Background The early disorder of diabetic retinopathy (DR) is the damage of retinal neural cells induced by high glucose and lack of oxygen.Previous studies show that bushenhuoxue serum can enhance the activity of glutamine synthetase (GS) in Müller cells under hypoxia,and glutamate-mediated retinal excitotoxicity also can be reduced by bushenhuoxue serum intervention.However,whether the concentration of glycine can be increased by bushenhuoxue serum is not clear.Objective This study was to investigate the protective effects of bushenhuoxue serum on retinal ganglion cells (RGCs) under hypoxia.Methods The Sprague Dawley (SD) rat serum containing bushenhuoxue was prepared.The RGCs of newborn SD rats were purified and identified by a twostep immunopanning procedure.After 72 hours,all RGCs were cultured in 96-well plates and divided into four groups:normal control group (cultured in adult SD rats normal serum),bushenhuoxue group (cultured in bushenhuoxue serum),hypoxia group (cultured in 1 mmol/L sodium dithionite); hypoxia + bushenhuoxue intervention group (cultured in bushenhuoxue serum+sodium dithionite).Glutamate and glycine contents in the extracellular fluid were detected by L-8800 automatic amino acid analyzer,and the content of lactate dehydrogenase (LDH) was assayed using LDH kits in 24,48 and 72 hours after culture.Results Cultured cells showed the green fluorescence under the immnofluorescence microscope.The contents of glutamate,glycine and LDH in the extracellular fluid were (0.0805±0.0010)mg/L,(0.0554±0.001 5)mg/L and (1 626.03 ±122.10)μmol/(min · L) in the normal control group in 24 hours after culture,and those in the hypoxia group were (0.022 5±0.001 1) mg/L,(0.014 6±0.001 1)mg/L and (1 458.68±94.23)μμmol/(min · L),showing significant reducing in the hypoxia group (q =-3.53,P =0.00 ; q =-2.45,P =0.00 ; q =-2.98,P =0.02).Compared with the normal control group,LDH and glycine contents in the extracellular fluid were significant raised in the hypoxia group 48 hours after culture (q =2.55,P =0.01 ;q =4.48,P =0.00).Seventy two hours after culture,the glutamate and glycine contents in the hypoxia group were higher than those of the normal control group (q =2.45,P =0.00 ;q =3.72,P =0.00).In 48 and 72 hours of culture,the contents of glycine were (0.017 4±0.001 5) and (0.019 2±0.001 2) mg/L in the hypoxia+bushenhuoxue intervention group,which were significantly higher than (0.016 0±0.001 2) and (0.018 0±0.000 8) mg/L in the hypoxiagroup (q=2.28,P=0.04;q=2.33,P=0.03),but the LDH level were (1 632.94±264.31) and (1 875.00±137.45)μmol/(min · L) in the hypoxia+ bushenhuoxue intervention group,which were lower than (1 688.49 ± 112.86) and (2 267.86 ± 175.21) μmol/(min · L) of the hypoxia group (q =-2.95,P =0.02 ; q =-2.35,P=0.00).No significant differences were seen in the glutamate content 24,48 and 72 hours after culture (P=0.55,0.28,0.46).A positive correlation was seen between the glutamate content and glycine content in the extracellular fluid (Kendall coefficient =0.519,Spearman coefficient =0.696,both at P =0.000).Conclusions The release levels of glutamate and glycine increase in the hypoxia RGCs,which probably is a compensatory response of RGCs.Bushenhuoxue serum can protect RGCs against injury by increasing the release of glycine and decreasing the LDH leakage from RGCs.

Objective: To explore the mechanism of anti-aging effect of acupuncture at Zusanli (ST 36). Methods: Thirty mice of 28 weeks old were randomly divided into 3 groups: control group, model group and acupuncture group. Sub-acute aging model was established by subcutaneous injection of D-galactose. The activity of Superoxide Dismutase(SOD) and content of serum Malondialdehyde(MDA) were measured. Morphological changes of jejunal mucosa were observed with light microscope. Result: SOD activity in the acupuncture group was highest among the three groups, whereas, MDA content was the lowest. In the model group, the ratio of normal intestinal villus was decreased, the intestinal villus was shrunk, and there were more epithelial cells desquamated. In acupuncture group, there were less abnormal intestinal villus and epithelial cell. Conclusion: Acupuncture at Zusanli (ST 36) has an effect in resisting aging by increasing the activity of serum SOD, decreasing the MDA contents, and improving the intestinal villus.

Objective To observe the effect of shenfu injection on ischemia/reperfusion injury in skeletal muscle of rats. Methods To make a model of ischemia/reperfusion injury in skeletal muscle of rats, and observe the plasma level of LDH, CK, MDA and the change of MDA, MPO, W/D in the skeletal muscle. Result Shenfu injection can reduce the heightened plasma level of LDH, CK, MDA and MDA, MPO, W/D of the skeletal muscle, lighten the injury of the ultrastructure. Conclusion Shenfu Injection can protect ischemia/reperfusion injury in skeletal muscle.

