BACKGROUND:The low output of seed cells and long cycle of traditional ceils culture methods in tendon animal models(rabbits and chicken) restrict the researches of tendon tissue engineering study.OBJECTIVE:To establish an ideal culture protocol of tail tendon in SD rats,to get more seed cells within less time for subsequent engineered tendon construction research.DESIGN,TIME AND SETTING:Controlled observation was performed in the National Key Laboratory of Biotherapy,Department of Stem Cells and Tissue Engineering,Sichuan University between February and November in 2006.MATERIALS:Rat tail tendon cells were harvested from 2 SD rats,aged 7-10 days;human prepuce fibroblasts were offered by National Key Laboratory of Biotherapy,Department of Stem Cells and Tissue Engineering,Sichuan University.METHODS:Tail tendon of SD rats was draw off and cut into pieces,which were then cultured in 10% fetal bovine serum+DFculture medium for getting primary tendoncyte by using suspension tissue culture method. The third generation cells were processed into immuocytochemistry stain with collagen type Ⅰ and Ⅲ,while human prepuce fibroblasts served as controls.Absorbance of stain result was measured by image-pro plus 5.02 for statistical analysis.MAIN OUTCOME MEASURES:Immuocytochemistry stain and absorbance measurement of SD rat tail tendon cells.RESULTS:The second generation of SD rat tail tendon cells were positive for type Ⅰ collagen stain,and negative for type Ⅲ collagen stain;human fibroblast were positive for both Ⅰ and Ⅲ collagen. In the rat tail tendon cells and human flbroblasts,the absorbance value of type Ⅰ collagen expression was dramatically higher than of type Ⅲ collagen(P＜0.05). There was no significant differences addressing the absorbance of type Ⅲ collagen expression between type Ⅰ and Ⅲ collagen of SD rat tail tendon cells and blank control group (P＞0.05).CONCLUSION:Cells cultured from SD rat tail tendon have biological characteristic of tendon cells. Tissue piece suspensionculture can obtain a quantity of primary or subcuitured cells of rat tail tendon within a short time.

Objective To compare skin barrier function among patients with facial acne,subacute eczema,melasma and solar dermatitis.Methods Three hundred patients,including 80 patients with facial acne,60 subacute facial eczema,80 facial melasma and 60 facial solar dermatitis,as well as 60 healthy controls were recruited in this study.Skin sebum content and transepidermal water loss (TEWL) were measured by a sebmeter and Tewameter TM 210 (Courage and Khazaka,Germany),respectively.Stratum comeum hydration was measured with a Scalar Moisture Checker (Scalar Corporation,Japan).Statistical analysis was carried out using analysis of variance and t test.Results Compared with the healthy controls,patients with facial acne showed increased skin sebum content and TEWL value but decreased stratum corneum hydration (all P ＜ 0.01),and patients with subacute eczema,solar dermatitis and melasma displayed lower sebum content and stratum corneum hydration but higher TEWL value (all P ＜ 0.01).Skin sebum content was significantly higher in patients with facial acne than in patients with subacute eczema,solar dermatitis and melasma ((184.65 ± 83.07) vs.(21.86 ± 18.94),(25.10 ±14.22) and (36.05 ± 32.84) μg/cm2,all P ＜ 0.01),but was similar between the patients with subacute eczema,solar dermatitis and melasma (P ＞ 0.05).In terms of stratum corneum hydration,patients with subacute eczema and solar dermatitis were statistically lower than those with acne and melasma (18.66％ ± 7.85％ and 20.91％ ± 8.05％ vs.24.32％ ± 8.16％ and 28.02％ ± 4.67％,all P ＜ 0.01),patients with facial subacute eczema were similar to those with solar dermatitis (P ＞ 0.05),and patients with facial acne were statistically lower than those with melasma (P ＜0.01).TEWL value was significantly higher in patients with melasma than in patients with acne,solar dermatitis and subacute eczema ((13.80 ± 4.t 3) vs.(20.86 ± 8.78),(22.85 ± 9.84) and (22.48 ± 10.37) μg/m2 h,all P ＜ 0.01),but similar between patients with acne,solar dermatitis and subacute eczema (P ＞ 0.05).Conclusions Skin barrier function is somewhat impaired in patients with facial acne,subacute eczema,melasma and solar dermatitis.Therefore,to recover skin barrier function may facilitate the treatment of these diseases.

