Objective To analyze the characteristics of toxin, the PCR-ribotyping(RT) and the multilocus sequence typing(MLST) of Clostridium difficile strains isolated from China-Japan Friendship Hospital in order to provide a basis for monitoring the outbreak of nosocomial Clostridium difficile infection.Methods A total of 321 samples were collected from the patients with suspected Clostridium difficile infection(CDI) in China-Japan Friendship Hospital(CJFH) during 2012 to 2013.All Clostridium difficile strains were isolated and identified by the standard phenotypic culture method.Cytotoxicity test was performed to detect toxin B.Toxin genes (tcdA and tcdB) and binary toxin genes (cdtA and cdtB) harbored by those strains were analyzed.RT and MLST were used for homologous analysis.Clinical data of the patients were collected to analyze the isolation rate of Clostridium difficile in different populations.Results Forty-eight strains of Clostridium difficile were isolated from 46 patients with diarrhea and three of them were isolated from the same patient.The incidence of CDI among all patients, outpatients and inpatients were 14.3%(46/321), 12.8%(5/39) and 14.5%(41/282), respectively.Toxin B was detected in all of the strains as indicated by the cytotoxicity test.Strains of sequence type 1(ST1) showed the strongest cytotoxicity of all the isolated Clostridium difficile strains.Ten out of the 48 strains (20.8%) were tcdA(-)/tcdB(+) strains, which belonged to either ST37 or ST81.The results of RT and MLST were consistent in assigning the strains into nine types, in which the predominant type was ST1/RT027 accounting for 27.1% (13/48).All of the ST1/RT027 strains presented a toxin gene profile of tcdA(+)/tcdB(+) and cdtA(+)/cdtB(+).Most of the ST1/RT027 strains were isolated from the Traditional Chinese Medicine Department of Respiratory, where smallnosocomial outbreaks of ST1/RT027 strain infection might happen.Conclusion CDI diagnosed in CJFH mainly belongs to nosocomial infection.Most of the isolated strains harbor tcdA(+)/tcdB(+) genes.Surveillance for the outbreaks of CDI caused by ST1/RT027 strains over producing toxins A and B should be strengthened in hospitals.

Objective To investigate the feasibility and effects of endoscopic surgical treatment of lumbar intervertebral disc herniation associated with veitebral osteochondrosis.Methods From June 2008 to December 2015,276 cases of lumbar intervertebral disc herniation associated with vertebral osteochondrosis were treated with endoscopic surgery,including 185 men and 91 women,with an average 39.2 years old (range,16-65 years old).The involved level included L2.3 in 2 cases,L3.4 in 9 cases,L4,5 in 126 cases and L5S1 in 139 cases.On preoperative axial CT,the diameter of ossification was more than half of the transverse or sagittal diameter of the spinal canal in 89 cases,and no more than half of the transverse and sagittal diameter of the spinal canal in 187 cases.All patients were operated on the side with serious symptom,181 cases were operated with mobile microendoscopic discectomy (MMED),and 95 cases were operated with percutaneous endoscopic surgery,including percutaneous transforaminal endoscopic discectomy (PTED) in 61 cases and the percutaneous interlaminar endoscopic discectomy (PIED) in 34 cases.The operation and complications were analyzed.Results The soft herniation,broken disc material and the periphery of compressing ossification were removed under the endoscope in all cases,until the nerve was well decompressed.However,the ossification was not complete resected.Dural sac tear occurred in 3 cases of MMED.In the early stage of PTED,2 cases converted to MMED because of intraoperative pain and difficulty,and one case had exiting nerve root injury.At the final follow-up of 12-60 months (average,20.6 months),visual analogue scale decreased from preoperative 8.5±1.2 to 1.0±0.9,Oswestry disability index decreased from preoperative 40.2±8.6 to 3.1±3.0.According to Macnab scale,the results were excellent in 89,good in 154 cases,moderate in 33 cases.Conclusion For most lumbar intervertebral disc herniation associated with vertebral osteochondrosis,good results can be achieve by removal of herniated and broken intervertebral disc and decompression of nerve with endoscope.Therefore,we speculate that the soft disc herniation and spinal stenosis are main pathogenic factors,and that the complete resection of ossification is not needed.

