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2013, Volume 44 Issue 4
06 August 2013 Previous Issue    Next Issue

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Long-term alcohol exposure inducing insulin resistance and retinal microvascular damage, a possible role of sphingomyelin XU Gao-lei WANG Zhi-xin ZHAO Jing-ya CHEN Wen-jing CUI Zhan-jun WANG Yan-fen WANG Hui DENG Jin-bo* GAO Xiao-qun 2013, 44 (4 ):  441-450.  doi: 10.3969/j.issn.0529-1356.2013.04.001 Abstract ( )   Objective To investigate the mechanism of insulin resistance and retinal damages induced by long-term alcohol exposure, especially the mechanisms which SM (sphingomyelin, SM) regulationin is involved in. Methods Sphingomyelin synthase 2 knockout (SMS2-/- ) mice and wild type (WT) mice were used to establish the long term alcohol exposure models. The total cases were 76 in each group. Blood glucose and blood insulin were measured with ELISA assay, in both SMS2-/- mice and WT mice. In the meantime, homeostasis model assessment of insulin resistance (HOMA-IR) was calculated. Finally, the pathologic alterations about retinal microvasculature were investigated with immunofluorescent labeling, hematoxylin and eosin staining and electron microscopy. Results Long-term alcohol exposure induced insulin resistance with dose dependency (P <0.05), and blood retinal barrier (BRB) damages, such as acellular capillaries increased (P <0.05), astrocytes decreased (P <0.05) and damages of endothelial cell in ultrastrctural level, with dose dependency. In addition, either insulin resistance index increase or BRB damages in SMS2-/- mice were less than in WT mice (P <0.05), suggesting SMS2-/- mice might attenuate insulin resistance and BRB damages. Conclusion The insulin resistance (IR) and BRB damages can be induced by long-term alcohol exposure. As an oxidative stress factor, alcohol exposure may lead to the activation of PKC-NFκB-(Bcl-Xs) pathway and produce the effects of IR and BRB damages.  References | Related Articles | Metrics

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Gene knockout of amyloid precursor protein promotes aluminum-induced cognitive impairment in mice WANG Dong-mei YUAN Shu-min ZHANG Xu ZHANG Lian-feng 2013, 44 (4 ):  451-455.  doi: 10.3969/j.issn.0529-1356.2013.04.002 Abstract ( )   Objective Amyloid precursor protein (APP) is involved in dementia, however, little is known about its role in the development of dementia. The APP knockout mice were used to investigate the effects of amyloid precursor protein on aluminum-induced cognitive impairment in mice. Methods  APP knockout mice and the littermates of no-negative mice aged 3 months were selected randomly and performed Morris water-maze tests after administration for 8 weeks. Aluminum was administrated in the diet. One group of APP knockout mice was used for vehicle group. One group of the littermates of no-negative mice was used as normal control (WT). The pathological changes in brain were detected by HE staining. The activities of glycogen synthase kinase 3β(GSK-3β)and Caspase-3 were also examined by Western blotting.  Results Compared with WT mice, Al-treated WT mice exhibited a decrease in the target-quadrant abidance by 28.1% and the crossing-target number by 18.8% in the probe test; APP knockout mice administrated showed a significant decrease in the target-quadrant abidance by 44.1% and the crossing-target number by 51% in the probe test. Compared with that of WT mice, the level of p-GSK-3β was decreased by 17.4% in Al-treated WT mice and by 46.4% in Al-treated APP knockout mice. Conclusion Gene knockout of APP promotes aluminum-induced neurotoxicity and cognitive impairment in mice. APP knockout leads to a significant increase of the activity of GSK-3β, which can accelerate the processing of dementia. Thus the protective effect of APP may be through inhibiting the activity of GSK-3β. References | Related Articles | Metrics

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N-methyl-D-aspartate receptor 2A expression intensity in brain regions of adult male SD rats BI Wen-jie XIAO Li ZHENG Xiang* 2013, 44 (4 ):  456-462.  doi: 10.3969/j.issn.0529-1356.2013.04.003 Abstract ( )   Objective N-methyl-D-aspartate receptor (NMDAR) is an important excitatory receptor within the central nervous system. Its 2A subunit is a critical functional subunit. There is no comprehensive data about NMDAR 2A expression intensity in rat brain. Methods In this experiment, we detected NMDAR 2A expression in the brain of adult male SD rat (n=6) with immunohistochemistry staining and image analysis. Results We drew the expression intensity atlas of rat brain based on the immunohistochemical staining intensity. Conclusion These data can provide a reference for in situ detection of NMDAR 2A in related experiments. References | Related Articles | Metrics

