Objective To make a research on the expression of perlecan in the formation of epidural scar after posterior lumbar laminectomy in the rat.Methods Sixty Wistar rats were randomly and evenly divided into five groups:the experimental groups including A(the first week after surgery),B(the second week),C(the fourth week),D(the sixth week)and the control group.Laminectomy was performed on lumbar vertebra in each experimental group.The hematoxylin-eosin(HE)staining was used to observe the formation of epidural scar.Immunohistochemistry was performed to detect the perlecan located in the epidural scar.Western blot and RT-PCR were made to quantify the expression of the perlecan after surgery.Results According to HE,with prolonged,the scar tissue gradually formed.Immunohistochemistry demonstrated that the perlecan was expressed in the fibroblast.Western blot and RT-PCR showed the high level expression of perlecan in the experimental groups compared to the control group(P＜0.01).Furthermore,the expression of perlecan gradually increased with the prolongation of exposure time and had significantly statistical significance (P＜0.01)between the first three experimental groups.However,compared to the fourth week group,the expression of perlecan in the sixth week decreased obviously(P＜0.01).Conclusion Perlecan was expressed in the fibroblast of the epidural scar,and which plays an important role in the formation of epidural scar.

Objective To investigate the impact of stable expression of perlecan shRNA lentiviral particles on proliferation of NIH3T3 cells. Methods Mouse fibroblasts were cultured.Lentiviral particles-green fluorescent protein (LV-GFP) was used to transfect the cultured NIH3T3 cells with multiplicity of infection (MOI) of 10,30 and 50.The GFP expression was observed with fluorescence microscopy after transfection for one week to estimate the proper MOI and the time of GFP expression needed.The transfection efficiency of LV-GFP with the proper MOI by fluorescence-activated cell sorting was detected.The stably transfected cell lines were developed by puromycin screening for more than 2 weeks.The third generation HFF in good condition was randomly divided into 3 groups:GFP group,shRNA group and control group.RT-PCR,Western blot and MTT assays were used to detect the expressions of perlecan mRNA and protein and cell proliferation in the 3 groups. Results Perlecan mRNA and protein showed high expressions in the control and GFP groups but low expressions in the shRNA group,with significant differences respectively between the shRNA group and the other 2 groups ( P ＜ 0.05).There was no significant difference between the 3 groups in the optical density at the first 2 days ( P ＞ 0.05).On 3 to 6 days the cells in the control and GFP groups grew normally while the cells in the shRNA group proliferated in a weak manner.the transfected cells in the shRNA group showed a significantly reduced proliferation rate compared with the other 2 groups ( P ＜ 0.05 ). Conclusion The growth of NIH3T3 cells can be inhibited significantly by transfection with perlecan shRNA lentiviral particles.

Objective To evaluate the assistant medial approach used in the surgical management of intraarticular fractures of calcaneus.Methods From January 2005 to December 2009,33 patients with intraarticular fracture of calcaneus were treated by open reduction and internal fixation through the expanded lateral L-shape approach and the assisted medial approach.They were 27 men and 6 women,aged from 18 to 62 years (average,36 years).There were 15 left sides and 18 right sides.According to Sanders' classification system,there were 27 patients with type Ⅲ displaced calcaneal fracture and 6 with type IV.Four cases were complicated with fracture of lumbar vertebra,one with pilon fracture and one with fracture of acetahulun.Fracture healing was observed and the Bǒhler and Gissane angles were measured.The American Orthopaedic Foot and Ankle Society (AOFAS) ankle-hindfoot scale system was used to evaluate the functional recovery.Results All patients were followed up postoperatively to assess their functional outcome.The follow-up duration was 6 to 24 months (mean,12 months).All incisions healed primarily and all fractures obtained bony union in an average of 8 weeks (from 6 to 24 months).All the Bǒhler and Gissane angles were corrected and AOFAS scores were improved.There were significant differences between preoperation and pestoperation in all indexes( P ＜ 0.05).Conclusion As the medial approach can be used to assist the expanded lateral L-shape approach in treatment of the intraarticular fractures of the calcaneus,the reduction and insertion of internal implants can be improved to achieve satisfactory functional outcome.

