Low back pain has a serious impact on people's lives in t he world.With the approaching of aging society in China,the number of low back pain increased rapidly.Though many of the pathological factors in spine might lead to low back pain,Modic changes (MCs) was one of the most important factors.Therefore,to get familiar with MCs in the etiology and pathology has a positive practical significance.Although a large number of studies have analyzed the epidemiological and imaging characteristics of MCs,there are relatively few studies on the histological and pathological fields of MCs.In recent years,researchers have gradually shed some light on etiology of MCs and found that low toxic bacterial infection and autoimmune reaction may play a vital role in the development of MCs.The damage of vertebral body or endplate is closely related to the etiopathogenesis of MCs.In addition,to reveal the pathological characteristic of MCs is also a good attempt for better understanding of MCs.There are various imaging changes in MCs,however,what is the pathological basis behind it.It is also unclear that why different types of MCs are capable of transformation? When MCs occur,the activation of some cytokines (such as IL-6,RANKL,etc.) and certain receptors (TLR,PPARγ,etc.) can interfere with the formation and metabolism of the nearby bone marrow cells,break the balance of formation and absorption in trabecular beam and further change their microstructures through various pathways.During this dynamic balance process,the different types of MCs can be transformed into each other.The specific type may not only depend on the intensity and duration of inflammatory stimulus,but also rely on the composition and metabolic status of the bone marrow.Although the study of animal models has made some progress,some limitations still exist in practical course.Due to the unclear mechanism of MCs,the study on etiology and pathology of MCs may play a great important role in clarifying this issue.

Objective To reveal and compare the immunogenicity of glycoprotein from nucleus pulposus and collagen from anulus fibrosus in rabbit intervertebral disc. Methods According to the similarity principle of genome biology and the amount of nucleus pulposus in different intervertebral disc, Glycoprotein from nucleus pulposus was harvested from the intervertebral disc of 20 rabbits and placed into the psoas by anterolateral approach. Similar implantation of collagen from anulus fibrosus was performed in another 20 rabbits. The implantations of autologous muscle in rabbits serve as a control group.After one week,two weeks, four weeks, six weeks and eight weeks, the rabbits were killed, the tissue was harvested. HE staining was applied to detect the degree of vascularization and infiltration of lymphocytes. Immunohistochemistry were used for analysis of T-helper cells (CD4+) and cytotoxic T cells (CD8+) in the tissue. Results HE staining showed the infiltration of lymphocytes was found just one week after implantation and vascularization was seen at the 2nd week in the glycoprotein implanted group. However in collagen implant group,small amounts of lymphocytic and atypical vascularization were seen at the 4th week. Immunohistochemistry revealed that the proportion of activated T cells (CD4+ and CD8+) was significantly higher in the glycoproteinimplanted group at all time points, as compared with the collagen-implanted group and sham group. The proportion of activated T cells was higher in collagen-implated group than that of in sham group . Conclusion The glycoprotein in nucleus pulposus is easier to induce the differentiation and mature of T cell. The antigenicity of glycoprotein in nucleus pulposus was stronger than that of collagen in anulus fibrosus in rabbit intervertebral discs.

Objective To screen and validate differentially expressed microRNAs in human degenerative nucleus pulposus cells (NPCs) and to predict their target genes,and to investigate the role of JNK pathway in degenerative intervertebral disc disease.Methods Eight degenerative nucleus pulposus tissues were harvested from 8 patients (6 males and 2 females) with lumbar degenerative disease,and three normal nucleus pulposus tissues were harvested from 3 patients (3 males) with lumbar fracture,intraoperatively.Differentially expressed microRNAs were screened by microRNA microarray analysis and validated by real-time qPCR.Target genes of highly expressed microRNAs were predicted by analyzing information from data bases:MicroCosm v5,TargetScan 5.1 and microRNA.org.Signal pathways associated with the target genes were analyzed,and qPCR was used to validate the screening results.Results Twenty differentially expressed microRNAs were identified.The microRNA-513a-5p and microRNA-494 were highly expressed in degenerative nucleus pulposus tissue,which was corresponding to the verification results,and the ratio was 2.222 2 and 2.948 5,respectively.Target genes of microRNA-513a-5p and microRNA-494 were MKK4 and JunD,respectively.MKK4 and JunD were both involved in JNK signaling pathway and located in upstream and downstream of this pathway,respectively.Conclusion In human degenerative nucleus pulposus cells,microRNA-513a-5p and microRNA-494 are highly expressed microRNAs,whose target genes are MKK4 and JunD,respectively.Both MKK4 and JunD are involved in JNK signaling pathway.These results indicate that JNK pathway may play important role in the pathogenesis of degenerative intervertebral disc disease.

