BACKGROUND: At present the strategy of nerve regeneration and repairng are main promoting nerve intrinsic regeneration capacity and improving the micro-environment. Studies have shown a number of combined treatment which could promote the regeneration and growth of nerve axon.OBJECTIVE: To explore the feasibility and effect of rat spinal cord injury repaired by peripheral nerve combined growth factor. METHODS: Sixty healthy adult female SD rats were randomly divided into 4 groups: nerve graft group, nerve graft combined growth factor group, spinal cord transaction group and laminectomy group. Taking T_9 as the center, a longitudinal incision was conducted in rat skin, revealing dural sac, spinal cord was transected and removed 3 mm, 2-cm segment of the eighth to tenth intercostal nerve was obtained from nerve graft group and nerve graft combined with growth factor group, autologous intercostal nerve was cross-transplanted into spinal defect (proximal white matter and distal gray matter, distal white matter and proximal gray matter) after pruning appropriately. The transplanted intercostal nerves were fixed with fibrin glue in nerve graft group, while those in nerve graft combined growth factor group were fixed with fibrin glue containing 2.1 mg/L acidic fibroblast growth factor, followed by dural suture～ Stump of broken ends was done in spinal cord transection group, while laminectomy was performed in laminectomy group. RESULTS AND CONCLUSION: At 90 days post-surgery, somatosensory evoked potential (SEP) and motor evoked potential (MEP) were determined, the motor function of hind limbs was evaluated by the Basso. Beattie.Bresnahan (BBB) test at 70 days. Both SEP and MEP were led in the laminectomy group, but not lead in spinal cord transection group; in nerve graft group, 3 rats showed bilateral SEP, 4 led unilateral SEP, 4 led bilateral MEP, 3 led unilateral MEP; in nerve graft combined with growth factor group, 5 led bilateral SEP and 2 led unilateral SEP, 5 led bilateral MEP and 2 led unilateral MEP. The SEP and MEP latency and amplitude in the nerve graft group and nerve graft combined growth factor group were significantly superior to the spinal cord transection group (P < 0.01), autologous rib nerve graft group was better than nerve graft combined growth factor group (P <0.01). In the laminectomy group, awake rats following anesthesia returned to normal exercise, rats in spinal cord transection group continued to extend limbs and rotated within 3 months, rats in other two groups recovered functions obviously 3 weeks post-surgery and gradually restored throughout the entire observation period. Nerve graft group and nerve graft combined growth factor group showed significantly increased BBB score compared with spinal cord transection were (P < 0.01), and the nerve graft combined growth factor group was superior to nerve graft group (P < 0.01). The peripheral nerve graft can promote the spinal function following spinal cord injury, while the nerve combined growth factor can better restore the function.