Objective To explore the correlation between caring behavior and critical thinking of nursing students in clinical practice. Methods Totally 203 nursing students in clinical practice were investigated with Critical Thinking Disposition Inventory- Chinese Version (CTDI- CV) and Caring Behavior Scale (CBS). Results The structural model obtained a good fit. Critical thinking and caring behavior were positively correlated with each other(β=0.46,P<0.01). Conclusions Nursing students′caring behavior could improve their critical thinking. With the cultivation of critical thinking of nursing students, nursing educators should pay attention to improving their caring behavior.

Objective To construct the vector that expresses the fusion protein of HIV-Tat protein and red fluorescent protein(mCherry) in mammalian cells,and observe by fluorescence microscopy the intracellular transduction and localization of recombinant protein in cells,in order to obtain a useful tool for the study of the uptake mechanism and intracellular localization of HIV-TAT.Methods With the designed primer coding mCherry sequence,the mCherry gene was amplified by PCR with the vector pmCherry-C2 as template,and inserted into vector pET14b-His-TAT to construct the expression vector pET14b-His-TAT-mCherry.The constructed vector was then transformed into E.coli BL21(DE3),which had been identified by PCR and double digested with restriction endonuclease,followed by sequencing.After IPTG induction,the recombinant protein of His-TAT-mCherry was lyzed and analyzed with SDS-PAGE.Purified His-TAT-mCherry recombinant protein was added to Hela cells and the fluorescence was observed to evaluate the transduction efficiency.Results The results of identification by PCR,digestion with restriction endonuclease and sequencing indicated that the vector His-TAT-mCherry was correctly constructed.His-TAT-mCherry fusion protein was expressed in mammalian Hela cell line and purified successfully,and the fusion protein showed cellular transduction activity.It was found by fluorescence microscopy that the red fluorescence protein located mainly over the cytoplasm,and also the membrane to some extent.Conclusion The expression vector is successfully constructed for HIV-TAT labeled with mCherry sequence.Effective expression and purification of this fusion protein is achieved.It has been observed that the constructed vector may be expressed in mammalian Hela cell under active condition.Thus,it might be useful in the study of uptake mechanism and intracellular localization of HIV-TAT.

Objective To investigate the radiosensitiation effect of berberine on human nasopharyngeal carcinoma ( NPC) in hypoxia condition and explore the underlying mechanisms. Methods MTT assay, clonogenic assay and flow cytometry were performed to analyze cell proliferation, colony formation and apoptosis, respectively. Male nude mice inoculated subcutaneously with CNE-2 cells were used to examine the radiosensitization effect of berberine in vivo. The expressions of HIF-1α and VEGF were assessed by Western blot. Results Berberine efficiently inhibited the proliferation of CNE-2 cells in time-dependent and dose-dependent fashions with an IC50 of ( 14?9 ± 2?2 ) μmol/L. Clonogenic survival assay showed that berberine ( 5 μmol/L ) sensitized CNE-2 cells to ionizing radiation in hypoxia and its SERD0 was 1?27. Under hypoxic condition, berberine alone (5, 15 μmol/L) could induce apoptosis (t=5?01, 9?02,P<0?05) and it further promoted 8 Gy radiation-induced apoptosis (t =5?31, 9?91,P <0?05). Moreover, berberine significantly delayed the tumor growth in the combination group (berberine +irradiation) compared with the mice received irradiation alone or PBS (t =2?96, 14?52, P <0?05). Immunobloting assay showed that berberine inhibited the upregulation of HIF-1α and VEGF induced by hypoxia in CNE-2 cells. Conclusion Berberine confers radiosensitivity on hypoxic NPC in vitro and in vivo, which is probably associated with the downregulation of HIF-1α and VEGF expressions.

