases. For 10 cases the high signal in T2WI was found at two discontinuous segments. For 2 cases the 1 high signal in T2WI was found at over two segments. For 6 cases the high signal in T2WI was found at over three segments. Conclusions Disc hemiation is the most common underlyding factor in cervical hyperextension injury. The spinal level with high signal in T2WI was correlative to the impacted facial site. The shear force at the inflection point with or without the anterior-posterior compression force accounted for the cervical hyperextension injury.

Objective To evaluate the roles of high-resolution computer tomography (HRCT) and bron- choalveolar lavage (BALF) in the diagnosis of connective tissue diseases associated with interstitial lung disease (CTD-ILD). Methods Clinical data of chest HRCT and BALF of patients with CTD-ILD from January 1997 to December 2007 in in-patient department of Peking University First Hospital, were retrospectively analyzed. Results ①Among 46 cases with the picture of chest HRCT, 19 (41.3%)showed usual interstitial pneumonia(UIP) -like pattern and 18 (39.1%) showed lobular and interlobular septa thickening. 8 (8/17) of ANCA vasculitis (AASV) and 5 (5/9) of rheumatoid arthritis (RA) manifested as UIP-like patterns respectively. In polymyositis/dermatomyositis(PM/DM) and Sjogren's syndrome (KS) patients, the organizing pneumonia(OP)-like pattern and lymphocytic interstitial pneumonia(LIP)-like pattern were 2/4 and 2/4 respectively. ②Among 32 cases undergoing BAL, 10/10 patients with AASV-ILD all showed that neutrophils were dominant in BALF, while, the other 22 patients showed that the ratio of neutrophils elevation (14/22, 63.6%) and the ratio of lymphocytes elevation (18/22, 81.8%) were comparable, and there were 12/22(54.5%)patients with both types of cell elevation. Among 13 cases with iymphocytes elevation in BALF who performed analysis of sub-type lymphocytes, 10/13 cases showed decreased CD4/CD8 ratio, 3/13 cases showed increased CD4/CD8 ratio which were all related with SS. ③Among 15 patients undertaken HRCT and BALF detection together, 7/7 UIP-like cases showed the ratio of neutrephils elevation in BALK While in non-UIP-like cases, 5/8 showed the ratio of lymphocytes elevation. Conclusion ①UIP-like patterns and patterns of lobular and interlobular septa thickening are the most common imaging features of HRCT in CTD-ILD, the former are mostly seen in AASV and RA. OP-like patterns and LIP-like patterns are commonly seen in PM/DM and SS respectively. ②Tbe increased neutrophil percentage is dominant in BAL fluid of patients with AASV-ILD, while the others show that the ratio of neutrophil and lymphocyte elevation are comparable, lymphocytes subtype analysis shows decreased CD4/CD8 ratio is dominant in CTD-ILD patients with lymphocytes increased. There is a significant relationship between increased CD4/CD8 ratio and SS-LIP. ③All of the cases with UIP-like patterns show the ratio of neutrophils elevation in BALF. While the ratio of lymphocytes elevation is dominant in non-UIP-like cases.

Objective To study the regulation of dendritic cells by recombinant glycated polylysine-coupled MIP-3α-FL double-gene targeting expression vector in liver cancer immune microenvironment.Methods H22 hepatocarcinoma cells were transfected with recombinant plasmid of MIP-3α-FL (shMIP-3α-FL) and injected into hepatoma model mice.The survival time,tumor size were compared.Flow cytometry was used to measure the number and phenotype of tumor infiltrating DCs.Results Western blot and ELISA demonstrated that the secretion of MIP-3α and FL in H22 cells was significantly increased after transfection with MIP-3α-FL.The survival time of the mice in the experimental group was significantly prolonged,the tumor size decreased.Flow cytometry showed that the number of tumor-infiltrating DCs in the experimental group was significantly higher than that in the control group;the expression of CD80 and CD86 in the infiltrating dendritic cells (TIDCs) was significantly higher than that of the control group.Conclusions The co-action of MIP-3α and FL can significantly promote DC accumulation,maturation,and conjugate glycosylated polylysine carriers increase the precision of targeting and enhance the antigenpresentation of the DCs.