BACKGROUND: Transforming growth factor (TGF)-β_1, a potent cell growth and proliferation regulatory proteins, plays an important role in development of anti-graft rejection and graft vascular disease. OBJECTIVE: To observe local injection of TGF-β_1 effects on transplant immune rejection following freezing disposal and nerve allograft. DESIGN, TIME AND SETTING: The randomized controlled animal study was performed at the Animal Experimental Center, Harbin Medical University from June 2007 to June 2008. MATERIALS: A total of 60 clean SD rats (recipients) were divided into 3 groups: autogenous nerve graft group, nerve allograft group, TGF-β_1 plasmid + nerve allograft group, 20 in each group. A total of 40 Wistar male rats served as donors. pAdTrack-CMV-TGF-β_1 plasmid, pAdEasy-1-Bj51833 cells were presented by the Orthopedic Laboratory of Fourth Hospital of Harbin Medical University. METHODS: Longitudinal posterolateral incision was made in 40 donor rats to expose sciatic nerve. The whole bilateral sciatic nerve was cut and placed in sterile frozen tubes for 1 week for use. Under the microscope, connective tissue was cut in the biceps muscle and semi-tendon and semi-membrane gap of recipient rats to expose the sciatic nerve. 1-cm sciatic nerve was cut 0.5 cm below the muscle from the plow-shaped hole. Transplantation of frozen autogenous nerve graft and nerve allograft (nerve at equal size) was separately performed in the autogenous nerve graft and nerve allograft groups. In the TGF-β_1 plasmid + nerve allograft group, pAdTrack-CMV-TGF-β_1 plasmid (40 μg) was injected into the local muscle and two sides of transected sciatic nerve of each rat following nerve allograft transplantation. MAIN OUTCOME MEASURES: Motor nerve conduction velocity, pathology and axonal counting were examined 3, 6, 9 weeks after surgery. RESULTS: Motor nerve conduction velocity was higher in the TGF-β_1 plasmid + nerve allograft group than in the nerve allograft group (P < 0.01), which did not show significant difference compared with the autogenous nerve graft group. Axonal counting was greater in the autogenous nerve graft and TGF-β_1 plasmid + nerve allograft groups compared with the nerve allograft group 9 weeks following surgery (P < 0.01). Using optical microscope and electron microscope, nerve fibers were normal and well arranged in the TGF-β_1 plasmid + nerve allograft group. Nerve fibers presented vascular proliferation, good myelin sheath. Abundant regenerated myelin sheath was found in nerve fiber. The number of Schwann cells was obviously increased, and there were prosperous cytoplasm, a large amount of rough endoplasmic reticulum, clear mitochondria. In regenerated axons, microfilament closely arranged, which was similar to the autogenous nerve graft group. In the nerve allograft group, the optical microscope and electron microscope showed a few nerve fibers, disorderly arranged, significant demyelination, axon degeneration and disappearance, without regenerated fibers. CONCLUSION: Local injection of TGF-β_1 plasmid could reduce immune rejection after cold sciatic nerve allograft transplantation.

BACKGROUND:It is proved by a number of experiments that such a structure as Bungner band-Schwann cell-basilar membrane,which is formed at 2 or 3 weeks after nerve injury,is the ideal microenvironment for neural regeneration. However,the sprouting of nerve fiber close to broken ends takes place at several hours after nerve injury,which shows that the regeneration of nerve fiber and the formation of required microenvironment don't occurred at the same time.OBJECTIVE:To investigate the best repairing time for peripheral nerve injury.DESIGN,TIME AND SETTING:A randomized control animal experiment was performed in the Animal Experiment Centre,Harbin Medical University from June 2007 to June 2008.MATERIALS:A total of 20 New Zealand rabbits were randomly divided into four groups,namely,an immediate repairing group and the other three groups that were repaired respectively at week 2,week 4 and month 3 after injury.METHODS:Peripheral nerve injury models of New Zealand rabbits were established. The immediate repairing group received suture immediately after injury;For the other three groups,the two broken ends of their nerves were fixed on sarcoiemmas temporarily and their wounds were sutured layer by layer. Then they were opened respectively at week 2,week 4 and month 3 after injury,to receive epineural suture with non traumatic 10-0 nylon suture under operating microscope,after which wounds were sutured again.MAIN OUTCOME MEASURES:Nerve electrophysiological observation,axon number,light microscope and electron microscope observation of sutured nerve segments in each group.RESULTS:Nerves repaired at week 2 after injury had a slower nerve conduction velocity than those at week 4 and month 3 after injury (P＜0.01);There was no difference of significance between the immediate repairing group and the group repaired at week 2 after injury (P＞0.05). According to the comparison among the four groups:it had the best repairing effect to repair nerve at week 2 after injury,with normal course and neat arrangement of nerve fibers,vascular proliferation in nerve fibers,myelin sheaths with better structure,Schwann ceils with active function,as well as regenerated axons with intensively arranged microfilaments;Repairing at week 4 after injury had the worst effect,with rare nerve fivers disorderly arranged,myelin sheath and axons significantly degenerated,most nerve fibers demyelinated with axons disappeared,and no regenerated nerve fibers seen;Repairing at month 3 saw the worse repairing effect,with more nerve fiber damaged and disorderly arranged,myelin sheath and axons significantly degenerated,nerve fibers rarely regenerated,less Schwann cells,as well as cytoplasm did not well develope;The effect of immediate repairing after injury was better,with nerve fibers unobviously damaged and well arranged,myelin sheath and axons lightly degenerated,large amounts of myelin sheaths regenerated in nerve fibers,Schwann cells increased obviously,as well as cytoplasms better-developed. Axon counting result was better in the group repaired at week 2 after injury than the otherthree groups,with the minimum in the group repaired at week 4 after injury.CONCLUSION:Repairing at week 2 after injury can get a better result than at any other time points,accordingly two weeks after nerve injury is the best time for repairing peripheral nerve injury.