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Neural protective effect of estrogen on the rat red nucleus by dissector quantitative analysis HAN Xiao JI Da-feng LI Yao-fu ZHANG Xia-nan QIN Jian-bing Lü Guang-ming* 2013, 44 (4 ):  463-467.  doi: 10.3969/j.issn.0529-1356.2013.04.004 Abstract ( )   Objective To investigate the protective effect of estrogen(E2) on red nucleus after spinal cord injury (SCI) inthe ovarectomized rat. Methods Eighty adult female rats were selected in this experiment and randomly divided into the normal group, rubrospinal tract injury group, estrogen replacement group, inhibitor of estrogen receptor (ICI) treatment group and E2+ICI cooperative treatment group. Sole rubrospinal tract injury model was established by cutting off the left dorsolateral funiculus at C3-C4 segment of rat spinal cord on one week after ovariectomy operation. The animal behavior was evaluated with forelimb probing experiment, and FR positive neurons were counted and analyzed on 1 week, 2 weeks and 4 weeks by using disecter after different treatments on animals. Results The forelimb probing test showed a higher use rate of left forelimb in the E2 replacement treatment group than that of other groups, but with no statistical significance (P>0.05). Except the normal group, the number of neurons in right red nucleus of rat midbrain decreased, especially on 4 weeks. The FR positive neurons of E2 replacement group had a more remarkable cell body, clearer outline and longer processes compared to the neurons in the RST injury group, ICI treatment group and E2+ICI cooperative treatment group. The stereological frame illustrated that more FR positive neurons survived in the E2 replacement group (P<0.05), but with no significant changes (P>0.05) among the RST injury group, ICI treatment group and E2+ICI cooperative treatment group. Conclusion The E2 replacement treatment may release red nucleus neuron retrograde injury and promote the neuron survival after rat SCI. References | Related Articles | Metrics

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Preparation and characterization of homeobox B9（HOXB9） polyclonal antibody NIU Miao-miao MA Bo TANG Yan LI Feng ZHAN Jun* ZHANG Hong-quan 2013, 44 (4 ):  468-474.  doi: 10.3969/j.issn.0529-1356.2013.04.005 Abstract ( )   Objective The purpose of this study was to prepare rabbit anti-homeobox B9(HOXB9) polyclonal antibody and to provide a tool for future study. Methods DNA fragments of human HOXB9 gene 5,terminal 7-458 base pairs was respectively cloned into PGEX-4T-1 vector expressing GST tag and pMal-C2x vector expressing MBP tag by polymerasechain reaction and recombinant DNA technology.The GST-HOXB9-N-terminal fusion protein and MBP-HOXB9-N-terminal fusion protein were expressed by E.coli BL21（DE3）after heat shock transformation and IPTG induction. The New Zealand rabbit was immunized with the purified GST-HOXB9-N-terminal fusion protein. We used ELISA assay to determine the titer of the antiserum.If the antiserum was effective, the rabbit blood was obtained from the carotid artery and immunoglobulinG(IgG)was purified from serum. Effectiveness and specificity of the antibody were identified by Western blotting and immunoprecipitation. Results The prepared antibody specifically recognized a single band of the target molecule, and had a strong capability of immunoprecipitation on the native HOXB9 protein.Using this antibody, we demonstrated that HOXB9 expressed highly in the mouse immune organs,pancreas,colon, lung, cerebellum, female reproductive system and testis,while expressed lowly in other organs. Conclusion We successfully prepared a rabbit anti-human HOXB9 polyclonal antibody. This polyclonal antibody is specific and effective, and may be used for Western blotting and immunoprecipitation analysis. References | Related Articles | Metrics

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Expression of acid-sensing ion channel 1a in endplate chondrocytes of the vertebral body in rats LI Xia ZHAO Yi-qing ZHOU Lu-yao HE Chun-hui Lü Wei-qun YUAN Feng-lai*  2013, 44 (4 ):  475-478.  doi: 10.3969/j.issn.0529-1356.2013.04.006 Abstract ( )   Objective To explore the expression of acid-sensing ion channels 1a(ASIC1a) in endplate chondrocytes of the vertebral body in rat. Methods Endplate chondrocytes were isolated from the 10 adult SD rats, and their phenotypes were determined by an inverted microscope and toluidine blue staining. RT-PCR and Western blotting were used to detect ASIC1a in endplate chondrocytes of the rat vertebral body. Fluorescence immunocytochemistry double marker was used to the location of ASIC1a in the cells. Results Positive staining for glycosaminoglycan was seen in endplate chondrocytes. RT-PCR analysis showed that ASIC1a mRNA were presentin endplate chondrocytes of the vertebral body. The total RNA, and the corresponding protein were detected. Conclusion The ASIC1a expresses in endplate chondrocytes of the vertebral body, which may be regarded as a new target of intervertebral disc degeneration (IVDD). References | Related Articles | Metrics

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Antibody diversity of hepatitis B immunoglobulin ZHOU Shi-quan ZHOU Ji-hang XU Yue-jun ZHUANG Xiao-ling LI Yi-wei LI Shi-bo LIU Xiao-guang* 2013, 44 (4 ):  479-484.  doi: 10.3969/j.issn.0529-1356.2013.04.007 Abstract ( )   Objective To classify IgG antibodies in hepatitis B immunoglobulin(HBIG) based on DNA-sequencing. Methods The total RNA was extracted as template from the peripheral blood of a healthy individual with high titer of HBsAb(equivalent to HBIG donor), and a two-step RT-PCR and Nested-PCR reaction was conducted sequentially with two sets of compatible primers, which obtained from a literature and specialized for amplifing all IgG heavy chain variable region(VH) genes that begin with VH3 subgroup. Then, the products were cloned to T-vector, and all white colonies were chosen for DNA-sequencing. Finally, the sequence ofevery insert was submitted to IMGT/V-QUEST network database for comfirming the genotype of VH (IGHV), D(IGHD) and JH(IGHJ), or blasted in Bioedit software for confirming the genotype of Cγ. Results A total of 56 clones containing insert were chosen and verified by DNA sequencing, and all comprised of VH3, D, JHand Cγ germline genes. These 56 sequences were divided into 49 kinds, in which 5 kinds came from 2 or 3 clones. Of 49 kinds of sequences, there appeared all 4 Cγ functional genes(IGHG2 topped with 28 clones), 11 of 23 VH3 functional genes(IGHV3-23 topped with 29 clones), 16 of 23 D functional genes(IGHD1-26 topped with 8 clones) and all 6 JH functional genes(IGHJ4 topped with 33 clones). Irrespective of constant-regions, there were 33 kinds of VH3-D-JH combinations. The［IGHV3-23]-［IGHD1-26］-［IGHJ4 combination was the topmost one, which appeared in 8 clones. Although 5 of which were linked to the same constant-region gene, IGHG2, their variable region sequences still had significant differences. Conclusion We succeeded in amplifying IgG heavy chain variable region genes rapidly and specifically from human peripheral blood with high titer of HBsAb, and found that it is practical to categorize antibody in HBIG or B cells in blood based on variable region DNA-sequencing, but the antibody diversity is so vast that it needs to magnify the number of clones al least several scales. References | Related Articles | Metrics

