敲减DTX4表达抑制C2C12细胞成肌分化

DTX4（Deltex 4 homolog）蛋白属于Deltex家族成员|Deltex家族是Notch信号通路的调节因子. 已知Notch信号通路在成肌分化中发挥重要作用. 然而,DTX4是否参与调控肌肉发育尚未有报道. 本研究探索DTX4对成肌分化的影响及作用机制. 实时定量PCR和蛋白质印迹分析揭示,伴随小鼠C2C12成肌细胞（myoblast）分化为肌管（myotube）过程,成肌分化标志蛋白肌球蛋白重链（myosin heavy-chain,MyHC）、肌细胞生成素(myogenin)表达逐渐升高,DTX4 mRNA及蛋白质表达水平也逐渐升高. 通过顺序专一的siRNA敲减DTX4表达后,C2C12成肌细胞肌管面积和肌管融合指数明显减少|MyHC、肌细胞生成素蛋白表达水平明显降低|但ERK信号通路未见明显变化.上述结果表明,敲减DTX4表达抑制C2C12细胞成肌分化.我们的结果提示,DTX4可能参与C2C12细胞成肌分化.     

CD133<Superscript>+</Superscript>CD44<Superscript>+</Superscript> subgroups may be human small intestinal stem cells

The identification and separation of small intestinal epithelial stem cells are still on the preliminary stage. In this study, we planned to utilize immunohistochemistry, fluorescence-activated cell sorting (FACS) and RT-PCR to investigate the possibility of CD133 and CD44 as markers of human small intestinal epithelial stem cells. The expressions of CD133, CD44 and Lgr5 were studied by immunohistochemistry. Four subgroups of CD133⁺CD44⁺, CD133⁺CD44⁻, CD133⁻CD44⁺, CD133⁻CD44⁻ were sorted out through FACS and the expression level of Lgr5 gene was measured by RT-PCR and polyacrylamide gel electropheresis (PAGE) with sliver stained. Ten cases of samples were available for analyzing. By immunohistochemical staining, few cells with positive expressions of CD133, CD44 and Lgr5 were distributed in the bottom of crypts with the expression locations somewhat overlapped. The average percentage of CD133⁺CD44⁺ cells was 0.0580 ± 0.0403%, while the corresponding contents of CD133⁺CD44⁻ cells, CD133⁻CD44⁺ cells and CD133⁻CD44⁻ cells were 0.4000 ± 0.1225%, 0.7000 ± 0.2646% and 76.5600 ± 3.5529% respectively. Ten times of positive expressions of Lgr5 were detected in the CD133⁺CD44⁺ groups, while 9/10, 8/10 and 4/10 times for CD133⁺CD44⁻, CD133⁻CD44⁺ and CD133⁻CD44⁻ subgroups respectively. With the help of Quantityone 4.62 software, the densities of corresponding place to Lgr5 and reference gene were obtained. The density ratios of corresponding place to Lgr5 to reference gene were significant difference between subgroups (P < 0.001). By means of LSD method, the density ratios in CD133⁺CD44⁺ subgroups had statistical differences from the other subgroups (P < 0.05). We concluded CD133⁺CD44⁺ cells may be human small intestinal epithelial stem cells, which need further researches to confirm.     

Induction of cytotoxicity by photoexcitation of TiO<Subscript>2</Subscript> can prolong survival in glioma-bearing mice

We have investigated the possibility that photoexcited titanium dioxide (TiO₂) could inhibit the growth of malignant cells. We studied the anti-glioma effects of nano-TiO₂ excited with ultraviolet A (UVA) irradiation both in vitro and in vivo. Transmission electron microscopy demonstrated that glioma cells take up TiO₂ by phagocytosis, and vital staining revealed that TiO₂ alone has no effect on glioma cell proliferation. However, if TiO₂ was combined with UVA irradiation the proliferation rate was decreased significantly compared to controls (P < 0.05). RT–PCR suggested that TiO₂ induction of glioma cell apoptosis is associated with changes in the expression of genes encoding Bcl-2 family members. We then investigated the in vivo antitumor effects of combined TiO₂ plus UVA treatment of established glioma tumors. TiO₂ plus UVA led to pronounced areas of necrosis, elevated indices of apoptosis, delayed tumor growth, and increased survival compared with the TiO₂-alone control group (P < 0.001). Log-rank survival analysis showed that median survival duration was prolonged (P < 0.001). These findings suggest that nano-TiO₂ based photodynamic therapy has potential in the treatment of glioma.     

