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2016 Vol. 46, No. 4 Published: 2016-04-25   235 Fusion expression and immune investigation of recombinant dengue virus EDIII tetravalent protein#br# REN Shoufeng, LIU Wenquan, CAO Guomei, LIANG Shaohui, PAN Changwang Objective: To construct dengue virus envelope protein EDIII tetravalent recombinant vaccines and to analyze its immunogenicity. Methods: Firstly, the gene sequences encoding dengue virus type 1-4 envelope protein EDIII were analyzed by bioinformatics methods, and the representative DENV1-4 EDIII was selected and connected by GS linker to obtain chimeric EDIII; Then, the chimeric gene coding EDIII was inserted into the multiple cloning site of the expression vector pColdIII to constructe the recombinant vector pColdIII-EDIII. The constructed pColdIII-EDIII plasmid was transformed into E.coli BL21 cells and induced by IPTG. The recombinant protein EDIII was purified by Ni-NTA affinity chromatography and confirmed by SDS-PAGE and Western blot analysis. The purified protein was used to immune BABL/c mice. After boosting twice, to detect the antibody titers in serum and the secretion of cytokines by ELISA to evaluate the immune response. Results: The mice immunized with EDIII could induce specific antibodies and the antibody titers was 1:40 000. The mouse spleen lymphocytes stimulated by antigen could produce cytokine including IFN-r, IL-4, IL-10, which were significantly higher in the experimental group. Conclusion: Our studies indicates that dengue virus envelope protein EDIII tetravalent vaccine can induce specific humoral and cellular immune response, and give priority to with the humoral immune response, which laid the foundation for the research of a new type of chimeric tetravalent vaccine in the future. 2016 Vol. 46 (4): 235-240,244 [Abstract] ( 692 ) HTML (1 KB)  PDF (3209 KB)  ( 1114 ) 241 Correlation between prothrombin gene polymorphism of rs5896 C>T and kidney stone JI Huijuan, CHEN Tao, JIANG Minghua, WANG Dawen, WANG Yijun, HU Yunliang. Objective: To investigate the correlation between prothrombin rs5896 C>T genetic polymorphism and kidney stone in the southern population of Zhejiang. Methods: Prothrombin gene polymorphism rs5896 was investigated in 102 patients with renal stone formation and 95 age-matched healthy volunteers without history of renal stone formation, using polymerase chain reaction-restriction fragment polymorphism (PCR-RFLP) and sequencing. The frequences of genetype and allele gene in the case and control were analyzed. Results: The frequencies of CC, CT, TT were 13.7%, 50.9%, 35.4% and 24.2%, 51.6%, 24.2% in kidney stone patients and healthy controls respectively. The difference between case and control was slight significance (P=0.028). The frequencies of C and T were 39.2%, 60.8% and 50.0%, 50.0% in kidney stone patients and heathy control. There was a different significance (P=0.031). Conclusion: It suggests that the rs5896 polymorphism of prothrombin gene is associated with the kidney stone in the southern population of Zhejiang, and it is possible that rs5896 C>T acts as a risk factor for the development of renal stone disease. 2016 Vol. 46 (4): 241-244 [Abstract] ( 625 ) HTML (1 KB)  PDF (1515 KB)  ( 948 ) 245 Clinical study of recombinant human endostatin combined with hepatic arterial chemoembolization in advanced primary hepatic carcinoma ZHAO Jianguo, WANG Ting, ZHAO Zhongwei, YE Wanli, WU Dongping, JI Jiansong, WANG Jianfang Objective: To investigate the clinical efficacy and adverse reactions of recombinant human endostatin (Endostar) combined with transcatheter arterial chemoembolization (TACE) in advanced hepatocellular carcinoma (aHCC) and provide the reference treatment for aHCC. Methods: Sixty-two patients with advanced hepatocellular carcinoma were enrolled  from January 2011 to December 2014 in our hospital which were divided randomly into observation group (32 cases) and control group (30 cases) according to the therapeutic methods, the patients of control group were treated with TACE, while the observation group was treated with TACE combined with hepatic dynamic arterial infusion of Endostar. The clinical efficacy and adverse reactions were observed and evaluated between two groups after treatment. Results: The effective rate of observation group was 75%, which was significantly higher than that of the control group with 46.67% (P<0.05), the PFS of observation group was significantly longer than the control group (P<0.05), the survival time was not significantly different (P>0.05) in two groups. No significant difference was found in the incidence of adverse reactions between two groups (P>0.05). Conclusion: Endostar combined with TACE can effectively improve the clinical efficacy and prolong the PFS without more adverse reactions in aHCC, which has important clinical value. 