Objective To establish a simple and feasible method in analyzing the molecular weight distribution of peptides in thymosin.Methods Improved Tricine-SDS-PAGE electrophoresis was applied to detect the molecular weight of peptides and the content of thymosinα1 in thymosin.Results Tested the thymosin preparations on the domestic market using this im-proved method.It was demonstrated that the peptide molecular weight distribution in thymosin preparations was between 3.5~8.5 kD,also could detect the concentration of 1 μg thymosinα1 in thymosin by using this improved method.Conclusion This improved method is suitable for the analysis of peptides molecular weight distribution and the concentration of thy-mosinα1,so it can be used for control quality of thymosin preparations.

Objective To investigate the expression and clinical significance of α-actinin-1 protein (ACTN1) in prostate cancer (PCa) and prostatic hyperplasia (BPH). Methods The clinical data of patients with PCa or BPH treated in our school affiliated hospital were collected between January 2007—October 2014, according to certain criteria. Immunohistochemistry method was used to detect the expression of ACTN1 in 30 samples of PCa and 30 samples of BPH tissues. Western blot assay was used to detect the relative expression of ACTN1 in 18 samples of PCa and 20 samples of BPH tissues in two groups. Results The result of immunohistochemistry showed that the positive expression rates of ACTN1 were 76.7%and 20%in PCa and BPH groups respectively. The difference was statistically significant (P<0.01). Western blot assay showed that the relative expression of ACTN1was significantly higher in PCa group (0.591±0.182) than that in BPH group (0.037 ± 0.052, P < 0.05). There was no significant difference in expression level of ACTN1 between different age groups. There was no significant difference in serum prostate specific antigen (PSA) level between patients with or without bone metastasis, and patients with or without lymph node metastasis (P > 0.05). There were significant differences in ACTN1 levels between different Gleason score and T staging groups (P<0.05). Conclusion The expression ofα-actinin-1 is significantly higher in PCa tissues than that in BPH tissues. There is the relationship between expression of ACTN1, Gleason scores and T staging.

ObjectivesTo detect the effects of P-selectin on deep venous thrombosis (DVT) in nephrotic syndrome (NS). and to evaluate the molecular magnetic resonance imaging (MRI) with a P-selectin targeted conlrost agent in diagnosis of thrombosis in the early phase. Methods(1) Forty-one patients with NS hospitalized in our department from 2005 to 2006 were enrolled in this study. They were assigned into DVT group and non-DVT group according to lower limbs radionuclide imaging (RNV) with 99mTc MAA. Blood P-selectin level was measured by ELISA method. (2) P-selectin was detected both in injured vein and blood immediately, 1 h and 3 h after the dog DVT model was established. (3) The P-selectin-targeted contrast agent was developed by conjugating anti-P-selectin lectin-EGF domain monoclonal antibody (PsL-EGFmAb) which was prepared by our lab. The potential of this contrast agent used in vitro molecular imaging experiment as well as in vivo experiment in dog DVT model was investigated. Results (1) Blood P-selectin level was elevated in patients with NS. It was much higher in DVT group than that in non-DVT group. (2) Blood P-selectin level was also elevated in DVT dogs and P-selectin expressed immediately in tunica intima of injured vein and subsequently in thrombus after the model established. (3) Mural thrombus showed higher signal visualization than surrounding muscle in 30 rain after contrast agent injection. These enhanced signals exhibited P-selectin specificity and persisted from the initiation of intima lesions to 3 h after development of thrombosis. There was signficant Differences in contrast-to-noise ratio (CNR) of the experiment group and the control group (11.50±2.32 vs 2.71±0.86, P＜0.01). The same results were derived from 30 rain to 1 hafter contrast agent being injected in distal to heart part of the injured vessel, and the signal decreased 24 h later. Differences in CNR of the experiment group and the control group were also statistically significant (10.40±2.15 vs 1.93±0.57, P＜0.01). Moreover, the contrast agent did not affect the vital signs of the dog. The function of the heart, lung, liver and kidney functions remained normal after contrast administration. Conclusions P-selectin*targeted new MR contrast can be used to early locate thrombus in vivo in an early stage, which does not compromise the function of the important organs. It may become a new method for early diagnosis of thrombosis.