To enhance the quality and efficiency of ozagrel by investigating the differences between the ozagrel polymorphs in bioavailability. Solid ozagrel in different polymorph forms were orally administered to SD rats. An HPLC method was established to determinate plasma level of ozagrel. The bioavailabilities of two polymorph forms were calculated and compared. The pharmacokinetic parameters of ozagrel, were as follows: Cmax was 32.72 ± 17.04 and 34.01 ± 19.13 mg · L(-1), respectively; AUC0-t was 61.14 ± 14.76 and 85.56 ± 18.08 mg · L(-1) · h, respectively; t½ was 1.53 ± 0.51 and 4.73 ± 3.00 h, respectively. There was no significant difference in pharmacokinetic parameters between form I and II polymorphs of ozagrel while the t½ of form II is longer, which indicates that the use of form II polymorph as pharmaceutical product may prolong the effective action time in clinics. This would help the polymorph quality control in drug production.
Animals ; Biological Availability ; Chromatography, High Pressure Liquid ; Methacrylates ; chemistry ; pharmacokinetics ; Rats ; Rats, Sprague-Dawley

Objective To evaluate the effects of electroacupuncture (EA) on synaptic interface structures around a cerebral infarct and evaluate how learning and memory relate to synoptic structure. Methods Forty-eight male, adult Wistar rats were divided randomly and equally into an EA group and a control group after models of ische-mic brain injury had been established. Each group was then divided into 1 week, 2 week and 3 week sub-groups. In the EA group EA was started 24 hours after the operation at the Baihui (DU20) and Da Zhui (BUI4) points and continued for one week, two weeks or three weeks. The control group was fed in normal cages and had no special treatment. Synoptic substructures in the hippocampal CA3 area of the ischemic side were assessed morphologically and quantitatively. Results The synaptic curvatures and postsynaptic density (PSD) an well as the width of the ac-tive area increased significantly in the EA group compared with the control group. Morris water maze learning ability in the EA group exceeded that in the control group. Spatial probe test results also significantly improved in the EA groups compared with the control groups. Conclusions Learning and memory improved in the injured rats following EA, and this is likely to be related to synaptic changes and synaptic plasticity increasing.

Objective To evaluate the performance of nested PCR in the detection of different fungi in paraffin wax embedded tissues. Methods Forty-four tissue samples were resected from rats infected with Fonsecaea monophora, patients with chromoblastomycosis, sporotrichosis or penicilliposis marneffei followed by preparation of paraffin wax embedded tissue sections for pathological examination and DNA extraction. Nested PCR was performed by using specific primers targeting the ribosomal DNA of Fonsecaea, Sporothrix and Penicillium marneffei, respectively. The sensitivity and specificity of nested PCR were analyzed and compared with those of pathological examination. Results The nested PCR showed positive results in 8 of 20 samples from rats with chromoblastomycosis, 7 of 10 samples from patients with sporotrichosis and all of the 10 samples from patients with penicilliposis marneffei, but not in the control samples. In the detection of Fonsecaea,Sporothrix schenki and Penicillium marneffei, the sensitivity was 40% ,70% and 100%, respectively, and the specificity was consistently 100%, for the nested PCR. Pathological examination revealed fungal elements in 95%, 70% and 80% of the corresponding samples, respectively. Conclusion Detection of fungal DNA in paraffin wax embedded tissue by nested PCR can be applied to the diagnosis of deep mycosis, especially to the diagnosis of penicilliposis marneffei.

