Adolescent ; Adult ; Child ; Child, Preschool ; Female ; Humans ; Male ; Manipulation, Orthopedic ; methods ; Radius Fractures ; therapy ; Ulna Fractures ; therapy

OBJECTIVETo evaluate the effect of interleukin-6 (IL-6) on the biological behaviors of the chondrocytes in the cartilage endplate of rabbits.

METHODSChondrocytes isolated from the cartilage endplate of New Zealand rabbits, verified for their biological characteristics by such means as toluidine blue staining for type II collagen, were treated with IL-6 at different concentrations. The proliferation of the chondrocytes was evaluated by MTT assay at different time points following the treatment, the cell cycle changes were determined by flow cytometry and the changes of aggrecan and type II collagen mRNAs detected by RT-PCR.

RESULTSAt the concentrations of 10, 50 and 100 ng/ml, IL-6 did not obviously affect the rate of chondrocyte proliferation. IL-6 at 50 ng/ml resulted in no obvious changes of the cell cycle of the chondrocytes, but significantly decreased the expression of collagen IIa mRNA.

CONCLUSIONIL-6 has no effect on the proliferation and cell cycle of the chondrocytes, but at higher concentrations, it inhibits matrix synthesis of the chondrocytes to promote intervertebral disc degeneration.

Aggrecans ; genetics ; Animals ; Cartilage ; cytology ; Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Chondrocytes ; cytology ; drug effects ; metabolism ; Collagen Type II ; genetics ; Female ; Gene Expression Regulation ; drug effects ; Interleukin-6 ; pharmacology ; Male ; RNA, Messenger ; genetics ; metabolism ; Rabbits

OBJECTIVETo analyze the risk factors of heterotopic (HO) ossification after total lumbar disc replacement and probe the preventive strategies for it.

METHODSThe radiographs and clinical data of 78 discs in 65 patients who received artificial lumbar disc replacement (ADR) from April 1998 to December 2003 were analyzed retrospectively by two radiologists and one orthopaedic surgeon and then postoperative HO were graded according to McAfee system. The bony formations in disc spaces, time of HO were found, and range of motion (ROM) of the operated levels were measured on radiographic films. In addition, the risk factors such as preoperative peri-annulus ossification, bony endplates injuries, and subsided or mal-position of the prostheses were also analyzed by Logistic regression analysis.

RESULTSPostoperative HO was found in 10 spaces of 9 cases. Class I of HO were occurred in 7 patients at means 2.1 years postoperatively with normal range of motion preserved. Three of them turned into class II or III with 10 degrees of mean ROM in the following 2.5 years. Another 2 (2/9) cases with preoperative peri-annulus ossification had bridging trabecular bone (class III) between the endplates and 9 degrees of ROM 2 years after surgery, then turned into class IV at 6 years with 0 degrees and 4 degrees of motion in the operated levels. As the risk factors of HO, preoperative annulus ossification (2 cases), bony endplates injuries (5 cases), mal-positioned prostheses (2 cases) and subsided prostheses (2 cases) were found simultaneity with significant positive relation to HO occurred (P < 0.05).

CONCLUSIONSFactors such as preoperative ossification of annulus, endplate injuries, prosthesis subsided and mal-position would have higher risks to have HO occurred after ADR, but ROM of most affected levels are preserved. Strict control indication and avoid all above risk factors can prevent HO occurring effectively.

Adult ; Aged ; Diskectomy ; Female ; Follow-Up Studies ; Humans ; Lumbar Vertebrae ; surgery ; Male ; Middle Aged ; Ossification, Heterotopic ; etiology ; prevention & control ; Prosthesis Implantation ; adverse effects ; Retrospective Studies ; Risk Factors

In the outbreak of COVID-19,triage procedures based on epidemiology were implemented in a local hospital in Changsha to control the transmission of SARS-CoV-2 and avoid healthcare-associated infection.This re-trospective study analyzed the data collected during the triage period and found that COVID-19 patients were en-riched 7 folds into the Section A designated for patients with obvious epidemiological history.On the other side,nearly triple amounts of visits were received at the Section B for patients without obvious epidemiological history.8 COVID-19 cases were spotted out of 247 suspected patients.More than 50％of the suspected patients were submi-tted to multiple rounds of nucleic acid analysis for SARS-CoV-2 infection.Of the 239 patients who were diagnosed as negative of the virus infection,188 were successfully revisited and none was reported as COVID-19 case.Of the 8 COVID-19 patients,3 were confirmed only after multiple rounds of nucleic acid analysis.Besides comorbidities,delayed sharing of epidemiological history added complexity to the diagnosis in practice.The triaging experience and strategy will be helpful for the control of infectious diseases in the future.