Objective:To investigate the risk factors for overall survival in operable hepatocellular carcinoma with portal vein tumor thrombus (PVTT-HCC) patients and establish a scoring system.Methods:Survival data in 253 PVTT-HCC patients were retrospectively analyzed in Guangxi Medical University Affiliated Tumor Hospital. Survival curves were analyzed using the Kaplan-Meier method and log-rank test. Cox stepwise regression analysis was used to identify independent preoperative risk factors affecting overall survival. A prognostic scoring system based on independent risk factors and their relative coefficients was established to screen patients with greater hepatic resection benefits, and the identification ability of the model was based on ROC.Results:A total of 253 patients with PVTT-HCC were enrolled in this study, there were 222 males and 31 females, with a median age 44 years. The median survival time in all patients was (13.00±2.15) months. Rate of overall survival was 51.8% at 1 year, 25.0% at 3 years and 17.7% at 5 years. Multivariable Cox regression analyses showed four risk factors including: AST≥40 U/L, ALP (≥80 U/L), tumor number (>1), and incomplete tumor capsule. A prognostic scoring system was established based on these variables. The area under curve of the scoring system was 0.780 (95% CI: 0.715-0.845). Patients were classified as low- or high-risk group for hepatic resection depending on whether their score was <3 ( n=77) or ≥3 ( n=176), respectively. High-risk patients had a median survival of 10 months, compared to 29 months in low-risk patients. Low-risk patients also had better survival rates at 1 year (75.3% vs 41.5%), 3 years (47.6% vs 15.2%), and 5 years (34.7% vs 10.5%), P<0.05. Conclusion:A prognostic scoring system for hepatic resection in PVTT-HCC patients has been developed based entirely on preoperative variables. Using this system, patients belong to the low risk group have better prognosis after surgery, which can provide a basis for surgical treatment of PVTT-HCC patients.

Objective To investigate the therapeutic effects and mechanism of anti-radiation pneumonia decoction(ARPD) on radiation induced lung fibrosis in rats.Methods One hundred and five male SD rats in a SPF grade were divided into Chinese medicine group,single radiation group and control group by random digits table method,with 35 in each group.After anesthetization,rats in Chinese medicine and single radiation groups were exposed to 6 MV X-rays at the dose of 15Gy.Rats in Chinese medicine group were treated with ARPD at the dosage of 10 ml·kg-1 ·d-1 once a day,but rats in single radiation group did not receive ARPD treatment.Rats in control group were treated with neither irradiation nor drugs.Five rats of each group were killed and the lung tissues and blood samples were collected at 15,30,60,75,90,105 and 140 d.The pathological changes of lung tissues were observed and the tissue protein and gene expressions of TGF-β1,PAI-1 and collagen type Ⅲ(C Ⅲ) were assayed by Western blot and RT-PCR.ELISA was used to detect serum TGF-β1 and plasma PAI-1.Tissue and serum HYP were determined by acid hydrolysis and alkaline hydrolysis methods respectively.Results Inflammation was found in the lung tissues of all the exposed rats.Obvious pathological lung fibrosis was found at 60 d,the inflammation and the fibrosis in treated group were slighter than those in single radiation group.In Chinese medicine group,the protein and gene expression levels of TGF-β1,PAI-1,C Ⅲ 30 d(Protein:t =2.49-3.74,t =2.63-4.57 and t =2.76-3.83;Gene:t =2.59-4.33,t =2.83-4.62 and t =2.83-3.96,P＜0.05),serum TGF-β1 and plasma PAI-1 15 dlater (t =2.85-6.27 and t =3.69-5.27,P＜0.05),and the levels of tissue and serum HYP60 dlater (t=3.65-4.40 and t =6.56-3.75,P＜0.05),all of them were lower than those in single radiation groups.There were significant positive correlations between tissue TGF-β1 and PAI-1 as well as C Ⅲ (Protein expression:r =0.604,0.759,P ＜0.05;Gene expression:r=0.519,0.816,P＜0.05).Conclusions ARPD may inhibit the pulmonary fibrosis by decreasing the levels of TGF-β1,PAI-1 and C Ⅲ.