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Effects of RhoA/ROCKI signal pathway on the taxol-induced cell-cycle arrest of cervical cancer cells  WANG Wei-li WANG Ping* WANG Feng HUANG Shuai-shuai REN Yu 2013, 44 (4 ):  485-491.  doi: 10.3969/j.issn.0529-1356.2013.04.008 Abstract ( )   Objective To explore the effects of RhoA/ROCKI signal pathway in taxol-induced cell cycle arrest of cervical cancer cells. Methods The MTT method was used to determine the minimal inhibition concentration of taxol in C33A cells. Cell cycle and expression of RhoA protein were assayed by flow cytometry. Expressions of ROCKI, cyclins and cyclin-dependent kinases (CDKs)were determined by Western blotting. Results After cells were treated with 1μmol/L of taxol, the cell number of G2/M phase and the expression of RhoA/ROCKI protein increased dramatically in a time-dependent manner. However, the expressions of CDK1,cyclin B1, A and D1 proteins were decreased markedly. Moreover, these effects were totally reversed when cells pretreated with the inhibitor of RhoA (C3 transferase), and partly reversed by the treatment of ROCKI inhibitor (Y-27632), although the significance still existed as compared with the control. No marked effects were observed on the expression of CDK2 in cells treated with taxol or RhoA/ROCKI inhibitors. Conclusion Taxolinduced G2/M arrest in C33A cells was related to the decreases of specific cyclins and CDK, which is tightly regulated by the increased RhoA activity, but ROCKI may only play the role partly in this process. References | Related Articles | Metrics

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Role of ZAC gene in the pathway of octreotide inhibiting proliferation of gastric cancer cells in vitro DAI Wen DING Yi ZHANG Qin-xian* 2013, 44 (4 ):  492-497.  doi: 10.3969/j.issn.0529-1356.2013.04.009 Abstract ( )   Objective To explore the role of ZAC gene in the pathway of somatostatin analogue, octreotide (OCT), inhibiting proliferation of gastric cancer cells. Methods The gastric cancer cells BGC823 and SGC7901 were treated with OCT at various concentrations for various times, respectively. MTT assay was used to screen effective condition of OCT for its antiproliferative action. After treatment with OCT (effective concentration/various times, effective time/various concentrations), the inducing effect of OCT on ZAC gene expression in gastric cancer cells was detected by Western blotting. Three fragments for ZAC gene RNA interference were inserted into pSUPER-EGFP-I vector to construct ZAC-shRNA expression vectors (pSUPER-EGFP-ZAC/1, pSUPER-EGFP-ZAC/2 and pSUPER-EGFP-ZAC/3), respectively. After identification by Hind Ⅲ/EcoRⅠdigestion and sequencing, the three ZAC-shRNA expression vectors were transfected into gastric cancer cells BGC823 and SGC7901, respectively. After G418 screening and RT-PCR identification, the gastric cancer cell lines in which ZAC gene was knock-down were established. Gastric cancer cells (control group) and those in which ZAC gene was knock-down (experimental group) were incubated with OCT at effective condition, the antiproliferative action of OCT was detected by MTT assay. Results The effective condition of OCT inhibiting proliferation of gastric cancer cells was 10nmol/L and treatmented for 24 hours. OCT induced ZAC gene expression in gastric cancer cells in a time-and dose-dependant manner. The results of Hind Ⅲ/EcoRⅠ digestion and sequencing indicated that ZAC-shRNA expression vectors were constructed successfully. The ZAC mRNA level in the gastric cancer cells transfected with shRNA-ZAC/2 was decreased significantly (P<0.05), which were the gastric cancer cell lines in which ZAC gene was knock-down. The proliferation of the gastric cancer cell lines in which ZAC gene was knock-down was significantly higher than that of corresponding BGC823 and SGC7901 (P<0.05). After OCT incubation, the proliferation of BGC823 and SGC7901 were decreased obviously (P<0.05); however, that of the cells in which ZAC gene was knock-down did not exhibit significant changes (P>0.05). Conclusion OCT induces ZAC gene expression in gastric cancer cells in a time-and dose-dependant manner. ZAC gene may play an important role in the pathway of OCT inhibiting the proliferation of gastric cancer cells. References | Related Articles | Metrics