Objective To investigate the risk factors of pulmonary hypertension (PH) after rapid ascent to high altitude in healthy young Chinese men .Methods A total of 552 non high altitude natives ( healthy young Chinese men ) were recruited in the study between June to August in 2012.The subjects were delivered to high altitude (3700 m) in 2 h by plane from low altitude (450 m).Demographic data including smoking , alcohol consumption , high altitude exposure histo-ry, body mass index (BMI) and age were collected within 24 h after arrival at 3700 m.Furthermore, the oxygen saturation ( SaO2 ) , blood pressure , heart rate , pulmonary artery systolic pressure were measured using the echocardiography system . Analysis of risk factors of PH was performed by logistic regressions .Results The incidence of PH was 23.10%.SaO2 was significantly lower in the higher-pulmonary artery systolic pressure group (higher-PASP)[(86.10 ±3.34)%] than that in the normal-PASP group [(89.09 ±2.65)%, P<0.01].Smoking, alcohol consumption, high altitude exposure history, BMI, age, systolic blood pressure , diastolic blood pressure , heart rate were not significantly different between the two groups mentioned above .Logistic regressions revealed that lower SaO 2 (OR:0.711,95% CI:0.647-0.782, P<0.01) was an independent risks factor of PH .Conclusion Low SaO2 is an independent risk factor of PH upon rapid arrival at 3700 m in healthy young Chinese men .However, smoking, alcohol consumption , high altitude exposure history , BMI, age, blood pressure and heart rate are not risk of PH .These observations will provide valuable clues to theoretical studies on PH and prevention of PH .

OBJECTIVETo investigate the clinical effects of 360 degree circular decompression and transpedicle screw fixation to treat the ossification of thoracic posterior longitudinal ligament by posterior approach.

METHODSFrom December 2009 to November 2013, 18 patients with ossification of thoracic posterior longitudinal ligament ossification were treated with 360 degree circle decompression and transpedicle screw fixation by posterior approach. There were 8 males and 10 females,ranging in age from 32 to 67 years old, with an average of 51 years old. Four patients were accompanied with ligamentum flavum ossification. Longitudinal ossifications in 5 cases were located in the upper thoracic vertebra and 13 cases in the middle and lower thoracic vertebra. Five cases were typical type, 4 cases were segmental type, 6 cases were continuous type and 3 cases were mixed type. All the patients had the posterior spinal canal decompression combined with longitudinal ligament resection, interbody fusion with bone graft and internal fixation surgeries. The operation time,blood loss and complications were recorded. JOA score were applied to evaluate the neurological function recovery pre-surgery, 2 days post-surgery and the latest follow-up. The surgery effects were evaluated by Epstein-Schwall method.

RESULTSThe operation time ranged from 3 to 6 hours (mean, 4.2 hours). The blood loss ranged from 800 to 4 000 ml (mean, 1 800 ml). All the patients were followed up, and the duration ranged from 6 months to 3 years, with a mean of 1.8 years. The JOA score increased from preoperative 4.30 ± 2.60 to 7.60 ± 2.40 2 days after surgery, and 7.80 ± 1.90 at the latest follow-up (t = 4.61, P < 0.001). The JOA scores between 2 days after surgery and the latest follow-up had no significant differences (t = 0.28,P = 0.78). The neurological recovery rate was 74% 2 days after surgery and 71% at the latest follow-up. There were 4 cases got an excellent result,10 good,3 fair and 1 poor according to Epstein-Schwall evaluation method. Four patients had cerebrospinal fluid leakage, 3 patients had intercostal nerve paralysis or pain, and 1 patient had superficial incision infection. The neurological function in 3 patients became worse in the second day posteratively , and among them, 2 patients were recovered at the latest follow-up and 1 patient had no changes. All the patients got fusion of bone graft and no internal fixation loosening and fractures occurred.

CONCLUSION360 degree circular decompression and transpedicle screw fixation can resect different types of thoracic longitudinal ligament ossification, and can achieve a good clinical effect.

Adult ; Aged ; Decompression, Surgical ; methods ; Female ; Fracture Fixation, Internal ; methods ; Humans ; Male ; Middle Aged ; Ossification of Posterior Longitudinal Ligament ; surgery ; Thoracic Vertebrae ; surgery