T Cells Contribute to Stroke-Induced Lymphopenia in Rats

Stroke-induced immunodepression (SIID) results when T cell and non-T immune cells, such as B cells, NK cells and monocytes, are reduced in the peripheral blood and spleen after stroke. We investigated the hypothesis that T cells are required for the reductions in non-T cell subsets observed in SIID, and further examined a potential correlation between lymphopenia and High-mobility group protein B1 (HMGB1) release, a protein that regulates inflammation and immunodepression. Our results showed that focal ischemia resulted in similar cortical infarct sizes in both wild type (WT) Sprague Dawley (SD) rats and nude rats with a SD genetic background, which excludes the possibility of different infarct sizes affecting SIID. In addition, the numbers of CD68-positive macrophages in the ischemic brain did not differ between WT and nude rats. Numbers of total peripheral blood mononuclear cells (PBMCs) or splenocytes and lymphocyte subsets, including T cells, CD4⁺ or CD8⁺ T cells, B cells and monocytes in the blood and spleen, were decreased after stroke in WT rats. In nude rats, however, the total number of PBMCs and absolute numbers of NK cells, B cells and monocytes were increased in the peripheral blood after stroke; nude rats are athymic therefore they have few T cells present. Adoptive transfer of WT splenocytes into nude rats before stroke resulted in lymphopenia after stroke similar to WT rats. Moreover, in vitro T cell proliferation stimulated by Concanavalin A was significantly inhibited in WT rats as well as in nude rats receiving WT splenocyte adoptive transfer, suggesting that T cell function is indeed inhibited after stroke. Lastly, we demonstrated that stroke-induced lymphopenia is associated with a reduction in HMGB1 release in the peripheral blood. In conclusion, T cells are required for stroke-induced reductions in non-T immune cells and they are the most crucial lymphocytes for SIID.     

miR-582-5p Is Upregulated in Patients with Active Tuberculosis and Inhibits Apoptosis of Monocytes by Targeting FOXO1

Macrophage apoptosis is a host innate defense mechanism against tuberculosis (TB). In this study, we found that percentage of apoptotic cells in peripheral blood monocytes from patients with active TB was lower than that from healthy controls (p<0.001). To understand whether microRNAs can modulate apoptosis of monocytes, we investigated differentially expressed microRNAs in patients with active TB. miR-582-5p was mainly expressed in monocytes and was upregulated in patients with active TB. The apoptotic percentage of THP-1 cells transfected with miR-582-5p mimics was significantly lower than those transfected with negative control of microRNA mimics (p<0.001), suggesting that miR-582-5p could inhibit apoptosis of monocytes. To our knowledge, the role of miR-582-5p in regulating apoptosis of monocytes has not been reported so far. Systematic bioinformatics analysis indicated that FOXO1 might be a target gene for miR-582-5p and its 3′UTR contains potential binding sites for miR-582-5p. To determine whether miR-582-5p could influence FOXO1 expression, miR-582-5p mimics or negative control of microRNA mimics were transfected into THP-1 cells. RT-PCR and western blot analysis showed that the miR-582-5p could suppress both FOXO1 mRNA and protein expression. Co-transfection of miR-582-5p and FOXO1 3′UTR-luciferase reporter vector into cells demonstrated that significant decrease in luciferase activity was only found in reporter vector that contained a wild type sequence of FOXO1 3′UTR, suggesting that miR-582-5p could directly target FOXO1. In conclusion, miR-582-5p inhibited apoptosis of monocytes by down-regulating FOXO1 expression and might play an important role in regulating anti-M. tuberculosis directed immune responses.     

