Trace gases and CO2 isotope records from cabo de rama, India

Concentrations of carbon dioxide (CO2), methane (CH4), carbon monoxide (CO), nitrous oxide (N2O) and hydrogen (H2), and the stable carbon (δ 13C-CO2) and oxygen (δ 18O-CO2) isotopic composition of CO2 have been measured in air samples collected from Cabo de Rama (CRI), India, for the period 1993-2002. The observations show clear signatures of Northern and Southern Hemispheric (NH and SH) air masses, mixed with their regional fluxes and chemical loss mechanisms, resulting in complex seasonal variation of these gases. The CRI measurements are compared with remote marine sites at Seychelles and Mauna Loa. Simulations of two major anthropogenic greenhouse gases (CO2 and CH4) concentrations using a chemistry-transport model for the CRI site suggest that globally optimized fluxes can produce results comparable to the observations. We discuss that CRI observations have provided critical guidance in optimizing the fluxes to constrain the regional source/sinks balance

Conditional Wwox Deletion in Mouse Mammary Gland by Means of Two Cre Recombinase Approaches

Loss of WWOX expression has been reported in many different cancers including breast cancer. Elucidating the function of this gene in adult tissues has not been possible with full Wwox knockout models. Here we characterize the first conditional models of Wwox ablation in mouse mammary epithelium utilizing two transgenic lines expressing Cre recombinase, keratin 5-Cre (BK5-Cre) and MMTV-Cre. In the BK5-Cre model we observed very efficient Wwox ablation in KO mammary glands. However, BK5-Cre Wwox KO animals die prematurely for unknown reasons. In the MMTV-Cre model we observed significant ablation of Wwox in mammary epithelium with no effect on survival. In both of these models we found that Wwox deletion resulted in impaired mammary branching morphogenesis. We demonstrate that loss of Wwox is not carcinogenic in our KO models. Furthermore, no evidence of increase proliferation or development of premalignant lesions was observed. In none of the models did loss of a single Wwox allele (i.e. haploinsufficiency) have any observable phenotypic effect in mammary gland. To better understand the function of Wwox in the mammary gland, transcriptome profiling was performed. We observed that Wwox ablation results in the deregulation of genes involved in various cellular processes. We found that expression of the non-canonical Wnt ligand, Wnt5a, was significantly upregulated in Wwox KO mammary epithelium. Interestingly, we also determined that components of the Jak/Stat3 signaling pathway were upregulated in KO mice and this correlated with a very robust increase in phospho-Stat3 signaling, which warrants further testing. Even though the loss of Wwox expression in breast and other cancers is very well documented, our findings suggest that Wwox does not act as a classical tumor suppressor as previously thought

Generation and Characterization of Mice Carrying a Conditional Allele of the Wwox Tumor Suppressor Gene

WWOX, the gene that spans the second most common human chromosomal fragile site, FRA16D, is inactivated in multiple human cancers and behaves as a suppressor of tumor growth. Since we are interested in understanding WWOX function in both normal and cancer tissues we generated mice harboring a conditional Wwox allele by flanking Exon 1 of the Wwox gene with LoxP sites. Wwox knockout (KO) mice were developed by breeding with transgenic mice carrying the Cre-recombinase gene under the control of the adenovirus EIIA promoter. We found that Wwox KO mice suffered from severe metabolic defect(s) resulting in growth retardation and all mice died by 3 wk of age. All Wwox KO mice displayed significant hypocapnia suggesting a state of metabolic acidosis. This finding and the known high expression of Wwox in kidney tubules suggest a role for Wwox in acid/base balance. Importantly, Wwox KO mice displayed histopathological and hematological signs of impaired hematopoeisis, leukopenia, and splenic atrophy. Impaired hematopoeisis can also be a contributing factor to metabolic acidosis and death. Hypoglycemia and hypocalcemia was also observed affecting the KO mice. In addition, bone metabolic defects were evident in Wwox KO mice. Bones were smaller and thinner having reduced bone volume as a consequence of a defect in mineralization. No evidence of spontaneous neoplasia was observed in Wwox KO mice. We have generated a new mouse model to inactivate the Wwox tumor suppressor gene conditionally. This will greatly facilitate the functional analysis of Wwox in adult mice and will allow investigating neoplastic transformation in specific target tissues