Objective To establish an animal model of annulus fibrosus (AF) partial defect for the repairing of interver?tebral disc (IVD) defect. Methods Image J 1.46r software was used to measure the T12/L1-L6/S1 intervertebral height in ovine lumbar spine X-ray films. AF thickness was measured by axial split disc. A 11 blade was used to make a trapezoid de?fect of upper bottom 3 mm, lower bottom 5 mm, height 5 mm and thickness 3 mm, whose lower bottom toward the nucleus pulposus (NP) in the left front of ovine lumbar IVD in vitro. The minimally invasive lateral approach was used to make the same type of trapezoid defect in the left front of the ovine lumbar IVD in vivo. The trapezoidal defect length of the axial divid?ing disc was measured, AF and a small amount of NP from trapezoidal defect in IVD were weighed, and the production of trapezoidal defect in IVD was evaluated. Results The lumbar intervertebral space height of ovine was (4.45 ± 0.28) mm. There were significant differences in the thickness of AF (4.08±0.50) mm , thickness (3 mm) and height (5 mm) of trapezoidal defect (P<0.05), respectively. There were no significant differences in trapezoidal defects in ovine lumbar IVD in vitro on the upper bottom (3.03 ± 0.09)mm, the lower bottom (5.03 ± 0.09) mm, the height (4.97 ± 0.10) mm, the thickness(3.02 ± 0.06) mm and the trapezoidal defect predetermined value on the upper bottom 3 mm, the lower bottom 5 mm, the height 5 mm and the thickness 3 mm (P>0. 05). The weights of the AF and NP taken out from ovine lumbar IVD in vitro and in vivo were (0.162 ± 0.011) g and (0.166 ± 0.014) g, and there was no significant difference between them (P > 0.05). Conclusion Through the operation of minimally invasive lateral approach, the method of making a trapezoidal defect in the experiments can establish animal model of AF partial defect, which meets the requirements for the repairing of IVD defect, and is simple, safe and reliable.

Objective To assess the prospect of integrated biphasic silk fibroin scaffold made by annulus fibrosus-nu?cleus pulposus tissue engineering in application as integrated intervertebral disc(IVD). Methods An integrated annulus fi brosus-nucleus pulposus(AF-NP)biphasic scaffold was made by silk fi broin using two different uncomplicated methods which were paraffin spheres-leaching method(outer AF phase)and phase separation method(inner NP phase). The scaf?fold was investigated by general observation, stereomicroscope and scanning electron microscopy(SEM). Its pore size, poros?ity, and compressive elastic modulus were determined. AF and NP cells were isolated from rabbit IVD and seeded into the corresponding phase of the scaffold respectively. The cell-scaffold complex was cultured for 48 hours. The biocompatibility of the scaffold was evaluated by SEM, live/dead staining while CCK-8 assay was used to assess cell proliferation. Results Stereomicroscope and SEM showed that AF phase and NP phase integrated perfectly without cross-linking. Both phases pos?sessed highly interconnected porous structure [pore size of AF and NP phase were(220.0±23.1)μm and(90.0±17.8)μm, re?spectively] and highly porosity(AF and NP phase were respectively 91%and 93%). In addition, this silk biphasic scaffold had impressive mechanical properties(150.7 ± 6.8)kPa. SEM revealed that disc cells attached to regions of pore walls, dis?tributed uniformly and secreted extracellular matrix. Live/Dead staining and cell count kit-8(CCK-8)analysis showed that the silk composite scaffold was non-cytotoxic to disc cells. Conclusion This silk biphasic AF-NP scaffold has satisfied pore size, porosity, biomechanical properties and biocompatibility, so it is ideal candidate for IVD tissue engineering.

Low back pain (LBP) is a common clinical condition, and non-specific LBP is believed to be associated with lumbar instability caused by paraspinal muscle (PSM) degeneration. Therefore, MRI evaluation of PSM degeneration can help predict the progression of low back pain-related diseases. At present, the most commonly used MRI parameters are the cross-sectional area (CSA) and fat infiltration (FI) of PSM. In order to assess the degree of PSM degeneration more accurately, modified parameters including functional CSA, relative CSA and muscle-fat infiltration index (MFI), have been designed. Advancements in MRI technology,such as the application of IDEAL-IQ, allow for early detection of changes in fat infiltration. Additionally, the proton density fat fraction (PDFF) derived from chemical shift encoding based water-fat MRI (CSE-MRI), also shows promise in studying PSM. Numerous MRI studies, both domestically and internationally, have aimed to establish the relationship between PSM degeneration and non-specific LBP, lumbar degenerative diseases, degenerative lumbar scoliosis, and osteoporosis. It has been found that PSM degeneration is affected by other factors such as intervertebral disc, articular process and endplate degeneration. Furthermore, PSM degeneration and scoliosis/kyphosis are also reciprocal causation. Higher fat content in the vertebral body and PSM is associated with increased bone fragility, while vertebroplasty following osteoporotic vertebral fracture can delay PSM degeneration and improve quality of life.