2016 Vol. 46 (4): 245-248 [Abstract] ( 683 ) HTML (1 KB)  PDF (1326 KB)  ( 916 ) 249 Expressions inhibition of haploid sperm nucleoprotein genes in mice sub-chronically exposed to benzene via inhalation ZHANG Yaya, QIN Qiongyu, LIN Chong, SUN Shuqiang, ZHU Hua, YANG Xinjun, YANhongtao Objective: To explore the major damage stage of spermatogenesis in mice submitted to subchronic inhalation of benzene. Methods: C57BL/6J mice were randomly divided into control, 1 ppm and 100 ppm benzene exposure group. Simulated occupational exposure condition, a subchronic inhalation exposure test was carried out using the dynamic control equipment. The pathological change of testicular tissue was observed with HE staining. The mRNA expression of marker genes of spermatogenesis was quantitatively tested with real-time polymerase chain reaction, the selected marker genes included Plzf of undifferentiated spermatogonia, Stra8 of differentiated spermatogonia, Sycp3 of primary spermatocyte, Tnp1 and Tnp2 of round spermatid, Akap4 of elongating spermatid, as well as Prm1 and Prm2. Results: Enhanced eosinophilic staining were observed in mice testicular tissue section of benzene exposure groups, a few karyolysis and exfoliative cells were found in 1 ppm exposure group, the phenomenon of mice testis germ cell degeneration and necrosis were more obvious in 100 ppm exposure group, the number of seminiferous tubules with no or few germ cells were increased obviously. It suggested that subchronic benzene exposure could induce pathological change in mice testis. The mRNA expression of Plzf, Stra8, Sycp3, Akap4 did not show statistical difference in all groups, Prm2 expression was significantly reduced in 1 ppm group than that of control group (P<0.01). The mRNA expression of transition protein Tnp1 and Tnp2, protamine Prm1 and Prm2 were significantly reduced in 100 ppm exposure group than that of the control and 1 ppm group. Conclusion: Subchronic benzene exposure can induce the pathological change in mice testis, down-regulate the mRNA expression of Tnps and Prms, mainly influence the process of histone replaced by protamine in round spermatid nuclei. 2016 Vol. 46 (4): 249-253 [Abstract] ( 632 ) HTML (1 KB)  PDF (1651 KB)  ( 850 ) 254 Expression of miR-21 in patients with the acute phase of Kawasaki disease and its clinical significance JIA Lianhong, PAN Lulu, LU Jiacheng, GENG Zhimin, WU Rongzhou, CHU Maoping. Objective: To study the expression of miR-21 in the serum of patients with Kawasaki disease and its clinical significance. Methods: Thirty-three cases of Kawasaki disease and 15 healthy control were collected from the Second Affiliated Hospital & Yuying Children’s Hospital of Wenzhou Medical University in 2014 and 2015. With cel-miR-39 as the external reference gene, the expression of miR-21 was detected with real-time quantitative PCR, and its significance as a marker in diagnosis was evaluated by ROC curve. Results: The expression of miR-21 in serum of KD patients was significantly higher than that in the healthy control group (P<0.05), and return to normal level after treatment. ROC curve indicated miR-21 had a sensitivity of 84.6% and a specificity of 71.4% for distinguishing KD patients from healthy children with an area under the curve (AUC) were 0.736 (95%CI: 0.531～0.941). Further analysis of the clinical pathological features indicated that the level of serum miR-21was associated with PLT. Conclusion: Serum miR-21 level is significantly elevated in KD patients, it may be used as a potential biomarker in diagnosis of KD. 2016 Vol. 46 (4): 254-257 [Abstract] ( 1003 ) HTML (1 KB)  PDF (1327 KB)  ( 997 ) 258 Effects of CYP3A4*1G genetic polymorphism on analgesia with sufentanil in lower abdominal surgery LI Li, LIN Zheng, XU Jianwei, QIAN Xiaoweri, LI Jun. Objective: To investigate the effects of CYP3A4*1G genetic polymorphism on analgesia with sufentanil in lower abdominal surgery. Methods: One hundred and twenty patients