A barrier-only boundary element delimits the formation of facultative heterochromatin in Drosophila melanogaster and vertebrates

Formation of facultative heterochromatin at specific genomic loci is fundamentally important in defining cellular properties such as differentiation potential and responsiveness to developmental, physiological, and environmental stimuli. By the nature of their formation, heterochromatin and repressive histone marks propagate until the chain reaction is broken. While certain active promoters can block propagation of heterochromatin, there are also specialized DNA elements, referred to as chromatin barriers, that serve to demarcate the boundary of facultative heterochromatin formation. In this study, we identified a chromatin barrier that specifically limits the formation of repressive chromatin to a distal enhancer region so that repressive histone modifications cannot reach the promoter and promoter-proximal enhancer regions of reaper. Unlike all of the known boundary elements identified for Drosophila melanogaster, this IRER (irradiation-responsive enhancer region) left barrier (ILB) does not exhibit enhancer-blocking activity. Not only has the ILB been conserved in different Drosophila species, it can also function as an effective chromatin barrier in vertebrate cells. This suggests that the mechanism by which it functions to spatially restrict the formation of repressive chromatin marked by trimethylated H3K27 has also been conserved widely during evolution.     

Ischemic injury of the liver in a porcine model of cardiac death assessed by in vivo microdialysis

This study aims to evaluate the ischemic injury of the liver in a porcine model of cardiac death assessed by in vivo microdialysis. A porcine model of cardiac death was established by the suffocation method. Metabolic indicators were monitored using the microdialysis technique during warm ischemia time (WIT) and cold ischemia time (CIT). Pathological changes in ischemic-injured livers were observed by haematoxylin–eosin staining. The predictive values of biochemical parameters regarding the liver donor were evaluated by receiver operating characteristic curve analysis. All statistical analyses were conducted using the SPSS 18.0 software (SPSS Inc, Chicago, Illinois, USA). The degree of warm ischemic injury of the livers increased with prolonged WIT. Serum glucose, glycerol, pyruvate, lactic acid levels and lactate-to-pyruvate (L/P) ratio increased gradually during WIT. Results from Pearson correlation analyses indicated that serum lactate level and L/P ratio were positively associated with the degree of warm ischemic injury of the livers. The degree of cold ischemic injury of the livers gradually increased after 12 h CIT. Serum glucose, lactic acid and L/P ratio achieved a peak after 6–8 h of CIT, but gradually decreased with prolonged CIT. The peak of glycerol occurred after 8 h of CIT, while no changes were found with prolonged CIT. Serum pyruvate level exhibited an increasing trend after 12 h CIT. Our results confirmed that serum glucose and lactate levels were negatively correlated with cold ischemic injury of the liver. However, serum glycerol and pyruvate levels showed positive correlations with cold ischemic injury of the liver. The liver donor was unavailable after 30 min WIT and 24 h CIT. The cut-off value of serum lactate level for warm ischemic injury of the livers was 2.374 with a sensitivity (Sen) of 90 % and specificity (Spe) of 95 %; while the L/P radio was 0.026 (Sen = 80 %, Spe = 83 %). In addition, the cut-off values of serum glucose, lactate, glycerol and pyruvate levels for cold ischemic injury of the livers were 0.339 (Sen = 100 %, Spe = 77 %), 1.172 (Sen = 100 %, Spe = 61 %), 56.359 (Sen = 100 %, Spe = 65 %) and 0.020 (Sen = 100 %, Spe = 67 %), respectively. Our findings provide empirical evidences that serum glucose, lactate levels and L/P ratio may be good indicators for the degree of warm ischemic injury of the livers after cardiac death; while serum glucose, lactate, glycerol and pyruvate levels may be important in predicting cold ischemic injury.     

The Rho kinase inhibitor Y-27632 facilitates the differentiation of bone marrow mesenchymal stem cells

The selective in vitro expansion and differentiation of multipotent stem cells are critical steps in cell-based regenerative therapies, but technical challenges have limited cell yield and thus the success of these potential treatments. The Rho GTPases and downstream Rho kinases (Rho coiled-coil kinases or ROCKs) are central regulators of cytoskeletal dynamics during the cell cycle and thus help determine the balance between stem cells self-renewal, lineage commitment, and apoptosis. Here, we examined if suppression of ROCK signaling enhances the efficacy of bone marrow-derived mesenchymal stem cells (BMSCs) differentiation into neurons and neuroglial cells. BMSCs were cultured in epidermal growth factor (EGF, 10 µg/l) and basic fibroblastic growth factor (bFGF, 10 µg/l) in the presence or absence of the Rho kinase inhibitor Y-27632 (10 µM). The expression levels of neuron-specific enolase (NSE) and glial fibrillary acidic protein (GFAP) were detected by immunofluorescence and Western blotting. The average number of NSE-positive cells increased from 83.20 ± 8.677 (positive ratio 0.2140 ± 0.0119) to 109.20 ± 8.430 (positive ratio 0.3193 ± 0.0161) per visual field in the presence of Y-27632, while GFAP-positive cell number increased from 96.30 ± 8.486 (positive ratio 0.18 ± 0.0152) to 107.50 ± 8.683 (positive ratio 0.27 ± 0.0115) (P < 0.05 for both). Both NSE and GFAP protein expression levels were enhanced significantly by Y-27632 treatment (NSE: 0.74 ± 0.05 vs. 1.03 ± 0.06; GFAP: 0.64 ± 0.08 vs. 0.97 ± 0.05, both P < 0.01) as indicated by Western blots. The Rho kinase inhibitor Y-27632 concomitant with EGF and bFGF stimulation promotes BMSC differentiation into neural cells. Control of Rho kinase activity may enhance the efficiency of stem cell-based treatments for neurodegenerative diseases.     