OBJECTIVE: To evaluate function and application of various biomaterials in tissue engineering repair skull defect, and to seek a reasonable skull substitutes. METHODS: Using "tissue engineering, skull defect, stem cells, cytokines, composite" as key words in Chinese, and using "tissue engineering, skull defect, mesenchymal stem cells, cytokines, materials" as key words in English, a computer-based online search was performed for publications from January 1993 to October 2009. Articles concerning biomaterials and tissue engineering skull defect repair were included. Articles describing repetitive study or Mete analysis were excluded. Function of various biomaterials in tissue engineering repair skull defect was assess in 17 articles. RESULTS: Stem cell technique is to obtain seed cells from a few myeloid tissues. Following in vitro induction and amplification, cells at a certain number could compound with stents, and construct tissue-engineered bone in vitro. The cells could sustain the biological features of osteoblasts, and fuse with bone tissue in the recipient site. Simultaneously, it could avoid the occurrence of immunological rejection. Two or over materials were compounded together, or biomaterial surface received various modification. These could promote adhesion between cells and materials, elevate cell bioactivity and maintain biological function. Following moved into seed cells using some vectors, the latter ossified in bone defect site, and secreted suitable bone growth factor, induced the differentiation of peripheral non-oriented osteogenitor cells into oriented osteogenitor cells, which could harvest adequate seed cells with osteogenic activity in a short time, and accelerate the healing of skull defects. The composite made by using three-dimensional virtual imaging and computer numerical control modeling techniques has the advantages of medical composite, precise anatomical consistency with surrounding tissues of skull defects, and perfect appearance. CONCLUSION: At present, no material can be accorded with the requirement of bone tissue engineering. Combination of some materials or modification of biomaterial surface can promote adhesion of cells and materials, elevate cell biological activity, and maintain biological function, which are presently hot focus of tissue engineering research of biomaterials.

Purpose　The aim is to develop a synthesis method of the silver carboxymethyl chitosan and to study its bacteriostasis to Staphlococcus aureus(S.aureus),Pseudomonas aeruginusa(P.aeruginusa),Escherichia Coli(E.coli),Klebsiella pneumoniae(K.pneumoniae) and Proteus vulgaris(P.vulgaris).Methods　Chitosan was modified by way of chemistry.The structure analysis of its derivate was analysed by infra-red absorption spectroscopy,using methods of dilution and concavering to study the bacteriostasis to some ordinary bacteria which cause infection in burn.Results　The infra-red spectragram of the derivate showed the chitosan had been modified by chloroacetic acid.When the concentrationl of silver carboxymethyl chitosan was 1.028 mg/ml and the concentration of the five bacteria were 104CFU/ml，the rate of bacteriostasis was 88%、80.2%、75.3% to S.aureus、P.aeruginusa and E.coli respectively.The MIC of silver carboxymethyl chitosan was similar to that of AgNO3 when they act on S.aureus and P.aeruginusa,but the former was lower than later when acting on E.coli.Conclusion　Silver carboxymethyl chitosan could inhibit some bacteria which caused infection in burn.It was a novel pharmaceutical in preventing and curing burn infection.

Objective To investigate the survival rate and its prognostic factors of laryngeal carcinoma. Methods We retrospectively analyzed 63 cases of laryngeal carcinoma. A total of 7 clinicopathologic factors were studied by univariate analysis and Cox multivariate model. Results The overall cumulative survival rate was 69.8% at 3 years,54.0%at 5 years.In univariate analysis, the survival was related to location of the tumor(P< 0.01), T status (P < 0.01), N status (P < 0.01), clinical stages (P < 0.01), pathologic grade (P < 0.01) and types of treatment (P < 0.01), while age was not correlated to prognosis (P > 0.05). In Cox multivariate model, T status (P < 0.01) and N status (P < 0.01) were independent prognostic factors. Conclusions T and N status were independent prognostic factors for patients with laryngeal carcinoma. Early detection and treatment should be given to improve the survival of patients.

Objective: To investgate the role of Toll-like receptor(TLR) 2 and TLR4 in pathogenesis of atopic dermatitis(AD) and the effect of topical tacrolimus ointment on expression of TLR2 and TLR4 in lesional AD skin.Methods: Immunohistochemistry was employed to study the expression of TLR2 and TLR4 in normal skin and lesional AD skin before and after using topical tacrolimus ointment.Results: The basal keratinocytes in normal skin constitutively expressed TLR2 and TLR4. In contrast,lesional epidermis from 9 patients with acute AD overexpressed TLR2 and TLR4 on the whole epidermis keratinocytes with membranous and cytoplasmic staining pattern.After using topical tacrolimus ointment for three weeks,TLR2 and TLR4 were expressed on basal and suprabasal keratinocytes with membranous and cytoplasmic staining pattern.Conclusion: These data suggest that TLR2 and TLR4 expressed by epidermal keratinocytes constitute part of the innate immune system of the skin,and increased TLR2 and TLR4 expression may be related to the skin innate immuno-inflammatory response in atopic dermatitis.Topical tacrolimus may directly or indirectly inhibit or downregulate TLR2 and TLR4 expression in KC and inhibit skin innate immuno-inflammatory response related to TLR-NF?B signal transduction and regulation in atopic dermatitis.