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Clinical significance of nucleolin expression in thyroid papillary carcinoma Lü Shun-zeng ZHAO Jing TIAN Hua* LI Fei LI Gui-bao FANG Yun-hai HU Yan-lai YIN Qun-sheng 2013, 44 (4 ):  498-502.  doi: 10.3969/j.issn.0529-1356.2013.04.010 Abstract ( )   Objective To explore the expression of nucleolin in thyroid papillary carcinoma, and discuss its clinical significance.Methods Nucleolin, chemokine receptor 7 (CCR7), matrix metalloproteinase 9 (MMP-9) were detected by immunohistochemical technique (SP) in 60 cases of normal thyroid tissues and 60 cases of thyroid papillary carcinoma tissues. The relationship among the expressions of nucleolin, CCR7, and MMP-9 and gender, age, or tumor size was analyzed. The correlation between nucleolin and CCR7/MMP-9 was analyzed. Results The expression rate of nucleolin in normal thyroid and thyroid papillary carcinoma tissues was 0, 100% respectively. The expression of nucleolin in thyroid papillary carcinoma with lymph node metastasis was mainly located in nucleus, cytoplasm and cell membrane while the expression of nucleolin was only located in nucleus in thyroid papillary carcinoma tissues without lymph node metastasis. Differences were statistically significant（P<0.05）. No significant correlation was found between the expression of nucleolin and gender, age or tumor size (P>0.05). In thyroid papillary carcinoma, the extra nuclear expression of nucleolin was positively correlated with the expression of CCR7 and MMP-9（P<0.01）. Conclusion Our study suggests that nucleolin can express in thyroid papillary carcinoma and the distribution of nucleolin differs between metastatic carcinoma tissues and nonmetastatic carcinoma. Nucleolin is closely associated with CCR7 and MMP-9. NCL can be used as an important indicator for cancer metastasis. References | Related Articles | Metrics

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Expression and clinical significance of trefoil factor 3 in patiens with papillary thyroid carcinoma ZHANG Xiao-jun WU Jing-fang* HUANG Jing ZHANG Jing ZHANG Jian-hong LIU Jun-chao LIU Bo XUE Gang* 2013, 44 (4 ):  503-508.  doi: 10.3969/j.issn.0529-1356.2013.04.011 Abstract ( )   Objective To investigate the expression and significance of trefoil factor 3(TFF3) in serum and tissue of patients with papillary thyroid carcinoma．Methods A total of 153 patients with thyroid tumor were divided into papillary thyroid carcinoma (PTC) group(PTC,n=46),adenoma group(A,n=51),papillary hyperplasia group(PH,n=36) and normal group,(N,n=153, normal thyroid tissue of every specimen). Sixteen normal human were served as the normal group in ELISA.The expressions of TFF3 protein and mRNA in surgical specimens were detected by immunohistochemistry(IHC) SP, in situ hibridazation (ISH) and ELISA method was used to determine the serum TFF3 level. Results 1. The positive profiless of TFF3 protein and mRNA were brown granules in cytoplasm. The positive  rate of TFF3 in 66 PTC group, 36 PH group, 51 A group and 153 N group were 92.42％, 47.22%,31.37％ and 25.40% respectively.There was a positive correlation between expression of TFF3 mRNA and protein,though a little lower of TFF3 mRNA(P<0.0001). The integral absorbance (IA) of TFF3 protein and mRNA in PTC was higher those in than other three groups (P<0.01). 2. The high expression rate of TFF3 protein/mRNA in patients with thyroid carcinoma was correlated with the TNM stage and lymph node metastasis  (P<0.01). 3. The concentration of TFF3 in serum with thyroid carcinoma was the highest among the four groups（P<0.001）. Serum concentrations of TFF3 in surgical specimens positive group was obviously higher than that in the negative group (P<0.05). Conclusion TFF3 protein/mRNA expresses strongly in PTC tissues. Detection of the serum TFF3 concentration may be a reference index in diagnosis of thyroid carcinoma. References | Related Articles | Metrics

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Expression of 14-3-3zeta and p-Bad proteins in stage T1 non-small cell lung cancer and the significance ZANG Dong-yu* LI Fu-zhi YANG Chun-yu 2013, 44 (4 ):  509-513.  doi: 10.3969/j.issn.0529-1356.2013.04.012 Abstract ( )   Objective To observe the expression of 14-3-3zeta and p-Bad in stage T1 non-small cell lung cancer(NSCLC), and to investigate the relationship between the expression of 14-3-3zeta and p-Bad in NSCLC. Methods One hundred and ten paraffin blocks and 50 fresh samples from NSCLC patients and normal lung were observed. Expression of 14-3-3zeta and p-Bad were detected by immunohistochemistry and Western blotting. Results Up-regulation of 14-3-3zeta was observed significantly in stage T1 NSCLC in contrast to that in the normal lung. The increased 14-3-3zeta protein level was correlated with differentiation degree and lymph node metastasis, but it was disassociated with age, gender and histological type. Decreased expression of p-Bad was observed in NSCLC in contrast to that in the normal lung. Expression of p-Bad was related to differentiation degree and lymph node metastasis, but it was disassociated with age, gender and histological type. Conclusion 14-3-3zeta and p-Bad play an important role in development and progression of NSCLC. References | Related Articles | Metrics