Objective: To investigate the effect of ubiquitin mutation at position 331 of tumor necrosis factor receptor related factor 6 (TRAF6) on the biological characteristics of colorectal cancer cells and its mechanism. Methods: lentivirus wild type (pCDH-3×FLAG-TRAF6) and mutation (pCDH-3×FLAG-TRAF6-331mut) of TRAF6 gene expression plasmid with green fluorescent protein tag were used to infect colorectal cancer cells SW480 and HCT116, respectively. The infection was observed by fluorescence microscope, and the expressions of TRAF6 and TRAF6-331mut in cells was detected by western blot. Cell counting kit-8 (CCK-8) and plate cloning test were used to detect the proliferation ability of colorectal cancer cells in TRAF6 group and TRAF6-331mut group, cell scratch test to detect cell migration, Transwell chamber test to detect cell migration and invasion, immunoprecipitation to detect the ubiquitination of TRAF6 and TRAF6-331mut with ubiquitinof lysine binding sites K48 and K63. Western blot was used to detect the effects of TRAF6 and TRAF6-331mut over expression on the nuclear factor kappa-B (NF-κB) and mitogen activated protein kinase mitogen-activated protein kinase (MAPK)/activating protein-1(AP-1) signal pathway. Results: The successful infection of colorectal cancer cells was observed under fluorescence microscope. Western blot detection showed that TRAF6 and TRAF6-331mut were successfully expressed in colorectal cancer cells. The results of CCK-8 assay showed that on the fourth day, the absorbance values of HCT116 and SW480 cells in TRAF6-331mut group were 1.89±0.39 and 1.88±0.24 respectively, which were lower than those in TRAF6 group (2.09±0.12 and 2.17±0.45, P=0.036 and P=0.011, respectively). The results of plate colony formation assay showed that the number of clones of HCT116 and SW480 cells in TRAF6-331mut group was 120±14 and 85±14 respectively, which was lower than those in TRAF6 group (190±21 and 125±13, P=0.001 and P=0.002, respectively). The results of cell scratch test showed that after 48 hours, the percentage of wound healing distance of HCT116 and SW480 cells in TRAF6-331mut group was (31±12)% and (33±14)%, respectively, which was lower than those in TRAF6 group [(43±13)% and (43±7)%, P=0.005 and 0.009, respectively]. The results of Transwell migration assay showed that the migration numbers of HCT116 and SW480 cells in TRAF6-331mut group were significantly lower than those in TRAF6 group (P<0.001 and P<0.002, respectively). The results of Transwell invasion assay showed that the number of membrane penetration of HCT116 and SW480 cells in TRAF6-331mut group was significantly lower than those in TRAF6 group (P=0.008 and P=0.009, respectively). The results of immunoprecipitation detection showed that the ubiquitin protein of K48 chain pulled by TRAF6-331mut was lower than that of wild type TRAF6 in 293T cells co-transfected with K48 (0.57±0.19), and the ubiquitin protein of K63 chain pulled down by TRAF6-331mut in 293T cells co-transfected with K63 was lower than that of wild type TRAF6 (0.89±0.08, P<0.001). Western blot assay showed that the protein expression levels of NF-κB, p-NF-κB and p-AP-1 in TRAF6-331mut-HCT116 cells were 0.63±0.08, 0.42±0.08 and 0.60±0.07 respectively, which were lower than those in TRAF6-HCT116 cells (P=0.002, P<0.001 and P<0.001, respectively). The expression level of AP-1 protein in TRAF6-HCT116 cells was 0.89±0.06, compared with that in TRAF6-HCT116 cells. The difference was not statistically significant (P>0.05). The protein expression levels of NF-κB, p-NF-κB and p-AP-1 in TRAF6-331mut-SW480 cells were 0.50±0.06, 0.51±0.04, 0.48±0.02, respectively, which were lower than those in TRAF6-SW480 cells (all P<0.001). There was no significant difference in AP-1 protein expression between TRAF6-331mut-SW480 cells and TRAF6-SW480 cells. Conclusion: The ubiquitin site mutation of TRAF6 gene at 331 may prevent the binding of TRAF6 and ubiquitin lysine sites K48 and K63, and then affect the expressions of proteins related to downstream NF-κB and MAPK/AP-1 signal pathways, and inhibit the proliferation, migration and invasion of colorectal cancer cells.
Humans ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Colorectal Neoplasms/pathology* ; Lysine/metabolism* ; NF-kappa B/metabolism* ; TNF Receptor-Associated Factor 6/metabolism* ; Transcription Factor AP-1/metabolism* ; Ubiquitin/metabolism*