ObjectiveTo investigate the clinical features and treatment of ascending paralysis after thoracolumbar fracture.MethodsThree male patients with 2 fracture levels at T12 and one at L1 were retrospectively studied.Their mean age was 41.3 years(range,39-42 years).All 3 cases were undertaken open decompression,reduction and internal fixation.Paralysis level began to ascend at 2-5 days after injury,with 2 cases up to C2,3 and 1 case up to T7.Two patients suffered irritating pain over the paralysis level before onset of ascending.Postoperative MRI images demonstrated well reduction and no compression of spinal cord.In the early phase after ascending,MRI obviously showed swelling in spinal cord and long T1 and long T2 signals shaped patchy and stripy distribution in the central area.One patient's MRI displayed that the spinal cord shrinked 16 days after trauma with abnormal high signal in the central area.ResultsTwo cases died of respiratory muscle paralysis and 1 case suffered paraplegia with no recovery 5 years after surgery.ConclusionAscending paralysis after thoracolumbar fracture is a rare complication with very poor prognosis.MRI is available for evaluating operational effects and affected level.The exact mechanism and effective treatment are still unclear and need further investigated.

OBJECTIVE: To investigate the effect of high mobility group box-1 protein (HMGB1) on the proliferative activity of human hepatoma cell line HepG2 and its potential regulating mechanism. METHODS: The cultured HepG2 cells were treated with recombinant HMGB1 (0, 10, 50, and 100 ng/mL, respectively) for 24 h. Cell proliferation was observed by MTT analysis. Western blot and reverse transcriptase-polymerase chain reaction were used to detect the expression of proliferating cell nuclear antigen (PCNA) and cyclin D1 protein and mRNA, respectively. RESULTS: Compared with the control group, HMGB1 at 10, 50, and 100 ng/mL obviously increased HepG2 cells proliferation, cyclin D1 and PCNA protein and mRNA expression after the treatment for 24 h, respectively (P<0.05). Anti-HMGB1 significantly inhibited the proliferation and cyclin D1 and PCNA mRNA and protein expression of HMGB1 on HepG2 cells (P<0.05). CONCLUSION Proliferation of HMGB1 on HepG2 cells may be associated with increasing cyclin D1 and PCNA expression. Anti-HMGB1 may have a therapeutic effect on hepatocellular carcinoma.
Cell Proliferation ; drug effects ; Cyclin D1 ; genetics ; metabolism ; HMGB1 Protein ; pharmacology ; Hep G2 Cells ; Humans ; Proliferating Cell Nuclear Antigen ; genetics ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Recombinant Proteins ; pharmacology ; Reverse Transcriptase Polymerase Chain Reaction

Objective To study the effect of human CCR4-NOT transcription complex subunit 7 (CNOT7) gene knockdown on the immune microenvironment of HepG2 cells and explore its significance. Methods We designed a cell transfection protocol and performed the experiment with three groups:CNOT7-targeted knockdown group, control group, and CNOT7 overexpression group. The transfection efficiency was assessed using inverted fluorescence microscopy, and the expression level of CNOT7, transforming growth factor-β1 (TGF-β1), and nuclear factor-kappa B (NF-κB) p65 proteins was determined by Western blotting. The concentration of TGF-β1 secreted in the cell culture supernatant was measured by ELISA. The sensitivity of tumor cells to the killing function of natural killer (NK) cells was detected by flow cytometry. Results Compared with the control group, the expression level of TGF-β1 and NF-κB p65 proteins was significantly decreased in the CNOT7-targeted knockdown group, and the TGF-β1 concentration in the culture supernatant was also significantly reduced. However, in the CNOT7 overexpression group, the expression level of the two proteins and TGF-β1 concentration were significantly increased. NK cells were co-cultured with tumor cells, and the apoptosis rate of HepG2 cells transfected with CNOT7-specific shRNA was significantly increased. However, in the CNOT7 overexpression group, the apoptosis rate was significantly decreased. Conclusion CNOT7 forms the immune microenvironment of hepatocellular carcinoma. Targeted knockdown of CNOT7 can reduce TGF-β1 secretion and enhance the killing function of NK cells toward HepG2 cells.