with ASA I or II, aged 20-65 years who underwent elective lower abdominal surgery under general anesthesia were recruited into this study. Patient-controlled analgesia (PCA) treatment was given after operation. Genotyping of CYP3A4*1G was carried out by pyrosequencing. The patients were assigned into 3 groups according to their genotypes: group I wild homozygote, group II mutation heterozygote and group III mutation homozygote. MAP and HR were monitored before induction of general anesthesia (T0), after intubation 1 min (T1), at skin incision (T2) and extubation (T3), and at 5 min after extubation (T4). Plasma cortisol (Cor) and angiotension II (Ang-II) were measured as well. PCA sufentanil consumption and adverse effects were recorded during the first 24 h after surgery. Results: ①MAP, HR, Cor and Ang-II at T1, T2 and T3 were lower in group I than those in group II and III (P<0.05). ②No significant differences in the scores of VAS were noted between the three groups (P>0.05). While similar degrees of pain control was achieved, patients in the *1G/*1G group (49.8±10.2) μg consumed significantly less sufentanil than that in either the wild-type group (64.6±10.9) μg or the *1/*1G group (62.5±12.7) μg (P<0.01). But there was no significant difference in this index between group II and III (P>0.05). Conclusion: CYP3A4*1G genetic polymorphism is one of the factors contributing to the individual variation in patient’s response to analgesia with sufentanil. 2016 Vol. 46 (4): 258-262 [Abstract] ( 687 ) HTML (1 KB)  PDF (1281 KB)  ( 743 ) 263 Relationship between XIAP expression and radiotherapy prognosis in nasopharyngeal carcinoma XIANG Zhenfei, FANG Jianchen, LIN Li, LU Miaozhen, XU Quan Objective: To investigate the relationship between X-linked inhibitor of apoptosis protein (XIAP) expression and prognosis of patients with nasopharyngeal cancer (NPC) that received radiotherapy. Methods: A retrospective analysis was performed with the data of 94 NPC patients from January 2010 to December 2012. The expression of XIAP in NPC tissues was detected with immunohistochemistry (IHC), and analyzed the prognosis relevance after radiotherapy. Results: The overall XIAP positive expression rate in NPC tissues was 69.15% (65/94) and the XIAP positive expression rate was negatively correlated with the p-TNM staging and the tumor differentiation (all P<0.05). Kaplan-Meier analysis showed that the high XIAP expression, high p-TNM staging and poorly differentiated were significantly correlated with decreased survival rate of NPC patients (all P<0.05). Multivariate Cox regression analysis showed that XIAP expression, p-TNM staging and degrees of differentiation were independent risk factors for the survival of NPC patients after radiotherapy. Conclusion: The high XIAP expression, p-TNM staging and degree of differentiation are independent risk factors for the survival of NPC patients after radiotherapy. 2016 Vol. 46 (4): 263-266,273 [Abstract] ( 830 ) HTML (1 KB)  PDF (2444 KB)  ( 838 ) 267 The effect of EGB on expression of TGFβ1, NF-κB, and FN in hepatic tissues from diabetic rats with non-alcoholic fatty liver disease WANG Jiayuan, FAN Jianshuang, WU Liang, HUANG Kate, WANG Rongrong Objective: To observe the effect of extract of ginkgo biloba (EGB) on hepatic tissue from DM rats with non-alcoholic fatty liver disease (NAFLD). Methods: Thirty male Sprague-Dauley rats were divided randomly into three groups: normal control group, type 2 diabetic group and EGB-treated group. After being fed with high-fat feeding for 4 weeks, the later groups were injected with low dosage strepozotocin (30 mg/kg) intraperitoneally to induce NAFLD model of type 2 diabetes mellitus. The EGB treated group was gavaged with EGB at the dosage of 50 mg·kg-1·d-1 for 12 weeks. The pathologic changes of liver was observed under light microscopy (LM) stained with HE and VG staining respectively. Additionally, the protein expression of NF-κB, TGFβ1, FN, α-SMA were assayed by immunohistochemistry (IHC), the mRNA expression of TGFβ1, NF-κB, FN from rats livers were assayed by semi-quantitative RT-PCR. Results: Liver specimen from diabetic rats stained by HE and VG showed obvious liver fatty degeneration, ballooning degeneration, dotty necrosis necrosis with accompanying inflammatory infiltration and fibrosis. Meanwhile, we discovered that there were a great number of α-SMA reactive hepatic stellate cells