Objective To investigate the possibility of establishing a Modic changes (MCs) animal model,and explore the pathogenesis of MCs through imaging,histology and molecular biology experiments.Methods Fifty four New Zealand rabbits (weight 2.5-3.0 kg,half male and half female) were randomly divided into 3 groups:sham group (n=l8),muscle embedment group (n=18) and NP embedment group (n=18).In NP embedment group,the L4-5 and L5-6 discs were exposed by the lumbar anterolateral surgical approach.A 16 G needle was used to puncture the L5-6 vertebral body close to the epiphyseal plate.The depth of the drilling was approximately 3 mm.A 5 ml syringe was then put into the L4-5 intervertebral disc and extracted the NP,which was injected into the drilled hole of the vertebral body.The muscle embedment group and sham group shared the same operating procedures and drilling methods with the NP embedment group.Some pieces of muscle acquired from paraspinal muscles were put into the drilled hole in muscle embedment group,while nothing was put into the drilled hole in sham group.After that,the bleeding stopping,tissue washing and suture were done in all groups.12 weeks,16 weeks and 20 weeks after the surgery,MRI scan was applied to each group.All the specimens were tested by HE staining,real-time fluorescence quantitative PCR and Western blot to observe the expression of inflammatory cytokines.Results After modeling for 12 weeks,16 weeks and 20 weeks,MRI showed low signal changes on T1WI and mixed high signal in the context of low signal changes on T2WI in the NP embedment group.However,the muscle embedment and sham group showed no significant signal changes.Gross observation and HE staining confirmed that there was abnormal tissue proliferation in the imbed site of the NP embedment group.RT-PCR and Western blot showed high expression of IL-4,IL-17 and IFN-γin the NP embedment group,which were positively correlated with the length of the postoperative period.There was no significant difference between the muscle embedment group and sham group.Conclusion The puncturing of vertebral body close to endplate and putting nucleus into it can create an animal model of MCs.Autoimmune factors may play an important role in MCs.

In order to develop a combined live vaccine that will be used to prevent against porcine parvovirus (PPV) and Pseudorabies virus (PRV) infection, the VP2 gene of PPV was inserted into the transfer vector plasmid pG to produce the recombinant plasmid pGVP2. The plasmid pGVP2 and the genome of PRV HB98 attenuated vaccine were transfected by using lipofectamine into swine testis cells for the homologous recombination. The recombinant virus rPRV-VP2 was purified by selection of green fluorescence plaques for five cycles. 6-week-old female Kunming mice were immunized intramuscularly with attenuated PRV parent HB98 strain, commercial inactivated vaccine against PPV, recombinant virus, DMEM culture solution. The injections were repeated with an equivalent dose after 2 weeks in all of the groups, and then challenged with the virulent PRV NY strain at 7 weeks after the first immunization. The recombinant virus rPRV-VP2 was successfully generated, and the recombinant virus could effectively elicite anti-PPV and PRV antibody and significant cellular immune response as indicated by anti-PPV ELISA and HI, PRV-neutralizing assay and flow cytometry. The challenge assay indicated that recombinant virus could protect the mice against the virulent PRV challenge. These results demonstrated that the recombinant virus can be a candidate recombinant vaccine strain for the prevention of PRV and PPV.
Animals ; Antibodies, Viral ; immunology ; Antigens, Viral ; administration & dosage ; genetics ; immunology ; Capsid Proteins ; administration & dosage ; genetics ; immunology ; Female ; Gene Expression ; Genetic Vectors ; genetics ; metabolism ; Herpesvirus 1, Suid ; genetics ; metabolism ; Mice ; Parvovirus, Porcine ; genetics ; immunology ; Swine ; Swine Diseases ; immunology ; prevention & control ; virology ; Viral Vaccines ; administration & dosage ; genetics ; immunology