Objective To investigate the protective effect of rosiglitazone on the rats with high altitude pulmonary edema.Methods Thirty-six SD rats were randomly (random number) divided into 6 groups (n =6 each):control group (Control),hypobaric hypoxia model group (HH),rosiglitazone groups (RSG) which were administered with 3 different doses [RSG-L:5 mg/ (kg · d),RSG-M:10 mg/ (kg·d),RSG-H:20 mg/ (kg· d)],dexamethasone group [Dex,4 mg/ (kg· d)].Rats were injected intraperitoneally with different doses of rosiglitazone (RSG),dexamethasone (Dex) or vehicle (Control and HH) for 3 days before placed in simulated altitude of 6 000 m hypobaric hypoxia animal chamber where the temperature and pressure were constant.After 72 h in the chamber,each rat was anesthetized.The water content of lung was determined with wet/dry weight ratio.Bronchoalveolar lavage fluid was measured by bradford method.The contents of GSH was measured by micro-ezymed labeled method.The contents of MDA was measured by TBA method.The enzymatic activities of SOD was measured by WST-1 method.The changes of the TNF-α,IL-6 and IL-10 in serum were determined by ELISA.Light microscope was used to observe the pathological changes of lung tissue.Results Compared with Control group,the wet/dry weight ratio of lung (5.08 ± 0.24) and total protein content of BALF (351.06 ± 44.55) μg/mL increased significantly (P ＜ 0.01) in HH group.There were red blood cells in the alveolar and interstitium,pink fluid exudation in the alveolar,the alveolar septum enhancement,and a large number of inflammatory cell infiltration;the SOD activity (10.65 ± 0.94) U/mgprot and the content of GSH (1.63 ±0.20) μmol/gprot in lung tissue were significantly decreased (P ＜ 0.01),the contents of MDA (2.1 5 ± 0.18) nmol/mgprot increased significantly (P ＜ 0.01),TNF-o (56.92 ± 2.87) pg/mL and IL-6 (217.80 ±48.01) pg/mL levels in serum were significantly increased (P ＜0.01),and IL-10 (76.85 ± 16.72) pg/mL level decreased (P ＜ 0.05).Compared with the HH group,the wet/dry ratio of lung and total protein content of BALF in different doses of rosiglitazone group significantly decreased (P ＜ 0.01),the pathological changes of the lung tissue was significantly improved,SOD activity and the content of GSH in lung tissue was significantly increased (P ＜ 0.01),the content of MDA decreased (P ＜ 0.01),The levels of TNF-α and IL-6 in serum were significantly decreased (P ＜ 0.01),while the IL-10 level was significantly increased (P ＜ 0.01).Conclusion Rosiglitazone could protect the high altitude pulmonary edema by alleviating the oxidative stress and inflammatory response.