in diabetic rats liver tissue. In addition, the protein expression of NF-κB, TGFβ1, FN (P<0.05, P<0.01, P<0.05) and the TGFβ1, FN mRNA expression (P<0.01, P<0.05) of diabetic rats livers were significantly higher than that of the normal control group. After treatment with EGB, the pathological change of liver and proliferation of hepatic stellate cell in diatetic rats relieved (P<0.05).Meanwhile, the protein expression of NF-κB, TGFβ1, FN (P<0.05) and the TGFβ1, FN mRNA expression (P<0.05) of diabetic rats livers decreased significantly compared with type 2 diabetic group. Conclusion: EGB can relieve NAFLD from diabetic rats. Down regulation of expression of NF-κB, TGFβ1, FN may be involved in it 2016 Vol. 46 (4): 267-273 [Abstract] ( 585 ) HTML (1 KB)  PDF (2457 KB)  ( 934 ) 274 Posterior individual fixation, combined with anterior mini-open debridement and strut graft for the treatment of lumbar spinal tuberculosis YING Xiaozhang, ZHENG Qi, SHI Shiyuan, SHEN Jian, WANG Yifan, ZHU Bo, HAN Guihe, ZHENG Mingfeng. Objective: To determine the efficacy of posterior individual fixation combined with anterior mini-open debridement and strut graft for the treatment of lumbar spinal tuberculosis. Methods: From February 2011 to May 2014, a total of 62 cases with lumbar spinal tuberculosis were treated with posterior individual fixation combined with anterior debridement and strut graft. The cohort comprised 33 males and 29 females with an average age of 51.3 years. The pre-operative Frankel grading showed that 2 cases were grade B, 8 grade C, 16 grade D, and 36 grade E. Anti-tuberculosis therapy was carried out for two to four weeks, and the operation plan was determined by symptoms, signs and image. The clinical efficacy was evaluated based on the kyphosis Cobb angle, the visual analogue scale (VAS), erythrocyte sedimentation rate (ESR) and the Frankel grade system before and after the surgery. Results: The average follow-up duration was 23.7 months (15-36 months). There were three patients presented with psoas abscess after three months on the contralateral side, and cured by drainage. At the final follow-up the Frankel grading showed that 1 was grade C, 5 were grade D, and 56 were grade E. The Cobb’s angle was decreased from (16.34°±3.19°) (pre-operation) to (4.52°±1.69°). The visual analogue scale (VAS) was decreased from (6.72±1.02) (pre-operation) to (2.06±0.73). The erythrocyte sedimentation rate (ESR) was decreased from (40.12±1.53) mm/1 h (pre-operation)to (10.67±0.72) mm/1 h. Graft union was observed 3 to 7 months after surgery, with a mean of 4.6 months. There was no vertebral tuberculous relapse or screw loose/abruption. Conclusion: It is effective to treat lumbar tuberculous with posterior individual fixation combined with anterior mini-open debridement and strut graft. 2016 Vol. 46 (4): 274-277,283 [Abstract] ( 583 ) HTML (1 KB)  PDF (2603 KB)  ( 857 ) 278 Expression of miR-187 in the tissues of lung cancer and its effect SUN Debin, LAN Xiu Objective: To explore the expression of miR-187 in the tissues of lung cancer and its effect.  Methods: By normalized to U6, the expressions of miR-187 in 32 lung cancer and matched non-tumor adjacent tissue specimens were examined by stem-loop real-time RT-qPCR method. The relationship between miR-187 expression and clinicopathological characteristics was further analyzed. After transfected with miR-187 mimics, the biological functions of miR-187 were determined by cell proliferation and cell cycle assay. Results: Expression of miR-187 in the tissue of lung cancer was obviously higher than that in non-tumor adjacent tissue specimens (t=5.236, P<0.0001). Clinical features correlation analysis showed that the down-expression of miR-187 was correlated with the pathological grading and the clinical staging (P<0.05). Functional studies indicated that the overexpression of miR-187 dramatically inhibited the proliferation of A549 cells in vitro and changed the situation of the cell cycle, arrested at G0/G1 phase. Conclusion: Down-expression of miR-187 in the patients of lung cancer may play a key role in the development and progression of lung cancer. 