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Comparision of observations between coronal histological and CT images of the lateral skull base LI Xi-ping XIA Yin HAN De-min* 2013, 44 (4 ):  514-518.  doi: 10.3969/j.issn.0529-1356.2013.04.013 Abstract ( )   Objective To obtain digital image data from sequential sectional coronary celloidin slices of the temporal bone in order to provide references for imaging sectional study and clinical diagnosis of the lateral skull base. Methods Two cadaveric heads were used to make sectional specimens after spiral CT scanning. After celloidin embedding, a brain slicer was used for making thin slices(100 μm). Digital camera was used to take pictures after each slicing.Representative slices were stained by HE method.Digital pictures,HE pictures and CT images were examined and comparied to identify characteristics of sectional structures. Results Two sets of sequential digital images were obtained with a resolution of 1920×2560 and spatial relationship among different structures was clearly shown. Sectional structures were demonstrated more clearly after contrasting observation. Conclusion Celloidin embedding is an ideal method for obtaining high resolution sequential digital imaging data of the lateral skull base. Coronary images are suitable to show ossicles, Prussak’s space,jugular vein fossa,the relationship between cochlear and middle cranial fossa, the jugular and petrous segment of internal carotid artery,the vertical segment of facial nerve,the relationship between internal acoustic canal and middle cranial fossa. References | Related Articles | Metrics

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Distribution and function of cellular retinoic acid binding protein 1 positive neural crest cells of mouse embryo YANG Yan-ping WU Shan-shan JING Ya* LI Hai-rong QIAO Cong-jin ZHANG Tao 2013, 44 (4 ):  519-524.  doi: 10.3969/j.issn.0529-1356.2013.04.014 Abstract ( )   Objective To investigate the formation and the distribution pattern of cardiac neural crest and its function during the development of cardiovascular system of mouse embryo. Methods Serial sections of forty-five mouse embryos during embryonic day(ED) 8 to ED 12 were stained immunohistochemically with antibodies against cellular retinoic acid binding protein 1 (CRABP1), α-smooth muscle actin (α-SMA), myosin heavy chain (MHC) and islet-1 (Isl-1). Results At ED8, CRABP1 had not been expressed in the ectoderm of neural fold. During ED8.5 to ED9, at the level of cardiac tube and branchial arch, immunohistochemical positive reactive cells for CRABP1 were observed in neural fold and a part of them delaminated from neural fold entered into the neighbor mesechyme. At ED10, CRABP1 positive cells in the bilateral mesenchyme of the neural tube migrated into branchial arch, exterior of aortic arch endothelium and the cardiac jelly of the outflow tract. From ED11 to ED12, CRABP1 expression in immunohistochemical reactive positive cells was down-regulated in the mesenchyme surrounding the aortic arch endothelium and in the endocardial cushion. While α-SMA immunohistochemical positive staining intensity in the smooth muscle cells of aortic arch wall was increased. Isl-1 positive cells were shown in the aortic-pulmonary septum and the walls of ascending aorta and pulmonary trunk septated by the septum, where CRABP1 was negative staining. Conclusion The time window of CRABP1 positive neural crest formation is from ED8.5 to ED9 in mouse embryo. After ED10, CRABP1 positive cells migrate and contribute to the formation of the tunica media smooth muscle of the aortic arch and endocardial cushion of the outflow tract. CRABP1 could not be used to mark the neural crest cells after migration. References | Related Articles | Metrics

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Expression of integrin CD11a, CD11b and CD11c during the rat heart development MU Ling-min WANG Wen-feng GUO ZHi-kun* ZHANG Guang-mou 2013, 44 (4 ):  525-529.  doi: 10.3969/j.issn.0529-1356.2013.04.015 Abstract ( )   Objective To study the expression changes of integrins CD11a, CD11b and CD11c during the rat heart development.Methods By using RT-PCR and immunohistochemistry, the genes and the protein expression of CD11a, CD11b and CD11c of rat cardiac tissue were detected on embryonic day 18(E18d), postnatal 5 days(P5d), postnatal 19 days(P19d), postnatal 40 days(P40d) and postnatal 1 year(P1y). Results Immunohistochemical results showed that CD11a, CD11b and CD11c expressed in cell plasma of rat heart tissue, and the level of their expression was gradually decreased from E18d to P1y. RT-PCR results showed that the expressions of CD11a, CD11b and CD11c between each group were positive. Among them, the changes of CD11a transcripts were not significant compared with that of P40d and P19d (P>0.05), and there were significant changes among other groups (P<0.05). The expressions of CD11b on E18d, P5d, P19d and P40d were of no significant changes compared with each other respectively (P>0.05), and there was statistical significant in the other groups. The expression of CD11c between groups was statistically significant (P<0.01). Conclusion The expression of CD11a, CD11b and CD11c exists a quantity change during the development of the rat myocardium, and different structures of integrin molecules showed similar expression rules during the heart development, so they may be play an important role on regulation in the development of myocardial cells. References | Related Articles | Metrics