Objective To prepare the anti-TLR4 C-terminal domain monoclonal antibody and investigate its effect in treatment of sepsis.Methods TLR4 C-terminal polypeptide (amino acid sequence:368-579,named as TLR4-C) was obtained through prokaryotic expression and Sephacryl S-100 gel purification,and then was used to immunize female Balb/c mice (6-8 weeks old).After cell fusion,antibody screening and purification,monoclonal antibody specific for the C terminal of TLR4 was obtained.Specificity of monoclonal antibody was detected by Western blot and cell immunofluorescence.In vitro antibody activity test,NR8383 was cultured for 1 h with adding antibody (100 μg/ml) and then 12 h after adding lipopolysaccharide (LPS) (10 ng/ml),and level of tumor growth factor (TNF)-α in the culture medium was tested by ELISA.In vivo septic animal experiment,40 SD rats were assigned to control antibody group (n =20) and anti-TLR4 monoclonal antibody group (n =20) according to the random number table.Each group was rejected 50 mg/kg corresponding antibodies via caudal vein for 1 h,and then LPS (10 mg/kg) via intraperitoneal injection for 4 h.Blood samples from caudal vein of ten rats in each group were collect to test the serum level of TNF-α.The rest rats in each group were used to measure the animal survival rate within 72 h.Results Three highly specific anti-TLR4 monoclonal antibodies were obtained and could combined with TLR4-C and TLR4 holoprotein.In vitro cell activity study indicated only one monoclonal antibody could obviously inhibit the release of TNF-α.In vivo animal experiment showed serum TNF-α level in anti-TLR4-C antibody group was (1.54 ± 0.18) ng/ml,significantly lower than (0.51 ± 0.10) ng/ml in antibody control group (P ＜ 0.01).Animal survival rate in anti-TLR4-C antibody group was 70％,higher than 30％ in antibody control group (P ＜ 0.05).Conclusion Anti-TLR4-C monoclonal antibodies have great capacity to neutralize TLR4 and good protective effect on LPS-induced sepsis.

This article was aimed to study the effect of polyphenols fromRubus suavissirnusS. Lee (RSLP) on spontaneous hypertensive rats (SHR) and to explore its mechanism of anti-hypertensive. The water extraction of RSLP was prepared. And the polyphenols was extracted with macroporous resin. The non-invasive blood pressure analysis system was used to detect the blood pressure. SHR model was selected to study the anti-hypertensive effect. The 16 normal Wistar rats were randomly divided into the control group and the normal RSLP high-dose group (RSLP-NH). The 40 SHR were randomly divided into the model group, Captopril group, RSLP-L group, RSLP-M group and RSLP-H group. SBP, DBP, HR, body weight and organ index were observed after the drug administration for 8 weeks and drug withdrawal for 2 weeks. The contents of SOD, MDA, GSH-Px, NO, NOS and ANP in serum were measured. The results showed that the blood pressure of SHR was significantly higher than that of the control group, which can be used for anti-hypertensive studies. Each RSLP group can obviously reduce the SBP and DBP of SHR (P < 0.05), but it had no effect on HR (P > 0.05). RSLP can elevate GSH-Px, SOD levels and reduce the activity of MDA (P < 0.05). RSLP can reduce NO, NOS and ANP contents in serum (P < 0.05). It was concluded that RSLP can significantly reduce the SBP and DBP of SHR, but it had no significant effect on HR. It can increase the activity of GSH-Px, SOD, NO, NOS levels, and reduce the contents of MDA, ANP in serum. It had certain inhibitory effect on the left ventricular hypertrophy.

Objective To investigate the mechanism of oxidative damage caused by lipopolysaccharide (LPS)induced sepsis in rat brain.Methods The rats were randomly divided into control group and model group (low LPS group and high LPS group).Twenty-four hours after the modeling,the rats were sacrificed before their brain tissue was taken out and prepared for the test.The changes in malondialdehyde (MDA),superoxide dismutase (SOD),glutathione peroxidase (GSH-Px),total antioxidant capacity (T-AOC),hydrogen peroxide (H2 O2 )and succinate dehydrogenase (SDH)were detected.The expression level of JNK and Nrf2 protein in brain tissue was detected by qRT-PCR and Western blotting. Results Compared with the control group,the MDA,SOD,GSH-px,T-AOC,H2O2 and SDH level increased significantly in the model group,and the difference in expressions of JNK and Nrf2 was statistically significant (P <0.05). Conclusion The LPS induced septic oxidative brain damage model in rats is successfully established,and the process may be regulated through the Nrf2 and JNK signal pathways.