BACKGROUND:In recent years,studies have shown the effects of tumor necrosis factor-a(TNF-а)on fibroblasts at different organizations in a tissue-specific and dose-dependent manner.OBJECTIVE:TO investigate the biologic effects of TNF-а and its signal transduction pathway-specific kinase inhibitors on mouse embryonic fibroblasts(NIH3T3).METHODS:NIH3T3 cells were cultured in vitro,and then the cells were assigned into 3 groups.Cells in the control group were cultured in DMEM high-glucose medium with 2%serum;those in the TNF-а group were cultured in 100 μg/L TNF-аmedium;those in the TNF-а+Anti-TNFRSF 1 B group were firstly cultured in medium with 50 μg/L Anti-TNFRSF 1 B for 1 hour,and then placed in the medium with 100 μg/L TNF-а.RT-PCR and Western blot methods were used to evaluate mRNA and protein expressions of type Ⅰ collagen and matrix metalloproteinase 3(MMP3)in each group.RESULTS AND CONCLUSION:In this experiment,NlH3T3 cells cultured with a certain concentration of TNF-а,the specificity kinase signal of transduction pathway presented with phosphorylation or protein activation,and the signal pathway was activated,which promoted MMP3 activation,and significantly reduced the expression of type Ⅰ collagen.The effect of TNF-а was certainly inhibited,but not completely eliminated after adding its signal transduction pathway inhibitor Anti-TNFRSF1 B.This further proves the role of TNF-а on NIH3T3 activation.

Objective To investigate the effects of Leptin on the base of anthropometry,densitometry,and biochemistry and reveal the role of Leptin in bone metabolism in postmenopausal women with osteoporosis (OP).Methods From January 2008 to January 2014,data of 82 postmenopausal females with or without OP were analyzed.They were randomized into two groups:OP (n=40) and conditional control group (CON) (n=42).The expression of Leptin receptors in osteoblasts was observed by immunohistochemistry.1 × 10 ng/ml,1 × 102 ng/ml,1 × 103 ng/ml,1× 104 ng/ml of Leptin were added respectively.Then the proliferation,differentiation and mineralization of osteoblasts were measure at 24 h,48 h and 96 h.The gene expressions of bone marrow mesenchymal stem cells (BMSCs) derived osteoblasts and adipocytes treated with Leptin were examined by quantitative analysis of Real time-PCR.Results The expression of Leptin receptors in osteoblasts was observed on plasma membranes and cytoplasm in the normal group and osteoporosis group.In osteoporosis group,different concentrations of Leptin enhanced cell proliferation,cell differentiation and cell mineralization.The growth rate of MTT and the concentration of ALP in the serum of were significantly increased with the effects of 1×10 ng/ml,1×102 ng/ml and 1×103 ng/ml of Leptin,and the action of 1×102 ng/ml Leptin was the most powerful with time dependence.Within 96 hours,the growth rate of osteoblasts increased gradually,and the concentration of ALP increased gradually in 3 weeks.The expression of RANKL/OPG in bone marrow BMSCs derived osteoblasts treated with Leptin were significantly increased.Conclusion Leptin receptor is present in osteoblasts,and Leptin affects biological behaviors of osteoblasts through receptors.The direct effect of Leptin may relate the expression of RANKL/OPG.Then,the balance between bone resorption and bone formation was broken and finally osteoporosis occurred.Lepin may promote osteogenesis and inhibit bone resorption in the differentiation of BMSCs.

[Objective]To investigate the role and mechanism of interleukin-1? on fibroblast synthesis of matrix metalloproteinase-3,and explore the impact of interleukin-1? on the formation of epidural scar.[Method]The NIH3T3 cells were divided into three groups:IL-1? group,IL-1? + SB202190 group and control group. After serum-free culture for 20 hours,IL-1? group was treated by IL-1? (10ng/ml) and IL-1? + SB202190 group was treated by the SB202190 (10?mol/L) for 1 hour before treatment with IL-1?(10ng/ml) and the control group was treated with 2% serum. Then those 3 groups were cultured for 24 hours under standard conditions (37℃ in a humidified atmosphere flushed with 5% CO2 in air). The expression of MMP-3 gene was examined by RT-PCR and Western blotting.[Result]The expression of MMP-3 significantly increased in the IL-1? group than that in the control group(P0.05). [Conclusion]IL-1? could stimulate the expression of MMP-3 in NIH3T3 cells which might inhibit the formation of epidural scar. Besides,the author found that the p38 pathway played an major role in this process.

Objective To study the effect on prevention of dura mater spinalis adhesion after laminectomy through an experiment by using keratin substance (KS) artificial fasciae.MethodSixty rats were randomly divided into three groups, each had 20. In all rats, total laminectomy were performed at T12 levels led to the defects of 2 mm?8 mm. The group A were sutured directly, which was regarded as control group; covered with artificial fasciae in group B; with self-faciae in group C. The animals were killed at the 2nd, 4th, 8th, 12th week after operation respectively, and macroscopy assessment was done with the help of the Rydell Criterion. The spine segment of T12 was removed en bloc including the paraspinal tissue, and all specimens were stained with hematoxylin and eosin. Improved Nussbaum Criterion was used for the following optical microscopy assessment. Transmission electron microscopy was performed on posterior epidural scar obtained from group-B at the 2nd, 4th, 8th, 12th week after operation respectively. The nonparametric Kruskal-Wallis test was used to determine the differences between the three groups, and a P value of 0.05).So compared to the control group, KS artificial fasciae and self-fasciae significantly reduced the formation of epidural fibrosis after laminectomy.ConclusionKS artificial fasciae have the best biological compatibility with the tissues with no adverse effects. The posterior hematoma and fibroblast (FB) are the main source of the epidural fibrosis.As placement of KS artificial fasciae over the laminectomy defect in rats could provide a physical barrier against invasion of fibrous tissue into the vertebral canal,and peridural fibrosis after laminectomy would be effectively prevented. KS artificial fasciae is safe and effective in limiting the direct contact by the postlaminectomy epidural fibrosis in rats.