Objective The biomimetic osteochondral scaffold contained calcified cartilage layer(CCL) was fabricated using slik fibroin (SF) and hydroxyapatite (HA) for materials.To investigate effects of biomimetic osteochondral scaffold contained CCL compounding with ADSCs on regeneration of the osteochondral defect on the rabbit knee,explore the feasibility of this design as a concept of osteochondral tissue engineering.Methods We fabricated a novel biomimetic osteochondral scaffold with CCL using SF and HA by the combination of paraffin-sphere leaching and modified temperature gradient-guided thermal-induced phase separation (TIPS) technique.The pore size,porosity,and compressive modulus of elasticity of the scaffold cartilage layer and the osteogenic layer were measured by scanning electron microscopy and microscopy CT.The osteochondral defect model on rabbit bilateral knees were established,and implanted with the non-CCL group (non-CCL scaffold compounding with ADSCs) and CCL group (CCL scaffold compounding with ADSCs).At 4,8 and 12 weeks after implantation,the rabbits were euthanized,respectively.Gross observation score,histological and immunohistochemical assessment,biochemical quantitative of new osteochondral tissue,micro-CT scans for new bone,were executed.We evaluated the regeneration of osteochondral defects in each group,and verified the role of CCL in vivo.Results The biomimetic osteochondral scaffold with CCL had a consecutively overlapping trilayer structure with different densities and pore structures,including a chondral layer (top layer),intermediate layer and bony layer (bottom layer).The cartilage layer had a well-oriented microporous structure with a uniform distribution with a pore size of (112.43± 12.65)μm and a porosity of 90.25％±2.05％.The subchondral bone layer had a good three-dimensional macroporous structure,good connectivity,pore size (362.23±26.52) μm,porosity of osteogenic layer was 85.30％± 1.80％.The cartilage regeneration in CCL+AD-SCs group was better than non-CCL+ADSCs group.The content of GAG and type Ⅱ collagen in new cartilage tissue in CCL+AD-SCs group was much more than non-CCL+ADSCs group.The new bone tissue analysis and biomechanical testing had no significant differences between the two groups.Conclusion The biomimetic osteochondral SF/HA scaffold contained CCL mimics the structure of normal osteochondral tissue with good 3-dimensional pore structure and biocompatibility.The scaffold complex autologous ADSCs successfully repair osteochondral defects in rabbit knee,and the presence of CCL accelerates the growth of cartilage.