Objective: To sieve the bioactive constituents of Radix Puerariae,serum pharmacochemistry research was performed.Method: Based on the establishment of HPLC fingerprints of Radix Puerariae,the constituents absorbed into blood were determined by comparing the HPLC fingerprints of the methanol extracts,tested serum samples and blank serum sample.Results: Four compounds absorbed into blood were detected,among which two were original constituents of Radix Puerariae(including puerarin),the other might be metabolites of the original constituents.Conclusion: These four constituents absorbed into blood were possible bioactive components of Radix Puerariae.Further studies on them will help clarify the bioactive constituents and mechanisms of Radix Puerariae.

Objective To investigate the relationship between the matrix metalloproteinase-3(MMP-3)and the stability of carotid artery plaque,and explore MMP-3's prediction role on the attack and relapse of acute ischemic cerebrovascular events.Methods 100 patients with the first ever acute cerebral infarction,100 patients with chronic cerebral circulation insufficiency(CCCI)and 40 persons without cerebrovascular diseases were enrolled in this study.According to the carotid ultrasound examination,100 cerebral infarction patients were divided into three subgroup: unstable plaque group(45 patients,mixed plaque,soft plaque),stable plaque group(35 patients,plaque Group)and endometrial coarse group(25patients).Matrix metalloproteinase-3(MMP-3)levels of all the subjects were measured by enzyme-linked immunosorbent assay(as basal level).All the subjects were followed up for one year to observe cerebral infarction events.Serum MMP-3 levels of each group,and the basic serum MMP-3 levels were compared among patients who were attacked or relapsed cerebral ischemic with those who had not been attack cerebral ischemic during this period of time.Results 5 patients in the cerebral infarction group had relapse (5%),2 patients in the CCCI group were attacked by cerebral ischemic(2%),and no one in the normal control group was attacked by cerebral ischemic.Serum MMP-3 levels in the acute cerebral infarction group were significantly higher than CCCI group,and both groups were significantly higher than normal control group (P ＜0.05).The basic serum MMP-3 levels in all patients who were attacked by cerebral ischemic were significantly higher than those who had not been attack by cerebral ischemic during this period of time(P ＜0.05).The serum MMP-3 levels of the unstable plaque group were significantly higher than stable plaque group.And both groups were significantly higher than endometrial coarse group(P ＜0.05).Conclusions Elevated levels of matrix metalloproteinase-3(MMP-3)might have something with the stability of carotid atherosclerotic plaque,and participate the attack and the relapse of acute cerebral infarction.Determination of MMP-3 might be used to predict the attack and relapse of acute cerebral infarction.

Objective To investigate the change of serum cystatin C (CC) level and its clinical significance in elderly patients with obstructive sleep apnea hypopnea syndrome (OSAHS),and to evaluate the diagnostic value of serum CC for renal impairment in elderly OSAHS patients.Methods Elderly snoring subjects (age ≥60 years) undergoing sleep apnea monitoring in our sleep laboratory were recruited.Patients were divided into simple snoring group,mild OSAHS group,moderate OSAHS group and severe OSAHS group according to the diagnostic criteria of American academy of sleep medicine.Anthropometric characteristics,lipid profiles,blood glucose,creatinine,estimated glomerular filtration rate (eGFR) and serum CC were compared between groups after age,gender and basic diseases were adjusted.The independent risk factors associated with serum CC were analyzed by Spearman's rank correlation and stepwise linear regression.Results Totally 89 elderly patients [70males and 29 females,aged (68.4±6.3) years in average] were collected.21 subjects were grouped as simple snoring,14 subjects as mild OSAHS,21 subjects as moderate OSAHS and 33 subjects as severe OSAHS.There were no differences in age,gender,basic diseases (hypertension and diabetes),lipid profiles,creatinine and eGFR among groups.The levels of body mass index,systolic blood pressure,diastolic blood pressure and serum CC were higher in severe OSAHS group than in simple snoring group [(28.3±3.9) vs.(24.6±5.9),(136.55±16.14) mm Hgvs.(122.43±16.64) mm Hg,(89.18±11.92) mm Hgvs.(78.10±9.55) mm Hg,(0.95±0.22) mg/L vs.(0.76±0.13)mg/L,respectively,all P＜0.05].Serum CC was significantly correlated with apnea hypopnea index (AHI),oxygen desaturation index,creatinine,eGFR,high sensitive C reactive protein (r=0.397,0.347,0.275,－0.354,0.254 respectively,all P％0.05).Regression analysis showed that AHI was an independent factor for serum CC (β=0.449,P＜ 0.001).Conclusions Serum CC level is increased in elderly patients with severe OSAHS.AHI is an independent factor for the increase of serum CC.Serum CC can be as an index for the early impairment of renal function in patients with OSAHS.