Synthetic osteogenic growth peptide promotes differentiation of human bone marrow mesenchymal stem cells to osteoblasts via RhoA/ROCK pathway

The osteogenic growth peptide (OGP) is a naturally occurring tetradecapeptide that has attracted considerable clinical interest as a bone anabolic agent and hematopoietic stimulator. In vitro studies have demonstrated that OGP directly regulates the bone marrow mesenchymal stem cells' (BMSCs) differentiation into osteoblasts. However, the exact mechanism of this process remains unknown. In the present study, we investigated the role of RhoA/ROCK signaling in differentiation along this lineage using human BMSCs. OGP treatment increased the mRNA level of bone morphogenetic protein-2 and alkaline phosphatase activity after osteogenic induction. Analysis of BMSCs induced in the presence of OGP revealed an increase in RhoA activity, and phosphorylation of FAK and cofilin. The ROCK-specific inhibitors, Y27632, blocked the OGP-induced regulation of BMSC differentiation. Taken together, these data suggest that OGP not only acts on BMSCs to stimulate osteogenic differentiation, but also in a dose-dependent manner, and this effect is mediated via the activation of RhoA/ROCK pathway.     

侧柏挥发性次生代谢产物的提取及GC-MS分析

利用气相色谱-质谱联用技术对水蒸气蒸馏提取的侧柏挥发性次生代谢产物进行了分析,研究结果表明年轮低于6年和6～12年侧柏心材质量分数最高的挥发性次生代谢产物均为柏木醇(40.14%,41.13%),年轮12～20年侧柏心材和边材质量分数最高的挥发性次生代谢产物均为8-丙氧基-香松烷(49.71%,44.03%)。侧柏年轮低于6年、6～12年、12～20年心材和边材的挥发性共有次生代谢产物为罗汉柏烯、雪松烯和花侧柏烯,只有6～12年侧柏心材挥发性次生代谢产物中含有愈创木烯且不含有α-杜松醇。     

Effects of chronic chromium(vi) exposure on blood element homeostasis: an epidemiological study

One hundred chromate production workers chronically exposed to low-level of hexavalent chromium [Cr(vi)] and eighty healthy individuals free from Cr exposure were recruited to the study. Personal sampling of airborne Cr was conducted and Cr content was quantified by Flame Atomic Absorption Spectrometry (FAAS). At the end of the sampling shift, blood samples were collected and element concentrations were measured by inductively coupled plasma mass spectrometry (ICP-MS) for Cr, Cd, Cu, Mo and Se and inductively coupled plasma atomic emission spectrometry (ICP-AES) for Ca, Fe, Mg and Zn. According to our results, 90% of the chromate production workers were exposed to airborne Cr in a concentration lower than 50 μg m(-3), which is the threshold limit value recommended by the American Conference of Governmental Industrial Hygienists and Chinese Ministry of Health. After Cr(vi) exposure, a significant increase in blood Cr, Cd, Fe, Mg, Mo, Se and Zn concentrations was observed, as well as a significant decrease in Ca concentration. A decrease in blood Cu was only observed among female workers. Blood Cr concentrations of the exposed workers (median = 15.68 ng mL(-1)) was four times higher than that of the controls (median = 3.03 ng mL(-1)), and significantly correlated with airborne Cr (r = 0.568, P<0.001). In addition, the inter-element correlations exhibited significant differences between the two groups. Our findings of the related health effects suggested that the underlying mechanisms of chronic Cr(vi) exposure on blood element homeostasis might be partly explained by oxidative stress in the body, dysfunction of Fe metabolism and renal injury.     

Haploinsufficiency of SGO1 results in deregulated centrosome dynamics,enhanced chromosomal instability and colon tumorigenesis