2016 Vol. 46 (4): 278-283 [Abstract] ( 656 ) HTML (1 KB)  PDF (1675 KB)  ( 981 ) 284 Expression and significance of ING4 in human gastric cancer CHENG Haiyan, SHEN Zhe, CHEN Weichang, YAN Su. Objective: To study the expression and significance of inhibitor of growth family member 4 (ING4) in many kinds of human gastric cancer cells and gastic cancer tissue. Methods: In the experimental group, there were three different typs of gastric cancer cells, MKN-1 (high differentiation), SGC-7901 (moderate differentiation)and BGC-823 (low differentiation). In the control group, there was a normal gastric mucosal cells GES-1. All cell lines were cultured in vitro. The mRNA and protein expression of ING4 were analyzed by RT-PCR and Western blot. Whether there was ING4 protein expression in gastric cancer tissue and adjacent tissue by immunohistochemistry. Results: The mRNA and protein expression of ING4 in SGC-7901 and BGC-823 groups were significantly lower than that in MKN-1 and GES-1 groups. The difference was statistically significant (P<0.05). The mRNA and protein expression of ING4 in BGC-823 group were significantly lower than that in SGC-7901 group. The difference was statistically significant (P<0.05). But there was no significant different between MKN-1 group and GES-1 group (P>0.05). The positive rates of expression of ING4 protein in gastric cancer tissue was significantly lower than that in adjacent tissue, the difference was statistically significant (P<0.05). Conclusion: ING4 protein as a tumor suppressor is associated with the development and prognosis of the gastric cancer, which provide theoretical basis for clinical use ING4 gene targeted therapy for gastric cancer in the future. 2016 Vol. 46 (4): 284-286 [Abstract] ( 631 ) HTML (1 KB)  PDF (1332 KB)  ( 756 ) 287 Effects of heterologous acellular dermal matrix combined with autologous skin grafts on the residual wounds with titanium mesh in skull defects LUO Xu, LIN Cai, WANG Xinling. Objective: To explore the therapeutic means of heterologous acellular dermal matrix (ADM) combined with autologous skin graft (ASG) to repair the residual skull defective wounds with titanium mesh exposed. Methods: Heterologous acellular dermal matrix (phase I) combined with autologous skin graft (phase II) transplantation operation was used in 10 patients of titanium mesh skull defect with exposed residual wound. The wound heal situation was observed. Results: ADM and ASG were all alive in 10 patients. Healing of type II incision on the wound was in grade I. The average operation duration was 30-45 min, while the average volume of blood loss was 30-50 mL. After follow-up observation for three months, skin elasticity at the surgical transplantation area was good without damage. Conclusion: The new therapeutic method for repairing the residual wound with exposure titanium mesh by means of ADM combined with ASG transplantation can avoid to some extent requiring more working time than other operations, such as regional perforating branch flap and free flap transplantation, and needn’t special equipment, adequate train and the complex preoperative preparation. 2016 Vol. 46 (4): 287-288 [Abstract] ( 637 ) HTML (1 KB)  PDF (1248 KB)  ( 740 ) 289 The application effect of piezosurgery in transcrestal maxillary sinus augmentation: a preliminary study YANG Ying, LIU Chuantong, JIN Qiong, LIU Guo Objective: To investigate the application effect of piezosurgery in transcrestal maxillary sinus augmentation (TMSA). Methods: Fifty-two patients need TMSA instomatology of Wenzhou Medical University were randomly divided into two groups: the experimental group and the control group. The piezosurgery was used in the experimental group for TMSA and the Summers’ technology was used in the control group for TMSA. The feeling of treatment was analyzed by visual analog scale (VAS), and then the differences in the perforation rate of maxillary sinus membrane and the height of lifting were analyzed with SPSS17.0 software. Results: A significant lower VAS score was found in the experimental group, compared with the control group (P<0.05). Meanwhile, the perforation rate of maxillary sinus membrane of experimental group was lower than that of the control group (P<0.05). However, the difference of the height of lifting in the two groups was not significant (P>0.05). Conclusion: The application of pizeosurgery is safe and reliable in TMSA. Compared with the Summers’ technology, the pizeosurgery can reduce discomfort feeling, and improve the treatment experience of patients. 