Objective To investigate the effect of tension stress on the changes of mRNA expression of interleukin-17B (IL-17) and inducible nitric oxide synthase (iNOS) in rat discs.Methods Sixty-four female Sprague-Dawley rats were randomly divided into 4 groups:sham (S),tail-suspended (TS),tail-suspended with needle puncture (TSNP) and single needle puncture (SNP) groups.Tail-suspension device made rats of TS group and TSNP group hindlimb suspended,which exerted tension force on the coccygeal vertebrae disc;and aseptic needle puncture on the tail disc between the seventh and eighth coccygeal vertebrae (Co7/Co8)of TSNP,SNP group induced the inflammatory environment.The discs were harvested 4 weeks later.Measurements included gross observation,histological observation and real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR) analysis.Results Gross and histological examination found that varying degrees of degenerative changes in the annulus and nucleus in TS,SNP and TSNP groups,but not in the S group.RT-qPCR results showed that the mRNA of IL-17B and iNOS were significantly higher in the TSNP and SNP groups than that in the S group.The values of 2-△△Q were 12.99 and 8.12 in TSNP groups,and the 2-△△Q values of SNP group demonstrated 22.49 and 14.98 compared to S groups.Sole application of tension stress brought by the tail-suspension slightly enhanced the expression of IL-17B and iNOS,the values of 2-△△Q were 1.22 and 1.49,which didn't reach a statistically significant change.Compared with SNP group,the mRNA expression of IL-17B and iNOS in TSNP groups were shapely decreased,the values of 2-△△Q were-9.50 and -6.86 respectively.Conclusion Relatively low magnitude tension stress might play a key role in anti-inflammatory process and the relief of discogenic pain.

Objective To observe the expression of Interleukin-17 (IL-17) in different types of herniated disc tissue,and to investigate Modic changes in corresponding injured segment and relationship between Modic changes and herniated types.Methods Fifty patients with single level disc herniation were enrolled in this study.According to clinical diagnosis criteria,imaging (CT and MR) examination results,and the view in surgery process the patients were divided into 2 groups:herniation group(35 patients),degeneration group(15 patients).According to MRI,patients were divided into 2 groups:Modic changes group(14 patients),non-Modic changes group (36 patients).The disc tissues were observed by HE staining and immunohistochemical staining,and the expression of IL-17 was detected by using real-time quantitative PCR.Results Pathological examination revealed that there were more granulation tissue and neovascularization in herniation group than in degeneration group.Immunohistochemical result showed that there were more IL-17-positive cells in herniation group than in degeneration group; there were more IL-17-postive cells in Modic changes group than in non-Modic changes group.The expression of IL-17 mRNA in herniation group was 6.7701 times more than that in degeneration group.The expression of IL-17mRNA in Modic changes group was 4.2836 times more than that in non-Modic changes group. Conclusion The herniated disc nucleus pulposus can cause more prominent local inflammation.Autoimmune responses maybe play an important role in pathophysiological mechanisms of Modic changes.

Objective:to investigate the clinical feature and dynamic changes of the cervical dural sac and spinal cord during neck flexion in Hirayama disease(juvenile muscular atrophy of distal upper extremity).Methods:Clinical data were taken and MRI in neutral neck position and a fully flexed neck position were performed on 27 cases of Hirayama disease.Results:(1)All patients were consistent with the diagnostic criteria of Hirayama disease who had asymmetric muscular atrophy and weakness of the hand and forearm.All patients were young males and right handed of whom 77.8% had initial symptoms before they were 19 years old.More patients(20 cases,74%)had muscular atrophy in the right hand than in the left at onset.The duration after disease onset was from 2-72 months[(26.48?15.57)months].(2)In neutral neck position by MIR examination,16 patients showed abnormal cervical curvature,14 showed atrophy of the lower cervical cord and 2 patients had intramedullary abnormal high signal.(3)In a fully flexed position of the neck,all patients showed forward displacement and flattening of the lower cervical cord,and a crescent-shaped high signal area behind the cord.(4)The crescent-shaped area was enhanced on T1-weighed imaging and disappeared after the patient returned to a neural position in one case.Conclusion:Hirayama disease occurs mainly in young males.There are obviously dynamic changes of the cervical cord during neck flexion in Hirayama disease by MRI examination,which can help the doctor make diagnosis in the early stage.