Chromosome instability (CIN) is found in 85% of colorectal cancers. Defects in mitotic processes are implicated in high CIN and may be critical events in colorectal tumorigenesis. Shugoshin-1 (SGO1) aids in the maintenance of chromosome cohesion and prevents premature chromosome separation and CIN. In addition, integrity of the centrosome may be compromised due to the deficiency of Cohesin and Sgo1 through the disengagement of centrioles. We report here the generation and characterization of SGO1-mutant mice and show that haploinsufficiency of SGO1 leads to enhanced colonic tumorigenesis. Complete disruption of SGO1 results in embryonic lethality, whereas SGO1^+/− mice are viable and fertile. Haploinsufficiency of SGO1 results in genomic instability manifested as missegregation of chromosomes and formation of extra centrosomal foci in both murine embryonic fibroblasts and adult bone marrow cells. Enhanced CIN observed in SGO1-deficient mice resulted in an increase in formation of aberrant crypt foci (ACF) and accelerated development of tumors after exposure to azoxymethane (AOM), a colon carcinogen. Together, these results suggest that haploinsufficiency of SGO1 causes enhanced CIN, colonic preneoplastic lesions and tumorigenesis in mice. SGO1 is essential for the suppression of CIN and tumor formation.Key words: SGO1, mouse genetics, chromosomal instability, centrosome, colon cancerSince its discovery several years ago, Shugoshin 1 (SGO1) has emerged as a crucial regulator of the cell cycle.^1–6 At cellular and molecular levels, SGO1 functions as a protector of centromeric cohesion of sister chromatids in higher eukaryotes.^5–7 Depletion of SGO1 by small interfering RNA (siRNA) leads to premature sister chromatid separation.^5–8 During mitosis, SGO1 localizes to centromeres in a manner that appears to be dependent on Bub1, Aurora B and survivin.^7–13 SGO1 works in concert with protein phosphatase 2A (PP2A) to protect centromeric cohesion during mitosis and meiosis.^14,15 It is implicated in microtubule dynamics and required for tension generation at the kinetochore.^2,6 In addition to the function of SGO1 in centromeres, sSGO1, a major splice variant of SGO1, has an important function in centrosome dynamics through mediating centriole cohesion.¹⁶ A recent study supports the centrosomal function of Sgo1 in further detail.¹⁷ Importantly, both cohesin and Sgo1 are shown to be involved in engagement of centrioles and thus in centrosomal integrity.¹⁷ Given the importance of centromeric cohesion and centrosome dynamics in the maintenance of chromosomal stability during cell division, it is conceivable that deregulated function of SGO1 would lead to major chromosomal instability.Chromosomal instability has long been appreciated as a driving force for tumorigenesis, since aneuploidy is prevalent in the majority of solid tumors.^18–20 However, several recent studies show that in certain physiological contexts, chromosomal instability induced by deregulated checkpoint genes is not always associated with tumor development in a straightforward manner.^21–24 For example, haploinsufficiency of CENP-E, a spindle checkpoint component, results in enhanced aneuploidy formation and a modest increase in spontaneous tumors in spleen and lung. However, CENP-E^+/− mice develop fewer tumors when these mice are challenged with carcinogens or in a genetically susceptible background, suggesting that chromosomal instability may suppress tumorigenesis in a context-dependent manner.²²Defects in the chromosome cohesion system and SGO1 may play a critical role in genomic instability, and cancers in human colon. Barber et al. attempted to identify genes involved in CIN in human colon cancer with a tumor DNA sequencing approach. They identified 11 somatic mutations distributed among five genes in a part that included 132 colorectal cancers. All but one of these 11 mutations were in the homologs of yeast genes that regulate sister chromatid cohesion, strongly suggesting a critical relationship between chromosome cohesion and CIN in colon cancer.²⁵ Consistently, SGO1 downregulation is implicated in human colon cancer. Among 46 colorectal cancer cases, hSGO1 mRNA expression was decreased in the tumor tissue in comparison with the corresponding normal tissue (p = 0.032).²⁶ However, direct evidence linking SGO1 to colonic tumor development was lacking. Furthermore, no genetic studies have been reported in mouse models with regard to functions of SGO1 in the maintenance of chromosomal stability and acceleration or suppression of tumor development.To determine the physiological function of SGO1, we have generated SGO1 haploinsufficient (+/−) mutant mice. Mouse embryonic fibroblasts (MEFs) from SGO1^+/− animals were found to contain lower levels of SGO1 than MEFs from wild-type embryos. SGO1 deficiency resulted in increased number of spindle centrosomal foci, enhanced chromosome missegregation and formation of micronuclei at an enhanced rate. Moreover, SGO1^+/− animals were prone to higher preneoplastic lesions and rapid development of colonic tumors after exposure to a colon carcinogen.     