2016 Vol. 46 (4): 289-291 [Abstract] ( 590 ) HTML (1 KB)  PDF (1232 KB)  ( 831 ) 292 The value of procalcitonin in guiding antibiotic therapy for acute exacerbation of chronic  obstructive pulmonary disease WU Tianxian, JIN Xuewen, ZHANG Yehui, HUANG Juhong, ZHENG Yulin. Objective: To investigate the value of procalcitonin (PCT) in guiding antibiotic therapy for acute exacerbation of chronic obstructive pulmonary disease (AECOPD). Methods: Totally 134 patients with AECOPD were randomly divided into PCT group (47 cases), Guide group (42 cases) and Experience group (45 cases). Three groups of clinical efficacy, the total medical costs, costs of antibiotic therapy and hospitalization days were statistically analysed. Results: Between the PCT group and the guide group in the total medical costs, costs of antibiotics and hospitalization days had no statistically significant difference (P>0.05). PCT group and guide group in the total medical costs, costs of antibiotics and hospitalization days were statistically significant difference compared with the experience group (P<0.05). There was no significant diference in cases of clinical efficacy among the three groups (P>0.05). Conclusion: PCT detection can effectively reduce the total costs of hospitalization, which can also reduce costs of antibiotic therapy and the duration of hospital stay in the treatment of AECOPD patients. 2016 Vol. 46 (4): 292-295 [Abstract] ( 680 ) HTML (1 KB)  PDF (1304 KB)  ( 1096 ) 296 Clinical and pathological analysis of 311 cases of periocular neoplasms LU Sijia Objective: To investigate the common clinical and pathological classifications and the distribution of periocular neoplasms. Methods: A total of 311 periocular neoplasms verified by the department of pathology at the third people’s hospital of Hangzhou from 2008 to 2015 were included in this study. Results: There were 139 male (44.69%) and 172 female (55.31%) in the 311 periocular neoplasms. The age distribution was 1 to 87 years old. Two hundred and fifty-seven (82.64%) of the lesions were benign, while 54 (17.46%) of the lesions were malignant or having malignant tendency. In 115 patients over 60 years old, 43 (37.39%) of them were malignant or having malignant tendency. The three leading benign lesions were verruca vulgaris, intradermal nevus and seborrheic keratosis; The three leading malignant lesions or lesions having malignant tendency were basal cell carcinoma, actinic keratosis and squamous cell carcinoma, the mean ages of occurrence of them were 66.85, 79.33 and 68.83 years old, respectively. The malignant lesions or lesions having malignant tendency were most commonly found on the lower eyelid (29.63%), followed by medial canthus, arcus superciliaris, upper lid and lateral canthus. Conclusion: The periocular area is important and sophisticated, it is susceptible to neoplasms. Basal cell carcinoma is the most common periocular malignant tumor. Pathological examination should be performed for periocular neoplasms to guide the diagnosis and treatment in clinics. 2016 Vol. 46 (4): 296-299 [Abstract] ( 603 ) HTML (1 KB)  PDF (1267 KB)  ( 1153 ) 299 Effect of nutritional intervention on nonalcoholic fatty liver disease ZHANG Rui, GU Binbin Objective: To observe the improvement of nonalcoholic fatty liver disease (NAFLD) with nutritional intervention. Methods: Fifty-six patients with NAFLD were given diet nutritional intervention for 6 months. The changes of macronutrient intake, biochemical indexes and body composition indexes were recorded and analyzed before and after intervention. Results: Nutritional intervention led to lower intakes in daily total energy and the energy rate from carbohydrate (P<0.01) and higher intake in the energy rate from protein (P<0.01), but the energy rate from fat did not significantly change. Body mass index, waist hip ratio, body fat mass and visceral fat area were significantly decrease, while body protein and skeletal muscle mass no obviously changed after intervention. Fasting blood glucose, triglyceride, total cholesterol and low-density lipoprotein were significantly lower than the level before intervention (P<0.01), but high-density lipoprotein did not significantly change. Conclusion: Reasonable nutrition intervention can help to loss weight and visceral fat, improve serum glucose and serum lipids and thus exerts a favorable effect on nonalcoholic fatty liver disease. 2016 Vol. 46 (4): 299-302 [Abstract] ( 614 ) HTML (1 KB)  PDF (1271 KB)  ( 863 )

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