B超引导下甲状腺肿块细针穿刺细胞学诊断的临床病理分析

目的：比较B超引导下细针吸取穿刺和传统细针吸取穿刺在甲状腺肿块性质确定中意义。方法：使用B超引导下细针吸取穿刺（UG-FNAB）和传统细针吸取穿刺技术（C-FNAB）对225例甲状腺肿块进行穿刺活检,然后进行涂片,染色,镜检,结合临床作出细胞学诊断,并与组织切片诊断对照,比较两种方法的确诊率。结果：在UG—FNAB检测病例中59.3％的结节位于甲状腺右叶,40．7％肿块位于左叶和峡部。87％的患者甲状腺机能正常,结节最长径平均为2．8±1．1cm,部分病例符合甲状腺恶性肿瘤的超声影像学改变。在本研究中诊断为甲状腺乳头状癌的病例细胞学和组织学特点均符合甲状腺癌形态学特点。在225例研究病例中,总确诊率为72．89％（164／225）,其中UG-FNAB组确诊率为90．58％（77／85）,C-FNAB组确诊率为62．14％（87／140）,两组确诊率比较有显著性差异（P〈0．05）。结论：本组研究数据显示比较C—FNAB,UG—FNAB在诊断甲状腺复杂性结节中具有更高的确诊率,特别是在C—FNAB方法不能得出明确诊断时。     

单纯治疗慢性前列腺炎对其合并早泄的影响

目的:探讨单纯治疗慢性前列腺炎对其合并早泄的影响。方法:选择372例前列腺炎继发早泄的患者进行单纯针对前列腺炎的治疗,评价前列腺炎的治疗效果(包括前列腺炎症状评分(NIH-CPSI)、前列腺液常规等)及早泄的治疗效果(包括患者性生活满意度评分、配偶性生活满意度评分及阴道内射精潜伏期等)。结果:慢性前列腺炎经综合治疗后,患者的NIH-CPSI评分及前列腺液白细胞计数均显著降低(P0.05)。前列腺炎治愈或好转后,大多数患者的早泄情况得到明显改善,患者性生活满意度、配偶性生活满意度均较治疗前显著提高,阴道内射精潜伏期亦较治疗前明显延长,差异均具有统计学意义(P0.05)。结论:单纯治疗慢性前列腺炎继可使大部分患者并发的早泄明显改善,而对少数前列腺炎好转后早泄症状改善不明显者,可联合应用SSRIs等药物治疗。     

UDP-glucose dehydrogenase modulates proteoglycan synthesis in articular chondrocytes: its possible involvement and regulation in osteoarthritis

Introduction

The objective of this study was to investigate the possible role of UDP-glucose dehydrogenase (UGDH) in osteoarthritis (OA) and uncover whether, furthermore how interleukin-1beta (IL-1β) affects UGDH gene expression.

Methods

UGDH specific siRNAs were applied to determine the role of UGDH in proteoglycan (PG) synthesis in human articular chondrocytes. Protein levels of UGDH and Sp1 in human and rat OA cartilage were detected. Then, human primary chondrocytes were treated with IL-1β to find out whether and how IL-1β could regulate the gene expression of UGDH and its trans-regulators, that is Sp1, Sp3 and c-Krox. Finally, p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580 and stress-activated protein kinase/c-Jun N-terminal kinase (SAP/JNK) inhibitor SP600125 were used to pick out the pathway that mediated the IL-1β-modulated PGs synthesis and gene expression of UGDH, Sp1, Sp3 and c-Krox.

Results

UGDH specific siRNAs markedly inhibited UGDH mRNA and protein expression, and thus led to an obvious suppression of PGs synthesis in human articular chondrocytes. UGDH protein level in human and rat OA cartilage were much lower than the corresponding controls and negatively correlated to the degree of OA. Decrease in Sp1 protein level was also observed in human and rat OA cartilage respectively. Meanwhile, IL-1β suppressed UGDH gene expression in human articular chondrocytes in the late phase, which also modulated gene expression of Sp1, Sp3 and c-Krox and increased both Sp3/Sp1 and c-Krox/Sp1 ratio. Moreover, the inhibition of SAP/JNK and p38 MAPK pathways both resulted in an obvious attenuation of the IL-1β-induced suppression on the UGDH gene expression.

Conclusions

UGDH is essential in the PGs synthesis of articular chondrocytes, while the suppressed expression of UGDH might probably be involved in advanced OA, partly due to the modulation of p38 MAPK and SAP/JNK pathways and its trans-regulators by IL-1β.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-014-0484-2) contains supplementary material, which is available to authorized users.