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Changes of homocysteine metabolism key enzyme in insulin resistance rat model FAN Xiao-ming GUO Jia-zhi LIU Shi-chang ZHANG Tie-jun KANG Li LU Di GUI Li* LI Shu-de* 2013, 44 (4 ):  530-534.  doi: 10.3969/j.issn.0529-1356.2013.04.016 Abstract ( )   Objective To research the homocysteine change in an insulin resistance model for SD rats, and to discusse the molecular mechanism of the homocysteine change. Methods Twenty healthy rats, six weeks of age, were randomly divided into the control group (n=10) and the insulin resistance group (n=10). Normal food was fed in control group；The insulin resistance model was established by high fat diet and gavaged by the high fat for 3 months. The concentrations of homocysteine, glucose, insulin, total cholesterol and triglyceride were measured in the fasting state. At the same time, the insulin resistance index (ISI) was calculated. The expressions of methylenetetrahydrofolate reductase (MTHFR), cystathionine β synthase (CBS) and methionine synthetase (MTR) were detected by immunohistochemistry, RT-PCR and Western blotting in the rat liver tissue. Results The concentrations of the blood glucose and blood insulin in the insulin resistance group were higher than that in the control group (P<0.05)， the insulin resistance index was increased, so the insulin resistance model was successed. The concentrations of triglyceride and homocysteine were increased (P<0.05). However, the concentration of the total cholesterol was no statistical significance (P>0.05). The protein and mRNA expressions of MTHER and CBS in insulin resistance group were lower compared with the control group by RTPCR, Western blotting and immunohistochemical staining(P<0.05). However, the expression of MTR protein and mRNA were not different between the insulin resistance group and the control group (P>0.05). Conclusion The homocysteine concentration is higher in the condition of insulin resistance, which may be related to decrease with the expression of MTHER and CBS in metabolic transformation of homocysteine. References | Related Articles | Metrics

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Establishment and optimization of the culture system of mouse oocyte in vitro maturation GE Li DU Hui LIU Li-wei SU Yan-ping* 2013, 44 (4 ):  535-540.  doi: 10.3969/j.issn.0529-1356.2013.04.017 Abstract ( )   Objective To study the effects of the duration of pregnant mare serum gonadotropin(PMSG) priming and in vitro culture (IVC) on nuclear maturation, and the effects of the different activation schemes on the abilities of parthenogenetic activation and development. Methods To detect the rate of nuclear maturation at different time points of IVC by using PMSG priming(each treatment had at least 3 replicates, 3 mice per replicate).In order to determine the optimum activation scheme, two schemes including: (1) ethanol combining with 6-dimethylaminopurine(6-DMAP) and (2) SrCl2 were used, CZB［fetal bovine serum(FBS) and CZB［bovine serum albumin(BSA)］ were used for embryo culture. In order to determine the optimum activation age, the oocytes matured at the different time points were activated. To investigate the effect of PMSG on the ability of oocyte development by shortening the duration of PMSG priming. Results The time of IVC when oocytes reached the highest rate of nuclear maturation (97.6% vs 91.9%) was prolonged from 14 hours to 16 hours if shortening the duration of PMSG priming from 46 hours to 24 hours. Shortening the duration of PMSG priming did not affect the rate of nuclear maturation, however, the rates of activation（91.2% vs 37.1%）and blastocyst（20.9% vs 0.0%）were significantly reduced. Two schemes used in the present study were able to induce the activation rate of oocytes to more than 90% in the in vitro maturation system, however, the rates of blastocyst were significantly different (P<0.05). The rates of activation (89.5%) and blastocyst (21.9%) reached the highest points from 24 hours to 26 hours during IVC. Conclusion A relatively ideal IVC system has been established in the present study. PMSG stimulation duration is 46 hours, oocytes cultured for 24 hours, activated with CZB (10mmol/L SrCl2) and embryos cultured in CZB (0.5% BSA). References | Related Articles | Metrics

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Morphological features of endothelial cells of arteries and cardiac valve in the pig GAO Hong-quan LI Xue-min WU Cai-qin GUAN Guo-ping WANG Fu-jun WANG Lu WANG Ke-qiang ZHANG Hong-qi* 2013, 44 (4 ):  541-545.  doi: 10.3969/j.issn.0529-1356.2013.04.018 Abstract ( )   Objective To investigate the morphological feature of endothelial cells of large and middle-sized arteries in the pig and provide basic data for experimental and clinical research of blood vessels. Methods Animals were perfused with heparin physiological saline and fixed with perfusion of formaldehyde at the physiological pressure. The ascending aorta,arotic arch, descending aorta, aortic valve, common carotid artery,internal thoracic artery, coronary arteries and their anterior and posterior descending branches were collected and observed under a scanning electron microscope. Results Endothelial cells were fusiform and the nuclei slightly extruded to the lumen. The long axis of the cells was consistent with the direction of the blood stream in the straight part of large and middle-sized arteries.In curved part of large artery and aortic valve, endothelial cells were ovary and rich in microvilli.There were regenerating endothelial cells in the posterior descending branch of the right coronary artery. In the common carotid artery,internal thoracic artery and coronary arteries,a few cells with long processes were seen, their cell bodies appeared as round, ovary, triangular or irregular shape, the number of processes varied from two to six, and some processes were bifurcated. Conclusion The size,shape and arrangement of endothelial cells and the length and density of microvilli are different at different locations. Shear stress may be one of major causes leading to these difference. References | Related Articles | Metrics