Protein kinase Cδ contributes to phenylephrine-mediated contraction in the aortae of high fat diet-induced obese mice

The down-regulation of α-adrenoceptor-mediated signaling casacade has been implicated in obesity but the underlying mechanism remains largely unknown. The present study investigated whether inositol 1,4,5-trisphosphate (IP3) receptor and protein kinase C (PKC) were involved in the reduction of α₁-adrenoceptor agonist phenylephrine-evoked contraction in aortae of high fat diet-induced obese (DIO) mice. C57BL/6 mice were fed with a rodent diet containing 45 kcal% fat for 16 weeks to induce obesity. Isolated mouse aortae were suspended in myograph for isometric force measurement. Protein phosphorylations and expressions were determined by Western blotting. In C57BL/6 mouse aortae, phenylephrine-induced contraction was partially inhibited by either IP3 receptor antagonist heparin or PKC inhibitor GFX, and the combined treatment with heparin and GFX abolished the contraction. Phenylephrine-induced contraction was significantly less in the aortae of DIO mice than those of control mice; only GFX but not heparin attenuated the contraction, indicating a diminishing role of IP3 receptor in DIO mice. Western blotting showed the reduced expression and phosphorylation of IP3 receptor and the down-regulated expression of PKC, PKCβ, PKCδ, and PKCζ in DIO mouse aortae. Importantly, PKCδ was more likely to maintain phenylephrine-mediated contraction in DIO mouse aortae because that (1) PKCδ inhibitor rottlerin but not PKCα and PKCβ inhibitor Gö6976, PKCβ inhibitor hispidin, or PKCζ pseudosubstrate inhibitor attenuated the contraction; and (2) PKCδ phosphorylation was increased but phosphorylations of PKCα, PKCβ, and PKCζ were reduced in DIO mouse aortae. The present study thus provides additional insights into the cellular mechanisms responsible for vascular dysfunction in obesity.     

穿透支原体LAMPs诱导NF-kB激活介导小鼠巨噬细胞凋亡

研究穿透支原体(Mpe)脂质相关膜蛋白(LAMPs)能否诱导小鼠巨噬细胞凋亡,并阐明其可能的分子机制,以了解Mpe潜在的致病性.用Annexin-V-FITC凋亡检测试剂盒和DNA Ladder方法检测Mpe LAMPs诱导体外培养的小鼠巨噬细胞系Raw264.7细胞的凋亡.以间接免疫荧光和Western blotting方法检测经Mpe LAMPs处理的小鼠巨噬细胞NF-κB的激活和NF-kB抑制剂吡咯啉烷二甲基硫脲(PDTC)对细胞凋亡的影响.结果表明:Mpe LAMPs能诱导小鼠巨噬细胞发生早期或晚期凋亡;Mpe LAMPs能诱导激活小鼠巨噬细胞的NF-κB,使其从细胞浆中转位到细胞核内;PDTC能显著地抑制经处理的小鼠巨噬细胞的NF-κB的激活,且能抑制Mpe LAMPs诱导的巨噬细胞发生凋亡.因此,Mpe LAMPs诱导小鼠巨噬细胞凋亡可能与NF-kB的激活有关,因而Mpe可能是一个重要的致病因素.     

中国雪豹的威胁与保护现状

大部分保护机构只有能力在特定种群层面上保护大型食肉动物, 而物种的灭绝风险却是在全球尺度进行评估的。因此, 要填补这一尺度断层, 多机构的工作与意见汇总非常必要。本研究综合了文献和18家中国雪豹(Panthera uncia)研究与保护机构共24位一线工作人员提供的信息, 经过两次集体讨论和填写评分表格的方式, 识别出21种威胁因素, 在全国层面和青海、西藏、新疆、四川和甘肃5个主要雪豹分布省区层面对威胁进行了排序, 并汇总了各保护机构实施的17项保护行动, 以及各项行动所应对的威胁。全国评分前三的威胁依次是基层保护部门能力不足(9.5分)、气候变化(8.0分)、当地社区保护动力不足(6.8分), 不同省区的威胁排序则体现出很大差异性。其中, 目前仍没有任何行动措施应对气候变化。虽然已有一些保护行动来应对基层保护能力不足和当地社区保护动力不足的问题, 如保护区能力建设、开展社区监测等, 但行动的覆盖尺度仍远远不够。