This experiment aimed to explore and research the process of preparing baicalein and wogonin through liquid fermentation with Bacillus natto. Active enzymes of produced by B. natto was used for the biological transformation of baclin and wogonoside, in order to increase the content of the haicalein and wogonin in the scutellaria. With the content of the baicalein and wogonin as evaluating indexes, the effects of carbon source, nitrogen source, the types and suitable concentration of inorganic salt, medium pH, granularities of medical materials, liquid volume in flask, shaking speed, liquid-to-solid ratio, fermentation time on the fermentation process were studied. The optimal process conditions for liquid fermentation of scutellaria were 1.0% of peptone, 0.05% of NaCl, pH at 6, the granularities of medical materials of the scutellaria screened through 40-mesh sifter, 33% of liquid, shaker incubator speed at 200 r x min(-1), liquid-to-solid ratio of 5:1, temperature at 37 degrees C, fermentation for 6 days, baclin's conversion rate at 97.6% and wogonoside's conversion rate at 97% in the scutellaria. According to the verification test, the process was stable and feasible, and could provide data reference for the industrial production.
Bacillus subtilis ; metabolism ; Biotransformation ; Fermentation ; Flavanones ; metabolism ; Flavonoids ; metabolism ; Glucosides ; metabolism ; Soy Foods ; microbiology

Objective To observe the clinical efficacy of acupoint thread embedding plus Western medication in treating major epilepsy.Method Sixty epilepsy patients were randomized into a thread embedding group and a Western medication group, 30 cases each. The Western medication group was intervened by Western medication; the thread embedding group was given acupoint thread embedding based on the same Western medication treatment. Seizure frequency, seizure score, general efficacy and the improvement of adverse reactions due to Western medications were analyzed.Result Compared to the Western medication group, the thread embedding group was more effective in controlling seizure frequency (P<0.01), and reducing seizure score (P<0.05) and adverse reactions of Western medications (P<0.05); the general efficacy of the thread embedding group was superior to that of the Western medication group in treating epilepsy (P<0.05).Conclusion Acupoint thread embedding plus Western medication is effective in treating epilepsy, and is superior to the use of Western medication alone.

Objective To investigate the relationship between peripheral blood mtDNA content and acute coronary syndrome. Methods A total of 244 patients who received coronary angiography(CAG)was enrolled in this study. They were divided into acute coronary syndrome(ACS)group(n=183)and control group(n=61)ac-cording to the CAG results. A quantitative real-time PCR-based method was used to measure relative content of mtDNA in peripheral blood. According to lesion blood vessel counts ,ACS patients were divided into single lesion group,double lesion group and three lesions group. Gensini score was used to quantitatively evaluate the severity of coronary stenotic lesions. Then we analyzed the relationship of mitochondrial DNA content with the lesion counts and the Gensini score. Results The ACS group had a lower mtDNA content ,as compared to controls (P <0.001).The lesion blood vessel count and mtDNA content in the single ,double and three vascular lesions group were all significanlty lower than that of the control group(P<0.05)and that of the double,three vascular lesions group were both significantly lower than that of the single vascular lesion group(P<0.05). According to the Gensi-ni score,all patients were divided into four groups,where with mtDNA content was decreased with the increase of the Gensini score(P < 0.01). The linear regression analysis showed that Gensini score was negatively correlated with mtDNA content(r=-0.644,P<0.001). ROC curve analysis showed that the area under the curve of mtDNA content for ACS diagnosis was 0.855(P<0.001),with the sensitivity and specificity of 0.756 and 0.833,respec-tively. Conclusion MtDNA content is closely associated with ACS and the Gensini score ,the latter indicating the severity of coronary stenotic lesions. MtDNA content detection has its value in the diagnosis of ACS.