Objective To evaluate the strategy and clinical effects of percutaneous endoscopic surgery for cervical disc herniation.Methods Fifty-one patients with cervical disc herniation were treated with percutaneous endoscopic surgery from June 2015 to March 2017,including 32 men and 19 women,with an average age of 52.2 years (range,28-66 years).Radicular symptoms were present in all patients,while 23 patients had mild myelopathy (Nurick Grade:0-3) and 3 patients of multilevel stenosis had severe myelopathy (Nurick Grade:4-5).According to axial image of preoperative magnetic resonance imaging (MRI),31 patients had lateral herniation that was located lateral to the edge of spinal cord,20 patients had central herniation that was located within the lateral edge of spinal cord.Among them,48 patients had soft herniation and 3 patients had ossified lateral herniation combined with foraminal stenosis.All surgery was carried out under general anesthesia,while posterior and anterior percutaneous endoscopic surgeries were performed for lateral herniation and central herniation respectively.Posterior endoscopic surgery was performed with "keyhole" fenestration at "V" point (the junction of lateral edge of lamina space and inner edge of facet).Lateral edge of thecal sac and nerve root were exposed and decompressed,soft herniation was explored and removed.Anterior endoscopic surgery was performed through puncture and 4mm tube between the visceral sheath and vascular sheath.The tube was inserted through disc to the base of herniation under fluoroscopy.The herniation was removed until the dura sac was exposed and relaxed.One stage open-door laminoplasty was performed for 3 patients with severe multiple segmental stenosis and huge central herniation.The operative time and blood loss were recorded,and patients were followed-up (range,6-18 months,average 12.1 months) to evaluate the clinical efficacy.Results The mean operative time of posterior endoscopic surgery was 90 min (range,45-150 min).The nerve root was not well exposed,and the fenestration was too lateral in 1 patient,with partial relieve of symptoms;and simple nerve root decompression was performed for 3 patients of ossified herniation combined with foraminal stenosis.Herniated or sequestered nucleus pulposus was removed for 27 patients,one of them had transient paralysis ipsilateral limb and 2 of them had linkage of cerebrospinal fluid.The Visual Analogue Score (VAS) score improved form preoperative 8.9±1.6 to 0.5±0.4,and the Oswestry Disability Index (ODI) score improved form 32.8±4.2 to 2.3± 1.9 at final follow-up.For anterior percutaneous endoscopic surgery,the mean operative time was 80 min (range,45-120 min).Herniated or free nucleus was successfully removed for all patients.The thecal sac was lacerated due to unclear exposure in 1 case.The VAS score improved form preoperative 6.9±2.3 to 0.9±0.8,and the ODI score improved form 40.1±8.6 to 5.6±3.0 at final follow-up,with improvement of myelopathy at least one Nurick Grade.During follow-up,the alignment of cervical spine was well preserved without kyphosis for two groups,while the height of intervertebral space decreased with 0.4±0.3 mm and 0.9±0.6 mm in posterior and anterior surgery respectively.Conclusion Percutaneous endoscopic surgery provides minimally invasive alternatives for some cervical disc herniation with predominant radicular pain.Posterior endoscopic surgery is suitable for lateral herniation,and anterior endoscopic discectomy is suitable for some central soft herniation without obvious collapse and instability.However,the long-term results of disc space collapsed after anterior approach remains unclear.

Objective:To evaluate the value and efficacy of microscope-assisted minimally invasive anterior lumbar discectomy and zero-profile fusion (ALDF) for lumbar degenerative diseases.Methods:Anterior lumbar distractors were designed to maintain the distraction of intervertebral space and expose the posterior edge of the intervertebral space. From June 2018 to December 2020, 41 cases of lumbar degenerative diseases were treated with this operation, including 19 men and 22 women, aged 29-71 years old (average 42.1 years old). All patients had intractable low back pain. Imaging examination showed lumbar disc degeneration with narrow intervertebral space, including disc herniation with Modic changes in 7 cases, spinal stenosis with instability in 16 cases and spondylolisthesis in 18 cases. The involved levels included L 2,3 in 1 case, L 3,4 in 3 cases, L 2-L 4 in 1 case, L 4,5 in 17 cases and L 5S 1 in 19 cases. An incision was taken that was pararectus for L 2-L 4 and transverse for L 4-S 1, with the intervertebral disc exposed via extraperitoneal approach. The intervertebral space was released and distracted after discectomy in intervertebral space, and self-made distractors were used to maintain the space. Under microscope, the herniation, posterior annulus and osteophyte were removed for sufficient decompression, with a suitable self-anchoring cage implanted into the intervertebral space. The visual analogue score (VAS), Oswestry dysfunction index (ODI), intervertebral space height, lordosis angle and spondylolisthesis rate were evaluated. Results:Operations were performed successfully in all the patients. The operation time was 70-120 min with an average of 90 min, and the intraoperative blood loss was 15-70 ml with an average of 30 ml. No severe complication such as nerve or blood vessel injury occurred. The patients were followed up for 12 to 36 months, with an average of 18 months. At the last follow-up, VAS decreased from 6.4±2.3 to 1.1±0.9, and ODI decreased from 44.9%±16.9% to 5.8%±4.7%. Intervertebral space height recovered from 7.2±2.8 mm to 12.1±2.1 mm and lordosis angle recovered from 6.9°±4.8° to 10.1°±4.6°. X-ray showed significant recovery of intervertebral space height, lordosis angle and spondylolisthesis rate, with obvious interbody fusion and no displacement of cage. For 18 patients of spondylolisthesis, the slippage recovered from 16.6%±9.3% to 7.6%±5.3%, with an average improvement of 54.2%.Conclusion:Microscope-assisted minimally invasive ALDF can provide sufficient decompression and zero-profile fusion for lumbar degenerative diseases with satisfactory results during short-term follow-up.

