2’-5’寡聚腺苷酸合成酶（OAS）及其临床意义的研究进展

干扰素作用于靶细胞膜表面的受体后,通过信号转导系统诱导一系列抗病毒蛋白产生,干扰病毒复制以达到抗病毒目的。2’-5’寡聚腺苷酸合成酶（2’．5’oligoadenylatesynthetase,OAS）是干扰素作用于细胞后产生的一种重要的抗病毒蛋白,几十年来,国内外学者对OAS家族及其抗病毒机制进行了大量研究并取得了一定的进展,OAS被dsRNA激活后,催化生成2-5A,2-5A激活核酸内切酶RNaseL,降解病毒RNA,阻断病毒蛋白合成,从而发挥抗病毒作用。体内外研究表明,OAS的表达量或活性的变化可用于评价机体对干扰素的反应,反映干扰素抗病毒效果,另外,它还可作为系统性红斑狼疮的病情活动度的一种检测指标。因此,OAS具有重要的临床应用价值。本文就OAS家族及其抗病毒机制,其测定方法与对于病毒性肝炎和系统性红斑狼疮疾病的临床意义展开综述,以期对OAS的研究和应用提供参考。OAS是典型的干扰素诱导产物,可反映机体内干扰素的抗病毒水平,具有广阔的应用前景。     

Distinct Contributions of TNF Receptor 1 and 2 to TNF-Induced Glomerular Inflammation in Mice

TNF is an important mediator of glomerulonephritis. The two TNF-receptors TNFR1 and TNFR2 contribute differently to glomerular inflammation in vivo, but specific mechanisms of TNFR-mediated inflammatory responses in glomeruli are unknown. We investigated their expression and function in murine kidneys, isolated glomeruli ex vivo, and glomerular cells in vitro. In normal kidney TNFR1 and TNFR2 were preferentially expressed in glomeruli. Expression of both TNFRs and TNF-induced upregulation of TNFR2 mRNA was confirmed in murine glomerular endothelial and mesangial cell lines. In vivo, TNF exposure rapidly induced glomerular accumulation of leukocytes. To examine TNFR-specific inflammatory responses in intrinsic glomerular cells but not infiltrating leukocytes we performed microarray gene expression profiling on intact glomeruli isolated from wildtype and Tnfr-deficient mice following exposure to soluble TNF ex vivo. Most TNF-induced effects were exclusively mediated by TNFR1, including induced glomerular expression of adhesion molecules, chemokines, complement factors and pro-apoptotic molecules. However, TNFR2 contributed to TNFR1-dependent mRNA expression of inflammatory mediators in glomeruli when exposed to low TNF concentrations. Chemokine secretion was absent in TNF-stimulated Tnfr1-deficient glomeruli, but also significantly decreased in glomeruli lacking TNFR2. In vivo, TNF-induced glomerular leukocyte infiltration was abrogated in Tnfr1-deficient mice, whereas Tnfr2-deficiency decreased mononuclear phagocytes infiltrates, but not neutrophils. These data demonstrate that activation of intrinsic glomerular cells by soluble TNF requires TNFR1, whereas TNFR2 is not essential, but augments TNFR1-dependent effects. Previously described TNFR2-dependent glomerular inflammation may therefore require TNFR2 activation by membrane-bound, but not soluble TNF.     

免疫毒素的抗肿瘤研究

免疫毒素是由具有导向能力的载体（抗体或细胞因子）和具有细胞毒性的分子（毒素）偶联而成的具有特异性细胞杀伤能力的杂合分子,是一种靶向药物。免疫毒素首先利用肿瘤细胞上导向分子的受体与细胞结合,然后进入细胞,再由毒素发挥蛋白质合成抑制作用,最终导致靶细胞死亡。与其他抗肿瘤药物相比,免疫毒素具有毒性强和特异性高的优点,在肿瘤治疗中显示出巨大的应用前景。简要概述了免疫毒素的作用机理、制备及其抗肿瘤研究进展。     

Quantitative analysis of isoprenoid diphosphate intermediates in recombinant and wild-type Escherichia coli strains

In biotechnology, the heterologous biosynthesis of isoprenoid compounds in Escherichia coli is a field of great interest and growth. In order to achieve higher isoprenoid yields in heterologous E. coli strains, it is necessary to quantify the pathway intermediates and adjust gene expression. In this study, we developed a precise and sensitive nonradioactive method for the simultaneous quantification of the isoprenoid precursors farnesyl diphosphate (FPP) and geranylgeranyl diphosphate (GGPP) in recombinant and wild-type E. coli cells. The method is based on the dephosphorylation of FPP and GGPP into the respective alcohols and involves their in situ extraction followed by separation and detection using gas chromatography–mass spectrometry. The integration of a geranylgeranyl diphosphate synthase gene into the E. coli chromosome leads to the accumulation of GGPP, generating quantities as high as those achieved with a multicopy expression vector. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. T. Vallon and S. Ghanegaonkar contributed equally to this work.     