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Expression of thyroid hormone receptors in the heart of neonatus rats with maternal hyperthyroidism CHEN Jing* LI Bin SONG Fang FENG Ruo-peng ZHAO Zi-wei 2013, 44 (4 ):  546-549.  doi: 10.3969/j.issn.0529-1356.2013.04.019 Abstract ( )   Objective To detect the expression of thyroid hormone receptors(TR), including TRα1,TRα2 and TRβ1. Methods The animal model of hyperthyroidism was established (population and quantity:24 female Wistar rats). Took the neonatal rat hearts of day 5, day 10 and day 15 after birth and stained paraffin sections with Masson staining. Detected expression of thyroid hormone receptors including TRα1,TRα2 and TRβ1 by real-time PCR. Results In the cardiac tissue, the TRα1 expression intensities were higher than TRa2 and TRβ1. In addition, there was a peak in day 10 of neonatal rats both in the hyperthyroidism and the normal ones. In the hyperthyroidism rat, the TRα1 expression intensities in day 5 and day 10 increased significantly,while it decreased in day 15 in hyperthyroidism. There was no significant difference of TRa2 expression in day 5, day 10 and day 15 between hyperthyroidism and normal. The TRβ1 expression intensity decreased in day 5 in hyperthyroidsm, however, it increased in day 10. Conclusions The maternal hyperthyroidism may influence the cardiac muscle of the neonatal rats by the different expression of thyroid hormone receptors. Due to the significant change of TRα1, it may play a very important role in myocardial damage of hyperthyroidism. References | Related Articles | Metrics

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Expression of peroxiredoxin I-thioredoxin, superoxide dismutases and catalases mRNA in hepatic ischemia-reperfusion injury of rats ZHANG Li-hua WANG Qie LIU Zhao WANG Lei* 2013, 44 (4 ):  550-553.  doi: 10.3969/j.issn.0529-1356.2013.04.020 Abstract ( )   Objective To observe the expression change of peroxiredoxin I-thioredoxin (PrxI-Trx), superoxide dismutases (SOD) and catalases (CAT) in hepatic Ischemia-Reperfusion injury of rats, and to explore their role in oxidative-stress response. Methods The hepatic ischemia-reperfusion injury model of rats was established by clipping the hepatic blood vessel released both hepatic left and middle lobes as well as bile duct pedicle with non-damage vascular clamp for 30 minutes. The serum and hepatic tissues were taken 6 hours after clipping. The morphological changes were observed under a microscope using HE staining.The serum alanine aminotransferase(ALT) was detected by Automatic biochemical analyzer. The PrxI-Trx, SOD and CAT mRNA expression were evaluated by RT-PCR. The protein level of PrxI, SOD and CAT were estimated by Western blotting. Results Compared with the control group, the ALT level and HE results of hepatic ischemia-reperfusion injury group showed that liver cells were significantly impaired. The PrxI-Trx, CAT and SOD mRNA expression were increased significantly. The protein level of PrxI, SOD and CAT also increased significantly. Conclusion PrxI-Trx, SOD and CAT may play an important role in inhibiting the oxidative-stress response induced by ischemia-reperfusion injury. References | Related Articles | Metrics

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Comparison and analysis of crown and cusp base area of the first molar crown by ancient Chinese crowds ZHOU Ya-wei HE Le-tian* 2013, 44 (4 ):  554-558.  doi: 10.3969/j.issn.0529-1356.2013.04.021 Abstract ( )   Objective To explore the differences of crown area of the first molar teeth and relative basal cusp area among the population of different feeding habits by measuring and analyzing crown basal area and relative basal cusp area of M1 and M1 of 184 teeth from five ancient Chinese archaeological sites. Methods High precision classification and measurement were done on crown area and cusp area by using the technology of modern digital image measurement, and data was detected and analyzed by using spss 19.0. Results The crown area of teeth on M1 of crowds which took in high portion of animal food was obviously larger than crowds which had plant food. The test of heterogeneity indicated that there was significant difference on the crown area between the two crowds. The relative basal cusp area of the two crowds had extraordinary similarity. The test of heterogeneity indicated no conspicuous difference. The order of the size of the first molar teeth of the ancient people in China which was represented by five archaeological sites showed consistency, that the relative basal cusp areas exhibited a sequence of protocone> paracone>metacone>hypocone in M1 and protocone>hypoconid>metaconid>entaconid>hypoconulid in M1. Conclusion The size of crown basal area of the first molar teeth shows diversity between groups, and the size of relative basal cusp area of the first molar teeth shows consistency between the groups. References | Related Articles | Metrics

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Genetic polymorphism of mitochondrial DNA hypervariable segment Ⅰof Mulam ethnic group in Guangxi WANG Xiao-qing WANG Chuan-chao DENG Qiong-ying* LI Hui 2013, 44 (4 ):  559-562.  doi: 10.3969/j.issn.0529-1356.2013.04.022 Abstract ( )   Objective To demonstrate the polymorphism of the mitochondrial DNA (mtDNA) hypervariable segment Ⅰ(HVSⅠ) and to clearly understand the maternal genetic structure of Mulam ethnic group in Guangxi. Methods Venous blood samples from 91 unrelated Mulam individuals were collected. HVS I of mtDNA was amplified using primers L15947 and R16488. PCR amplicons were sequenced in both forward and reverse directions using the ABI 3730. Polymorphisms were scored relative to the revised Cambridge reference sequence (rCRS). The number of polymorphic sites, number of haplotypes, the mean number of pairwise differences and genetic distances between populations were calculated. Polygenetic tree was drawn based on dA distance. Results MtDNA HVSⅠsequences showed high levels of diversity. Seventy-four haplotypes were identified among the 91 samples. The nucleotide diversity was 0.0188±0.010 and the mean number of pairwise differences was 6.618±3.154. Genetic distances between populations and the cluster map show that Mulam falls together with southern groups, but away from northern populations. Among southern populations, Mulam clusters were most closely with the Zhuang and Sui. Conclusion Mulam is a typical southern Kam-Tai population, and may experience population expansion or selection. MtDNA HVS-Ⅰ reveals highly genetic diversities in Mulam, which may have potential use in population genetics and forensic practice. References | Related Articles | Metrics