Objective To find out the display rules of the key part of the proximal femur and fracture line and obtain the best viewing position and angle by placing the femoral neck at different positions and different angles through X-ray.Methods Six dry specimens of cadaveric femur were from Department of Anatomy,Tianjin Medical University.Three models of complete femoral neck model,tin line fracture model and steel saw fracture model were made respectively.The tin line fracture model was based on the Pauwells angle,using the tin wire(1mm) wrapped around the femoral necks to make three kinds of fracture models (Pauwells angle 30°,50° and 70°);steel saw fracture model was made by hacksaw,then reposition in situ,to make three kinds of fracture models same as the tin line fracture model.The projection manner included different positions and different angles,different projection positions include:parallel with the femoral shaft,perpendicular to the femoral shaft,parallel to the femoral neck and perpendicular to the femoral neck;different projection angles included:from 40°,30°,20°,15°,10° and 5° in the head side to 5°,10°,15 °,20 °,30 °and 40° in the foot side and vertical angle 0°.For the complete femoral neck model,we observed the imaging characteristics of the key parts of the femoral head and neck (tension trabecular bone and pressure trabecular bone;lesser trochanter;intertrochanteric line;length of neck of femur and femoral head shape),looking for the display rules,and obtained the best viewing position.For tin line fracture model and steel saw fracture model,we tried to find the best display angle and position by the different projection position and angle.Results The results of complete femoral neck model:Lesser trochanter:in perpendicular to the femoral shaft position showed the best,gradually increased with the foot side deflection;tension trabecular bone and pressure trabecular bone:in parallel with the femoral shaft position 10°on the head side;intertrochanteric line:no difference between parallel with the femoral shaft with perpendicular to the femoral shaft;Femoral neck shape:deflect to both sides,head of femur was out of shape.The results of tin line fracture model:the Pauwells angle 30° model showed the best position in parallel with the femoral shaft position 20° on the head side;the Pauwells angle 50° model showed the best position in parallel with the femoral shaft position 5° on the head side;the Pauwells angle 70° model showed the best position in parallel with the femoral shaft position 10° on the head side.The results of steel saw fracture model were the same with the tin line fracture model.Conclusion There was the best viewing angles and positions for the key anatomy of the proximal femur and different Pauwells angle classification of femoral neck fracture.The image doctor could make more accurate projection,according to the different types of femoral neck fracture.

Objective To investigate the expression and clinical implication of miR-494 and its target gene JunD in intervertebral disc herniation (IDH).Methods Six intervertebral disc tissue samples of spinal burst fracture were collected during operation,and then the nucleus pulposus cells were cultured.TNF-α of different concentration (0,10,50 and 100 ng/ml) were added to the cells to induce apoptosis.The apoptosis rate of nucleus pulposus cells and the expression of miR-494 were detected at different time after the stimulation (0,8,16 and 24 h) using qRT-PCR and flow cytometry respectively.Then,AntigomiR-494 was transfected into nucleus pulposus cells after lentivirus packaging,followed by the use of TNF-α (100 ng/ml,16 h) to induce apoptosis.The experiments contained blank control group,AntigomiR-494 + TNF-α group,and negative control + TNF-o group.Flow cytometry was used to detect the apoptosis in each group,and Western blot the expressions of JunD and cytochrome C.The luciferase double report based analysis and bioinformatics methods were used to investigate the relationship between miR-494 and JunD gene.Results The expression of miR-494 and the apoptosis rate of nucleus pulposus cells increased along with the increase of concentration of TNF-α and length of stimulation (P ＜ 0.05).After transfection,the expression of miR-494 and the apoptosis rate in AntigomiR-494 + TNF-α group were significantly lower than those in negative control + TNF-α Group (P ＜ 0.05).The results of Western blot showed that the expression level of JunD protein in AntigomiR-494 + TNF-α group was significantly higher than that of the negative control group (P ＜ 0.05),and the expression level of cytochrome C protein was significantly lower than that of the negative control + TNF-α group (P ＜ 0.05).Luciferase double report gene validation and bioinformatics prediction confirmed that miR-494 directly targeted JunD.Conclusion TNF-α can induce apoptosis of nucleus pulposus cells in a time and dose dependent manner.The expression of miR-494 increases with the increase of TNF-α concentration and length of stimulation.MiR-494 might be the key regulator of the apoptosis of nucleus pulposus cells under the induction of TNF-o.MiR-494 gene knockout can protect the nucleus pulposus cells,through the possible mechanism of upregulating target gene JunD and mediating the cytochrome C apoptosis pathway.MiR-494-JunD-cytochrome C signaling pathway might be one of the potential mechanisms of intervertebral disc degeneration.