PKCλ/ι signaling promotes triple-negative breast cancer growth and metastasis

Triple-negative breast cancer (TNBC) is a distinct breast cancer subtype defined by the absence of estrogen receptor (ER), progesterone receptor (PR) and epidermal growth factor receptor 2 (HER2/neu), and the patients with TNBC are often diagnosed with higher rates of recurrence and metastasis. Because of the absence of ER, PR and HER2/neu expressions, TNBC patients are insensitive to HER2-directed and endocrine therapies available for breast cancer treatment. Here, we report that expression of atypical protein kinase C isoform, PKCλ/ι, significantly increased and activated in all invasive breast cancer (invasive ductal carcinoma or IDC) subtypes including the TNBC subtype. Because of the lack of targeted therapies for TNBC, we choose to study PKCλ/ι signaling as a potential therapeutic target for TNBC. Our observations indicated that PKCλ/ι signaling is highly active during breast cancer invasive progression, and metastatic breast cancers, the advanced stages of breast cancer disease that developed more frequently in TNBC patients, are also characterized with high levels of PKCλ/ι expression and activation. Functional analysis in experimental mouse models revealed that depletion of PKCλ/ι significantly reduces TNBC growth as well as lung metastatic colonization. Furthermore, we have identified a PKCλ/ι-regulated gene signature consisting of 110 genes, which are significantly associated with indolent to invasive progression of human breast cancer and poor prognosis. Mechanistically, cytokines such as TGFβ and IL1β could activate PKCλ/ι signaling in TNBC cells and depletion of PKCλ/ι impairs NF-κB p65 (RelA) nuclear localization. We observed that cytokine-PKCλ/ι-RelA signaling axis, at least in part, involved in modulating gene expression to regulate invasion of TNBC cells. Overall, our results indicate that induction and activation of PKCλ/ι promote TNBC growth, invasion and metastasis. Thus, targeting PKCλ/ι signaling could be a therapeutic option for breast cancer, including the TNBC subtype.Breast cancer is a clinically heterogeneous disease and both intra and inter-tumor heterogeneities provide great challenges for developing successful therapies. Expressions (or absence thereof) of estrogen receptor (ER), progesterone receptor (PR) and epidermal growth factor receptor 2 (HER2)/neu are widely used to clinically classify breast tumors into multiple therapeutic groups.¹ The ER/PR-positive and the HER2-positive breast cancer patients could be benefited from endocrine and HER2-targeted therapies.¹ However, triple-negative breast cancers (TNBCs), which represent ∼12–17% of all breast cancer,² lack ER, PR and HER2/neu expressions² and are not responsive to therapies targeting these receptors. Therefore, the only systemic therapy available for TNBC is chemotherapy.³ Furthermore, TNBC is associated with aggressive pathologic features like higher histology grade and mitotic index⁴ and often found to be associated with higher rate of metastasis and recurrence leading to limited clinical outcome.^{5, 6, 7, 8} Recurrence of TNBC tends to recur within a few years after successful initial treatment^{6, 9} and often develops metastasis to the bone, brain and lungs with poor prognosis.^{2, 6} Thus, identification of signaling pathways that regulate malignant progression of breast cancer subtypes, especially TNBCs, would be therapeutically important.In recent years, PKC signaling has been implicated in modulating invasion and metastasis of multiple tumors.^{10, 11} The PKC family consists of multiple serine/threonine kinases and the relative contribution of individual PKC isoforms during cancer progression varies due to pleiotropism.¹² PKC isoforms regulate diverse cellular functions such as cell-cycle regulation, cellular survival, cell–cell communications and apoptosis.¹³ In particular, atypical PKC isoforms, PKCζ and atypical protein kinase C lamda/iota (PKCλ/ι), are known to be important for chemotaxis, cell polarity, migration and wound healing processes.^{14, 15} Aberrations in all these processes are manifested in tumor progression and metastasis.¹⁴ Consistent with these notions, recent studies indicated that atypical PKCs are associated with various human cancers.^{10, 11} Importantly, the PKCλ/ι gene is located at the 3q26.2 genomic region, which is most frequently amplified in human cancer^{16, 17}, and overexpression of PKCλ/ι has been implicated in cancer development in multiple tissues including the lung,^{18, 19} pancreas,²⁰ stomach,²¹ colon,²² esophagus,²³ liver,²⁴ bile duct,²⁵ ovary,¹⁷ prostate²⁶ and brain.²⁷ Recently, few studies have been reported higher expression of PKCλ/ι in ER/PR- and HER-positive breast cancer and also in lymph node metastases.^{28, 29} Kojima et. al.²⁸ showed that PKCλ/ι expression is highly induced in the ER/PR- and HER2-positive IDCs compared with ductal carcinoma in situ (DCIS) and normal breast. PKCλ/ι forms apical-junctional complexes (AJCs) with other polarity proteins such as partitioning defective 3 homolog (PAR3) and partitioning defective 6 homolog (PAR6),^{30, 31, 32, 33} and invasiveness of breast tumor cells was shown to be associated with loss of PKCλ/ι localization from their apical domains.²⁸ In addition, predominant nuclear localization of PKCλ/ι in both normal and atypical ductal hyperplasia (ADH) lesions prompted the concept that PKCλ/ι might be in an inactive state in these lesions.²⁸ However, expression and activation of PKCλ/ι in TNBCs and the functional importance of PKCλ/ι signaling in relation to invasive breast cancer progression and metastasis are very poorly understood.^{10, 11}Here, we studied PKCλ/ι signaling during invasive progression of TNBC. We utilized expression evaluations in triple-negative IDCs as well as metastatic breast cancers of human patients, in vitro and in vivo functional assays, and global gene expression analysis of human patient samples. We concluded that PKCλ/ι signaling is an important regulator for invasion and metastatic progression of human breast cancers including triple-negative subtypes.     