BACKGROUND: Simple nanotube surface modification of titanium implant has been shown to promote adhesion, proliferation and differentiation of osteoblasts. Collagen coating can promote osteoblast adhesion and osseointegration in vivo. OBJECTⅠVE: To observe the effects of type collagen combined titanium dioxide nanotube composite coating modified titanium surface on osteoblast adhesion in vitro and osseointegration in vivo. METHODS: The titanium dioxide nanotube was fabricated on the pure titanium surface, then type Ⅰ collagen was combined with the nanotube structure to form composite coating. Scanning electron microscope observation was used to characterize the surface topography of the pure titanium, titanium dioxide nanotube and type Ⅰ collagen combined titanium dioxide nanotube surfaces. Contact angle test was employed to evaluate the hydrophilicity of different samples. MC3 T3-E1 murine preosteoblasts were seeded on the three kinds of materials for 4 hours. Cell adhesion morphology was examined by scanning electron microscope. Adherent cell counting was detected under inverted fluorescence microscope. Expression of actin cytoskeleton and vinculin was observed under laser scanning confocal microscope. The gene expression of vinculin and osteoprotegerin mRNA was detected by real-time quantitative PCR. The three kinds of samples were implanted into the tibia of Sprague-Dawley rats (provided by Laboratory Animal Center, Ⅰnstitute of Radiation Medicine, Chinese Academy of Medical Sciences) , and tibia samples were removed after 4 weeks of implantation for biological push-out test and histological observation. RESULTS AND CONCLUSⅠON: (1) Scanning electron microscope: There was mechanical scratch on the pure titanium surface. There was controllable, and uniform vertical arrangement of nanotubular structures with a diameter of approximately 70 nm on the titanium dioxide nanotube surface, and collagen adhered surrounding the nanotubular structures on the type Ⅰ collagen combined titanium dioxide nanotube substrate, and partial tubule orifices were closed. (2) The hydrophicility of type Ⅰ collagen combined titanium dioxide nanotube was significantly larger than those of the other two materials (P < 0.05) . (3) Compared with the pure titanium and titanium dioxide nanotube surfaces, the type Ⅰ collagen combined titanium dioxide nanotube substrate displayed increased adherent cell number, much well-organized cytoskeleton, enhanced immunofluorescence intensity of vinculin protein staining, and increased expression levels of vinculin and osteoprotegerin mRNA levels (all P < 0.05) . (4) Ⅰn vivo test revealed that the maximum push-out force in the type Ⅰ collagen combined titanium dioxide nanotube group was significantly higher than that in the pure titanium and titanium dioxide nanotube groups (P < 0.05) . Hematoxylin-eosin staining results showed that there were few bones, but many fibrous connective tissue surrounding the implant in the pure titanium group; there were more newly-born bones, and less fibrous connective tissue surrounding the implant in the titanium dioxide nanotube group; there were dense newly-born bones, and few thin fibrous connective tissue surrounding the implant in the type Ⅰ collagen combined titanium dioxide nanotube group. (5) These results indicate that type Ⅰ collagen combined titanium dioxide nanotube surface can facilitate osteoblast cell adhesion and promote osseointegration in vivo.

Objective To assess the influence between managements in emergency room(ER) andoutcome of severe traumatic brain injury (TBI),in order to provide inference for treatment.Methods A retrospective analysis was performed in severe TBI patients and recorded next indexes.(1) The managements in ER,including endotracheal intubation and oxygenation,fluid resuscitation,and mannitol intake.(2) The vital signs arriving at ICU,including systolic pressure and blood oxygen saturation.(3) Prognostic indicators including inhospital mortality and days during ICU,the scores of Glasgow outcome scale (GOS) at discharge and 6 months after injury.Results In 140 severe TBI patients,65 patients (46.4％) died during ICU.The mortality of patients with endotracheal intubation [65.0％ (39/60)] was significantly higher than that without endotracheal intubation [32.5％(26/80)](P＜ 0.01).The mortality in whether fluid resuscitation and using mannitol had no significant difference [44.7％ (46/103) vs.51.4％ (19/37),49.2％ (31/63) vs.44.2％ (34/77)] (P ＞0.05).In days during ICU,there was no significant difference among the three treatment measures (P＞ 0.05).In GOS grade at discharge and 6 months after injury,the proportion of 4 and 5 grade were 8.3％ (5/60) and 25.0％ (15/60) in patients with endotracheal intubation,while 27.5％ (22/80) and 52.5％ (42/80) in patients without endotraeheal intubation (P ＜ 0.01).In fluid resuscitation and using mannitol patients,there were no significant difference(P ＞ 0.05).Conclusion Treating severe TBI patients in ER,endotracheal intubation should be carefully chosen,fluid resuscitation and mannitol may not be given.