BACKGROUND: Artificial nerve grafts of human hair keratin are a kind of biological products. It has low antigenicity,absorbability and stimulation to nerve fiber growth following specific biochemistry. It is hoped to have better effect than otherartificial nerve grafts.　 OBJECTIVE: To investigate the anatomy and histocompatibility of artificial nerve grafts of the human-hair keratin, and toobserve its effects on the repair of peripheral nerves. 　 DESIGN, TIME AND SETTING: Randomized, controlled, animal experiment. The study was performed at the Animal CentralLaboratory of Affiliated Hospital of Qingdao University Medical College between November 2006 and June 2008. MATERIALS: Artificial nerve grafts of human hair keratin is a compound of human hair processed by specific controlledbiochemistry based on ground substance, embedded with a layer of biological membrane. It has low antigenicity, absorbabilityand stimulation to nerve fiber growth following specific biochemistry. METHODS: Eighteen Wistar rats were randomly divided into three groups. The sciatic nerve, 10 mm, was removed andtransplanted with human-hair keratin graft, skeletal muscle and untreated hair, respectively.MAIN OUTCOME MEASURES: The characteristics of histomorphology and anatomy were observed at 8, 12, 24 weeks afterthe surgery. RESULTS: White tissues appeared between the broken ends of the sciatic nerve at 8 post-operative week in the graft group,and appeared in the graft space in human-hair keratin at the 12th week. At the 24th week, a large amount of infantile myelinatednerve fibers were observed under optical microscope regenerating around the human hair, which was partially degraded andabsorbed. Schwann cells were observed under an electron microscope and myelinization. 　 CONCLUSION: The artificial nerve grafts of the human-hair keratin are well compatible with the body tissues, and couldinduce nerve regeneration.

Objective To study the prevalence of cholelithiasis among spinal cord injured (SCI)male patients, and the correlation of cholelithiasis with the SCI patients′ age, weight,and the level of injury, severity and duration of the SCI. Methods One hundred male SCI patients including 58 patients with ASIA class A and B and 42 with ASIA class C and D,with aged 20-65 years old(average 46.5 years) were followed up.They suffered from a spinal cord injury more than one year. The control group consisted of 100 male volunteers without both SCI and biliary diseases, with aged 20-68 year old(average 42.6 years). All patients in the two groups underwent ultrasonographic imaging to evaluate the gallbladder and the biliary tract. Results The prevalence of cholelithiasis was 26.0% in the SCI group patients and 10.0% in the control group,the difference was statistical difference (P 0.05). Conclusions SCI represents a major risk factor for the development of cholelithiasis . There are no correlation of cholelithiasis with the SCI patients′ age , weight,SCI level,severity and duration of SCI.

BACKGROUND:Cervical Open-door laminoplasty with unilateral mass screw fixation and laminectomy with bilateral mass screw fixation are both procedures that treat cervical spondylotic myelopathy by expanding the space available for the spinal cord. Effectiveness and safety of two operative methods remain unclear. OBJECTIVE:To investigate the biocompatibility of implant and host between open-door laminoplasty with unilateral mass screw fixation and laminectomy with bilateral mass screw fixation to treat multi-segment cervical spondylotic myelopathy. METHODS: Data of 117 patients with multi-segment cervical spondylotic myelopathy (≥ 3 segments) were retrospectively analyzed. Sixty-five cases underwent open-door laminoplasty with unilateral mass screw fixation (laminoplasty group). Fifty-two cases underwent laminectomy with bilateral mass screw fixation (laminectomy group). Clinical outcomes were evaluated by the Japanese Orthopaedic Association scoring system and by recovery rate. Cervical curvature index and cervical range of motion were assessed by X-ray films in both groups. RESULTS AND CONCLUSION:The average folow-up time was 28 months (range 12-59 months) in both groups. No C5 nerve root palsy occurred in the two groups. Japanese Orthopaedic Association scores were greater during final folow-up than pre-fixation in both groups (P < 0.01). No significant difference in Japanese Orthopaedic Association score, recovery rate, and final folow-up cervical curvature index was detected in both groups (P > 0.05). Cervical range of motion was lower during final folow-up than pre-fixation in both groups (P < 0.01). Results confirmed that open-door laminoplasty with unilateral mass screw fixation and laminectomy with bilateral mass screw fixation have similar effectiveness in the improvement of neurological function, relieving pain and reducing complications, but wil greatly reduce the cervical range of motion.