Objective:To evaluate the safety and validity of enriched autologous bone marrow mesenchymal stem cells (BMSCs) and annular suture for repairing defect after lumbar discectomy.Methods:Enrichment of autologous BMSCs: autologous bone marrow blood was collected from 5 patients undergoing lumbar surgery, and nucleated cells were enriched on gelatin sponge particles by selective cell retention technique. From October 2016 to March 2019, 109 patients with lumbar disc herniation underwent discectomy with mobile microendoscopic discectomy technique, including 61 males and 48 females, aged 24-59 years. Discectomy group: 26 cases received simple discectomy. Suture group: 39 cases received annular suture after discectomy. BMSCs+suture group: 44 cases received intradisc transplantation of gelatin sponge particles enriched with autologous BMSCs and annular suture after discectomy. The perioperative conditions were recorded, with visual analogue scale (VAS), Oswestry dysfunction index (ODI), Pfirrmann grade of disc degeneration, disc height and degree of herniationevaluated after operation.Results:In enrichment test with flow cytometry, the enrichment multiple of nucleated cells and target cells was 6.4±0.9 and 4.2±0.6 respectively, and BMSCs grew well in vitro. The operation time was 35-55 mins. 7 cases in the suture group were transferred to the discectomy group and 10 cases in the BMSCs+suture group were transferred to BMSCs group due to unsuccessful suture. There were no significant differences in VAS, ODI, Pfirrmann grade of disc degeneration, disc height and degree of herniation among the groups. There was no significant difference in intraoperative bleeding, postoperative drainage and length of hospital stay. The incision was healed without redness and swelling. 18 patients were followed up for 6 months, and 91 cases were followed up for 1-3 years (25.0±5.6 months). There was no interbody fusion, heterotopic ossification or infection during follow-up. VAS and ODI decreased significantly after operation in all patients. At final follow-up, the VAS improvement rate of BMSCs+suture group (81.7%±7.9%) was higher than discectomy group (73.0%±8.9%), suture group (74.0%±6.9%) and BMSCs group (75.3%±8.4%); the ODI improvement rate of BMSCs+suture group (91.9%±8.8%) was higher than discectomy group (86.2%±8.1%) and suture group (86.4%±5.5%). According to MRI, the Pfirrmann grade of disc increased 0.7 in discectomy group, 0.6 in suture group, while it did not increased significantly in BMSCs+suture group and BMSCs group, and the progress of Pfirrmann grade in BMSCs+suture group and BMSCs group were lighter than discectomy group and suture group.The disc height decreased in each group, the loss rate of disc height in BMSCs+suture group (17.2%±4.3%) was less than discectomy group (29.3%± 6.3%) and suture group (20.6%±5.7%); and suture group was less than discectomy group. The degree of herniation was reduced by more than 50% in all groups, while 1 case in discectomy group had herniation without clinical symptoms.Conclusion:Autologous BMSCs and annulus suture are safe and effective in repairing the defect after lumbar discectomy, which may help to slow down the degeneration of intervertebral disc.

Objective To explore the gene expression profile in paraspinal muscle degeneration(PMD)and identify key ferroptosis genes.Methods RNA sequencing was performed on paraspinal muscle tissue of 3 normal and 3 PMD patients respectively to obtain differentially expressed genes.Through protein-protein interaction(PPI)and gene functional enrichment analysis,the intersection of ferroptosis genes was identified to identify key hub genes associated with ferroptosis.The diagnostic value for PMD disease was analyzed by receiver operating characteristic(ROC)curves.Results A total of 292 differentially expressed genes were identified in PMD.Among them,125 genes were significantly downregulated and 167 genes were significantly upregulated.Bioinformatics analysis revealed that 14 differentially expressed genes were associated with ferroptosis.Among them,ferroptosis genes MUC1,ATF3 and CDKN1A were key hub genes with good specificity and sensitivity for diagnosing PMD.Functional enrichment analysis revealed that they may mediate the occurrence and progression of PMD by regulating cell apoptosis,ferroptosis and skeletal muscle tissue development and differentiation.Conclusion Ferroptosis genes MUC1,ATF3 and CDKN1A can serve as biomarkers for diagnosing PMD,providing theoretical basis for decoding the pathological mechanism of PMD and developing new drugs.