Objective To analyze tumor immune microenvironment and related mechanisms in liver cancer.Methods We included 10 cases of hepatocellular carcinoma,hepatitis B patients and healthy volunteers from January 2015 to December 2017 in Shanxi Grand Hospital.We first detected the peripheral and local GM-CSF level in each group,detected myeloid-derived suppressor cells (MDSCs) GM-CSF and pathway-related protein expression.from liver cancer,tumor margin and normal liver tissue through flow cytometry and immunohistochemistry,Finally,we transfected the CCR4-NOT transcriptional complex subunit 7 (CNOT7) recombinant plasmid in the hepatoma cell line,and then detected the related protein expression.Results There was no significant difference for peripheral blood GM-CSF level between liver cancer group,hepatitis group and control group (P＞0.05).The level of local GM-CSF was (32.2±8.9) ng/L,which was higher than that of hepatocellular carcinoma (9.7±2.7) ng/L and normal liver tissue (11.6±2.9) ng/L.The difference was statistically significant (P＜0.05).The proportion of MDSCs at the edge of the tumor was (9.9 ±3.6) ％,which was higher than that of liver cancer (4.0± 1.5) ％ and normal liver tissue (6.3±2.3) ％,and the difference was statistically significant (P＜0.05).Immunohistochemistrydata was consistent with previous data.Compared with normal liver tissue,CNOT7 and STAT3 were highly expressed in liver cancer tissues,while STAT1 was lowly expressed.HepG2 human hepatoma cells were selected for transfection.Compared with the empty plasmid group,CNOT7 expression was decreased in the knocking out group at the same time STAT1 expression was increased,STAT3 and GM-CSF expression was decreased.Conclusion In hepatocellular carcinoma,the secretion of GM-CSF increased and the number of MDSCs increased.Knocking out CNOT7 reduced GM-CSF secretion and activate the JAK/STAT signaling pathway.

BACKGROUND:Anatomic medul ary locking (AML) femoral prosthesis is circular cylinder and has satisfactory efficacy. However, some scholars found the complications such as thigh pain, loss of bone at the proximal end of the femur, and wearing-related osteolysis. F2L femoral prosthesis is cone-shaped and also has satisfactory efficacy, but the thigh pain incidence is relatively low. ＜br> OBJECTIVE:To compare the intermediate-long term results of AML versus F2L in total hip arthroplasty. ＜br> METHODS:Between November 1997 and January 2005, we retrospectively reviewed 60 patients (66 hips) undergoing total hip arthroplasty using biological femoral prosthesis. At fol ow-up examination, 58 hips in 52 patients were available for clinical and roentgenographic review. 26 AML devices were placed in 24 patients, and 32 F2L devices were placed in 28 patients. The AML group were reviewed with an average of 12.7 years fol ow-up (range 10 years and 3 months to 15 years and 5 months), while the F2L group were reviewed with an average of 9.5 years fol ow-up (range 8 years and 3 months to 11 years and 1 month). The clinical results were evaluated with Harris methods and X-ray examination. Kaplan-Meier analysis was performed to evaluate the survival of femoral component. End point was radiographical loosening or revision of the femoral component for any reason. ＜br> RESULTS AND CONCLUSION:There were no significant difference between AML and F2L about Harris score in the latest fol ow-up (P>0.05). After surgery, the incidence of thigh pain was significantly lower in F2L group than that in AML group (P＜0.05). In AMKL group, the stress-shielding 1 level was observed in 21 hips (81%), and 2 level in five hips (19%);in F2L group, the stress shielding 0 level was observed in 20 hips (62%) and 1 level in 12 hips (38%). There were significant differences between the two groups (P＜0.05). The stress shielding showed significant differences between the two groups (P＜0.05). The incidence of osteolysis in F2L group was significantly lower than that in AML group (P＜0.05). Kaplan-Meier analysis showed that, the survival rate of both AML and F2L components were 1.0 (95%confidence interval:0.98-1.00). Experimental findings indicate that, both AML and F2L femoral prosthesis have a satisfactory long-term efficacy after total hip arthroplasty, and the incidence of thigh pain and osteolysis is significantly lower in F2L group.

Objective To investigate the immediate oral feedback after objective structure clinical examination (OSCE) for postgraduate year 1 & 2 surgery residents (PGY1 & 2).Methods 37 PGY1 and 38 PGY2 wereevaluated.The examination was composed of 6 stationsand limited to15 minutes per station.Each station was evaluated by centesimal system score.Immediate oral feedback was given in the last2 minutes.A questionnaire was given to each resident and examiner at the end of OSCE.All data analyses were conducted using SPSS version 22.0,repeated measures ANOVA and LSD test were used,and correlations were tested by the Pearson correlation test.Results The average scores for PGY1 & 2 were (68.97 ± 5.40) and (68.35 ± 5.00),the between-and inter-round differences in average score were not statistically significant.There was no significant correlation about theevaluation of the residents' performance during OSCE between the examiners and the residents.The necessity and effectiveness of immediate oral feedback were confirmed by both the examiners and the residents.Conclusions Immediate oral feedback isfeasible with limited impact on OSCE score,but the plan should be furtherrefined.Follow-up study isnecessary to identify the long-term effect on the clinical competency.