Objective:To investigate the effect of catheterin-related antimicrobial peptides(CRAMP)on the damage of cardiac microvascular endothelial cells induced by high glucose.Methods:Adult mouse heart microvascular endothelial cells were isolated and cultured.A model of microvascular endothelial cell injury was established by high glucose culture.The endothelial cells were randomly divided into 4 groups as the following.In the control group, 27.5 mmol/L mannitol was given as isoosmotic control as compared with the high glucose group.In the high glucose group(HG group), cells were cultured with 33.3 mmol/L high glucose for 48 h, and then treated without CRAMP.In 0.15 mg/L CRAMP treatment group, cells were cultured with 33.3 mmol/L high glucose for 48 h, followed by 0.15 mg/L CRAMP treatmen for 24 h. In the 0.5 mg/L CRAMP treatment group, cells were cultured with 33.3 mmol/L high glucose treatment for 48 h, and then treated with 0.5 mg/L CRAMP for 24 h. Cell proliferation was examined by staining with CKK-8 cell counting kit.The secretion of inflammatory factors in microvascular endothelial cells was detected by ELISA kit.Reactive oxygen species assay kit detects the level of reactive oxygen species in cells.Cell apoptosis was detected by apoptosis kit.Tubule formation and tubule number were measured by cells cultured on the matrix glue membrane, then detected by microscopic observation.The nitric oxide(NO)test kit measures levels of NO.The expression of nitric oxide synthase(eNOS)was detected by western blotting.Results:The cell proliferation activity was significant lower in the HG group than in control group[(52.2±5.4)% vs.(100.0±7.3)%]. The cell proliferation activity was higher in the 0.15 and 0.5 mg/L CRAMP groups than in the HG group[(72.0±3.4)% vs.(52.2±5.4)%; and(84.2±5.8)% vs.(52.2±5.4)%( F=75.300, P<0.001)]. The expression of tumor necrosis factor-α was significantly higher in the HG group than in the control group and in 0.5 mg/L CRAMP group[HG group of(239.1±32.1)μg/L, the control of(22.1±3.7)μg/L, 0.5 mg/L CRAMP of(84.6±9.4)μg/L]( F=197.300, P<0.001). The level of reactive oxygen species was significantly higher in the HG group than in control group and in 0.5 mg/L CRAMP group[(20.8±2.4)in HG group, (4.8±1.7)in control group, (10.2±1.5)in CRAMP group]( F=105.700, P<0.001). The number of apoptotic cells was significantly higher in the HG group than in control group and in 0.5 mg/L CRAMP group[(21.2±3.1)% in HG group, (2.2±0.6)% in control group(9.5±1.2)% in CRAMP group]( F=141.900, P<0.001). The length and number of tubules were lower in the HG group than in control group and in CRAMP group[for the length: (87.8±9.1)μm in HG group, (337.0±37.2)μm in control group(206.5±16.3)μm in CRAMP group( F=160.800, P<0.001); for the number: (9.1±1.9)in HG group, (22.0±3.4)in control group, (16.8±2.2)]in CRAMP group( F=36.200, P<0.001)]. The level of NO was lower in the HG group than in control group and in CRAMP group[(0.25±0.05)in HG group, (1.05±0.16)in control group, (0.75±0.06)in CRAMP group( F=83.200, P<0.001)]. The protein expression and mRNA levels of endothelial nitric oxide synthase(eNOS)were lower in the HG group than in the control group and in CRAMP group[for eNOS protein: (0.07±0.03)in HG group, (0.81±0.05)in control group, (0.54±0.07)in CRAMP group, F=275.700, P<0.001; and for eNOS mRNA: (0.11±0.07)in HG group, (1.00±0.22)in control group, (0.57±0.12)in CRAMP group, F=50.600, P<0.001]. Conclusions:CRAMP protein can inhibit the damage of cardiac microvascular endothelial cells by increasing eNOS-mediated NO signal pathway.