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Physical development of Tujia children and adolescents HUANG Da-yuan* ZHANG Hui-juan XIONG Jian WU Guo-yun LIANG Cheng-qing 2013, 44 (4 ):  563-569.  doi: 10.3969/j.issn.0529-1356.2013.04.023 Abstract ( )   Objective To explore the feature and physical growth patterns of the Tujia nationality children and adolescents in Wuling mountain area. Methods According to the anthropometric method, seventeen physical development items for 1865 Tujia children and adolescents（953 males and 912 females）in Wuling mountain area were investigated, and eleven physical indexes were calculated by formula．Results The characteristic of growth development of Tujia children and adolescents was that the mean value of measuring items gradually increased with age. The growth curves of both Tujia boys and girls were on the rise and appeared crossover phenomenon. The age at peak height velocity was at 11-13 years old for boys while 9 and 11-12 years old for girls. The age changing law of physical development indexes of Tujia children and adolescents were similar to that of other nationalities, while there were differences in the development degree of physical body. Conclusion The physical development of Tujia children and adolescents is in conformity with the general growth pattern，there is gender difference in physical development between male and female of Tujia children and adolescents, and the physical development of Tujia children and adolescents is relatively behind the development of Han and other nationalities. References | Related Articles | Metrics

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Comparative study of the dextran-DTPA-Gd and gadopentetate dimegiumine in interstitial magnetic resonance lymphography WU Jing SUN Yue WAN Sheng-biao LIU Shi-en FENG Yuan-yong LI Wei XU Wen-jian JIANG Tao* SHANG Wei*  2013, 44 (4 ):  570-575.  doi: 10.3969/j.issn.0529-1356.2013.04.024 Abstract ( )   Objective To compare the feasibility of interstitial magnetic resonance(MR) lymphography using subcutaneous injection of dextran-DTPA-Gd and gadopentetate dimeglumine. Methods The 1.2ml dextran-DTPA-Gd (3.96×10-3 mol/L) was injected subcutaneously into the interdigital skin fold of the dorsal aspect of the right hind leg in 12 New Zealand rabbits. The 1.2ml gadopentetate dimeglumine(0.4998mol/L) was injected into the left hind leg in the same rabbit after 30 minutes. MR images were obtained before injection and at 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60 minutes and 2,4,24 hours after injection by the 3D TOF CE-MRA sequence.The signal intensities of popliteal lymph node were measured, the enhancing rates(E%) were calculated and the signal-time course curve was drawn. Results The bilateral popliteal lymph node showed the same signal intensities. The lymphatic vessels in the right hind limb drainage area and popliteal lymph node were strengthened significantly and the E% of popliteal lymph node was 212.7% at 10 min after injecting dextran-DTPA-Gd, reached the peak(314.1%) by 35min, was 208.2% at 4 hours and disappeared 24 hours later. The signal intensities of the left lymphatic and popliteal lymph node were weak and the E% of popliteal lymph node was 78.8% 10 min after injecting gadopentetate dimeglumine, reached the peak(98.3%) by 20min, was 29.0% 4 hours, and disappeared 24 hours later. The E% was statistically significant (P<0.05). Animals in each group during the observation period were no significant adverse reactions, biochemical examinations were not statistically significant (P>0.05). Histological examination of the injected organs showed no obvious pathological changes. Conclusion Compared with small molecular gadopentetate dimeglumine, the dextran-DTPA-Gd is an effective lymphatic contrast agent, which can specific strengthen the lymph nodes and lymphatic. References | Related Articles | Metrics

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Improvement of Masson staining in renal fibrosis of mice WEI Sheng-nan ZENG Xiang-jun LI Hui-hua* 2013, 44 (4 ):  576-579.  doi: 10.3969/j.issn.0529-1356.2013.04.025 Abstract ( )   Objective To get an improved Masson 3 staining method for staining renal fibrosis. Methods Bouin‘s fluid was used to re-fix the paraffin fixed sections. Ethanol∶acetic acid（2∶1) which was stored at -20℃ was used to treat the sections; Brilliant green was used to substitute aniline blue. Duration of brilliant green staining was decreased. Results Renal interstitium of fibrosis and non-fibrosis were distinguished. Collagen fibers appeared green (counter staining by brilliant green) and cytoplasm and muscle fibers were red. Conclusion In improved Masson 3 staining, tissues of fibrosis and non-fibrosis are distinguished by brilliant green, the staining results are stable and uniform. The method demonstrates a satisfied compared staining result. References | Related Articles | Metrics

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Review of the major components of neuron dendrites and their function in noncanonical transportation QIANG Wen-bin ZHANG Xiao-min LUO Jian-hong* 2013, 44 (4 ):  580-584.  doi: 10．3969/j.issn.0529－1356.2013.04.026 Abstract ( )   Objective Proteins synthesis and transportation in cells are complex processes and have been strictly controlled, especially in neurons which possess many specialized structures. The non-canonical transportation in dendrites is different from canonical transportation in the soma, as the former mainly consists of dendrite mRNA, dendrite endoplasmic reticulum and Golgi outposts. In recent years, many studies have been undertaken to elucidate the appearance, function and roles of organells in pathology during non-canonical transportation. References | Related Articles | Metrics