Growth of a Molecular Base for Feeding: The Mind Body Dualism

It is often difficult to identify the ‘who, when, and where’ of advances in medicine and surgery because it's a rare advance indeed (such as the use of digitalis by William Withering) that can be clearly related to the astuteness of one person at one time and place.     

Intra‐ and interspecific differences in nutrient recycling by European freshwater fish

1. We measured N and P excretion rates of 470 individuals belonging to 18 freshwater fish species widespread in Western Europe. We assessed the effect of body mass on excretion rates at both the intra‐ and interspecific levels. 2. The high variability in per capita N and P excretion rates was mainly determined by differences in body mass. The scaling coefficients of allometric relationships for both N and P excretion rates were significantly lower than 1 (mean ± SE, 0.95 ± 0.04 and 0.81 ± 0.05, respectively). 3. The slope of the allometric relationship between fish mass and nutrient excretion rate was significantly different among species. We did not detect any influence of phylogenetic conservatism on fish mass and on excretion rates. Further investigations are needed to understand the biological determinants of these differences. 4. This high intra‐ and interspecific variability in per capita excretion rates, coupled with differences in fish body mass, produce marked differences in biomass‐standardised excretion rates. These results thus indicate the necessity for further experimental and in situ investigations on the consequences of nutrient recycling by fish in freshwater ecosystems.     

Expression and characterization of the chemokine receptors CCR2 and CCR5 in mice

The chemokine receptors CCR2 and CCR5 play important roles in the recruitment of monocytes/macrophages and T cells. To better understand the role of both receptors in murine models of inflammatory diseases and to recognize potential problems when correlating these data to humans, we have generated mAbs against murine CCR2 and CCR5. In mice CCR2 is homogeneously expressed on monocytes and on 2--15% of T cells, closely resembling the expression pattern in humans. In contrast to humans, murine NK cells are highly CCR5 positive. In addition, CCR5 is expressed on 3--10% of CD4 and 10--40% of CD8-positive T cells and is weakly detectable on monocytes. Using a model of immune complex nephritis, we examined the effects of inflammation on chemokine receptor expression and found a 10-fold enrichment of CCR5(+) and CCR2(+) T cells in the inflamed kidneys. The activity of various chemokines and the antagonistic properties of the mAbs were measured by ligand-induced internalization of CCR2 and CCR5 on primary leukocytes. The Ab MC-21 (anti-CCR2) reduced the activity of murine monocyte chemotactic protein 1 by 95%, whereas the Ab MC-68 (anti-CCR5) blocked over 99% of the macrophage-inflammatory protein 1alpha and RANTES activity. MC-21 and MC-68 efficiently blocked the ligand binding to CCR2 and CCR5 with an IC(50) of 0.09 and 0.6--1.0 microg/ml, respectively. In good correlation to these in vitro data, MC-21 almost completely prevented the influx of monocytes in thioglycollate-induced peritonitis. Therefore, both Abs appear as useful reagents to further study the role of CCR2 and CCR5 in murine disease models.