Objective:To determine the expression of circular RNA-SEC31A(circSEC31A) in pancreatic cancer and investigate the effects on the invasion and migration of pancreatic cancer cells and the underlying molecular mechanism.Methods:Differentially expressed circRNAs between pancreatic cancer cells (BXPC-3, PANC1, CaPan-2, SW1990) and human normal pancreatic cells (HPDE) were identified by qRT-PCR. Then, two cell lines with high circSEC31A expression were selected to conduct next experiments. According to the sequence of the back-splicing site in circSEC31A, siRNAs for downregulation of circSEC31A were designed and transfected by liposome to silence circSEC31A in pancreatic cancer cells, and grouped as followed siR-circSEC31A#1 and siR-circSEC31A#2. Meanwhile, siR-NC group transfected with non-specific siRNA served as control. Transwell assays and wound healing assays were operated to evaluate the functional role of circSEC31A on the invasion and migration of pancreatic cancer cells. RNA Pull-down assay with circSEC31A probe and oligo control probe was used to screen the miRNA combining with circSEC31A and the effects of miRNA on cell invasion and migration of pancreatic cancer cells were validated. The effect of miR-200c-3p and circSEC31A silencing on the expression of PDK1 mRNA was identified by qRT-PCR. The protein expression of PDK1, downstream Akt and p-Akt after circSEC31A silencing was verified by Western blotting assays.Results:The relative expression level of circSEC31A in HPDE (1.000±0.120) was obviously lower than that in BXPC-3 (1.920±0.130), SW1990 (2.93±0.528), PANC1 (4.557±0.692) and CaPan-2 (5.247±0.194), and all the differences were statistically significant ( P<0.001). Compared with the PANC1 siR-NC group (1301.3±94.6) and CaPan-2 siR-NC group (1835.0±70.1) per 100 high power field, transwell assays showed that the numbers of invasive pancreatic cancer cells was highly decreased in PANC1 siR-circSEC31A#1 group (727.3±92.9), siR-circSEC31A#2 group (792.0±18.1), CaPan-2 siR-circSEC31A#1 group (718.0±90.6), siR-circSEC31A#2 group (692.7±84.8). Wound healing assays showed that silencing circSEC31A decreased the wound healing rate of pancreatic cancer cells in PANC1 siR-circSEC31A#1 group (20.667±3.215)%, siR-circSEC31A#2 group (20.000±4.583)%, CaPan-2 siR-circSEC31A#1 group (28.000±8.185)%, siR-circSEC31A#2 group (29.667±5.686)%, compared with the PANC1 siR-NC group (55.000±4.359)% and CaPan-2 siR-NC group (69.000±3.606)%. RNA Pull-down assays showed that compared with PANC1 oligo probe group (1.000±0.091) and CaPan-2 oligo probe group (1.000±0.153), miR-200c-3p was significantly enriched in the PANC1 circSEC31A probe group (2.237±0.175) and CaPan-2 circSEC31A probe group (2.166±0.156). Compared with PANC1 siR-NC group (939.3±57.0) and CaPan-2 siR-NC group (786.7±51.5) per 100 high power field, the numbers of invasive pancreatic cancer cells were up-regulated in PANC1 siR-miR-200c-3p group (1206.0±99.1) and CaPan-2 siR-miR-200c-3p group (1838.0±105.7), while the low numbers of invasive pancreatic cancer cells were observed in PANC1 siR-miR-200c-3p+ siR-circSEC31A group (932.7±116.4) and CaPan-2 siR-miR-200c-3p+ siR-circSEC31A group (785.3±58.8). Compared with PANC1 siR-NC group (1.000±0.103) and CaPan-2 siR-NC group (1.000±0.107), the relative expression of PDK1 mRNA in PANC1 siR-miR-200c-3p group (1.898±0.159) and CaPan-2 siR-miR-200c-3p group (2.102±0.337) was upregulated. Furthermore, the expression of PDK1 mRNA was decreased in the siR-miR-200c-3p+ siR-circSEC31A group (0.980±0.070, 1.015±0.079). Western blot assays showed that the expression of PDK1 protein in PANC1 siR-NC group, siR-circSEC31A#1 group, siR-circSEC31A#2 group was 0.767±0.086, 0.281±0.191, 0.333±0.062 and in CaPan-2 siR-NC group, siR-circSEC31A#1 group, siR-circSEC31A#2 group was 0.712±0.038, 0.353±0.061, 0.308±0.018. The expression of p-Akt protein in PANC1 siR-NC group and siR-circSEC31A group was 0.741±0.050, 0.114±0.027, 0.139±0.041. In addition, p-Akt protein expression in CaPan-2 siR-NC group and siR-circSEC31A group was 0.823±0.052, 0.141±0.045, 0.280±0.089. PDK1 and p Akt expression in siR circSEC31A group was obviously lower than those in sir NC group. All the differences between either groups above were statistically significant ( P<0.05). Conclusions:circSEC31A is upregulated in pancreatic cancer cells, which facilitates the invasion and metastasis of pancreatic cancer cells via miR-200c-3p/PDK1/Akt signaling pathway, supporting that circSEC31A may function as a new diagnostic and therapeutic target for pancreatic cancer patients.

OBJECTIVETo investigate the relationship between radiographic parameters and the 4th lumbar(L4) degenerative spondylolisthesis.

METHODSFrom April 2010 to April 2012, 60 patients with the L 4 degenerative spondylolisthesis (DLS) were enrolled in DLS group, 56 healthy volunteers were recruited in control group. A series of radiographic parameters were measured in the two groups, including disc height (DH), disc degeneration index(DDI), L4 vertebral inclination angle(L4-VA), pelvic incidence (PI), L4 vertebral size (L4-VS), lumbar lordosis angle (LLA), facet joint angulation (FJA) of cephalad and caudad portions, delta FJA of cephlad and caudad portions, asymmetry variation of FJA, bone mineral density(BMD). Student's test was used to compare difference of parameters between two groups. Multivariate logistic regression analysis was used to reveal risk factors of the development of DLS.

RESULTSFifty-three cases of L4 spondylolisthesis in DLS group were classified into grade I, 7 cases of L4 spondylolisthesis were classified into grade II. The average Boxall index was 0.17 ± 0.05. There were significant difference of DH, DDI, L4-VS, L4-VA, LLA, PI, FJA, BMD between DLS group and control group (t = 2.28-9.33, P = 0.021-0.043) . There were significant differences of delta FJA of cephlad and caudad portions in L3-4, L4-5 between DLS group and control group (t = 3.398 and 28.122, P = 0.000 and 0.039). There was no significant difference of asymmetry variation of FJA in L3-4, L4-5 between DLS group and control group (t = 0.209-0.465, P = 0.295-0.858). Multivariate logistic regression analysis showed that LDS was more frequent among patients with smaller L4-VS(OR = 1.01, 95%CI = 1.000-1.024, P = 0.048), larger L4-VA (OR = 1.88, 95%CI = 14.000-14.600, P = 0.037), larger LLA (OR = 1.90, 95%CI = 1.600-15.800, P = 0.040), larger PI (OR = 2.58, 95%CI = 18.000-19.600, P = 0.029) and the more sagittal FJA (OR = 2.46, 95%CI = 1.400-16.400, P = 0.035) than those in control group.

CONCLUSIONSDLS is signifantly correlated with L4-VS, L4-VA, LLA, PI, FJA . They may be risk factors of the development of DLS.

Aged ; Bone Density ; Case-Control Studies ; Female ; Humans ; Intervertebral Disc Degeneration ; diagnosis ; Lumbar Vertebrae ; diagnostic imaging ; Male ; Middle Aged ; Radiography ; Risk Factors ; Spondylolisthesis ; diagnosis