Peripheral T Cell Cytokine Responses for Diagnosis of Active Tuberculosis

BACKGROUND: A test for diagnosis of active Tuberculosis (TB) from peripheral blood could tremendously improve clinical management of patients. METHODS: Of 178 prospectively enrolled patients with possible TB, 60 patients were diagnosed with pulmonary and 27 patients with extrapulmonary TB. The frequencies of Mycobacterium tuberculosis (MTB) specific CD4(+) T cells and CD8(+) T cells producing cytokines were assessed using overnight stimulation with purified protein derivate (PPD) or early secretory antigenic target (ESAT)-6, respectively. RESULTS: Among patients with active TB, an increased type 1 cytokine profile consisting of mainly CD4(+) T cell derived interferon (IFN)-γ was detectable. Despite contributing to the cytokine profile as a whole, the independent diagnostic performance of one cytokine producing T cells as well as polyfunctional T cells was poor. IFN-γ/Interleukin(IL)-2 cytokine ratios discriminated best between active TB and other diseases. CONCLUSION: T cells producing one cytokine and polyfunctional T cells have a limited role in diagnosis of active TB. The significant shift from a "memory type" to an "effector type" cytokine profile may be useful for further development of a rapid immune-diagnostic tool for active TB

WWOX sensitises ovarian cancer cells to paclitaxel via modulation of the ER stress response

There are clear gaps in our understanding of genes and pathways through which cancer cells facilitate survival strategies as they become chemoresistant. Paclitaxel is used in the treatment of many cancers, but development of drug resistance is common. Along with being an antimitotic agent paclitaxel also activates endoplasmic reticulum (ER) stress. Here, we examine the role of WWOX (WW domain containing oxidoreductase), a gene frequently lost in several cancers, in mediating paclitaxel response. We examine the ER stress-mediated apoptotic response to paclitaxel in WWOX-transfected epithelial ovarian cancer (EOC) cells and following siRNA knockdown of WWOX. We show that WWOX-induced apoptosis following exposure of EOC cells to paclitaxel is related to ER stress and independent of the antimitotic action of taxanes. The apoptotic response to ER stress induced by WWOX re-expression could be reversed by WWOX siRNA in EOC cells. We report that paclitaxel treatment activates both the IRE-1 and PERK kinases and that the increase in paclitaxel-mediated cell death through WWOX is dependent on active ER stress pathway. Log-rank analysis of overall survival (OS) and progression-free survival (PFS) in two prominent EOC microarray data sets (Tothill and The Cancer Genome Atlas), encompassing ~800 patients in total, confirmed clinical relevance to our findings. High WWOX mRNA expression predicted longer OS and PFS in patients treated with paclitaxel, but not in patients who were treated with only cisplatin. The association of WWOX and survival was dependent on the expression level of glucose-related protein 78 (GRP78), a key ER stress marker in paclitaxel-treated patients. We conclude that WWOX sensitises EOC to paclitaxel via ER stress-induced apoptosis, and predicts clinical outcome in patients. Thus, ER stress response mechanisms could be targeted to overcome chemoresistance in cancer

WWOX protein expression varies among ovarian carcinoma histotypes and correlates with less favorable outcome

BACKGROUND: The putative tumor suppressor WWOX gene spans the common chromosomal fragile site 16D (FRA16D) at chromosome area 16q23.3-24.1. This region is a frequent target for loss of heterozygosity and chromosomal rearrangement in ovarian, breast, hepatocellular, prostate carcinomas and other neoplasias. The goal of these studies was to evaluate WWOX protein expression levels in ovarian carcinomas to determine if they correlated with clinico-pathological parameters, thus providing additional support for WWOX functioning as a tumor suppressor. METHODS: We performed WWOX protein expression analyses by means of immunobloting and immunohistochemistry on normal ovaries and specific human ovarian carcinoma Tissue Microarrays (n = 444). Univariate analysis of clinical-pathological parameters based on WWOX staining was determined by χ(2 )test with Yates' correction. The basic significance level was fixed at p < 0.05. RESULTS: Immunoblotting analysis from normal ovarian samples demonstrated consistently strong WWOX expression while 37% ovarian carcinomas showed reduced or undetectable WWOX protein expression levels. The immunohistochemistry of normal human ovarian tissue sections confirmed strong WWOX expression in ovarian surface epithelial cells and in epithelial inclusion cysts within the cortex. Out of 444 ovarian carcinoma samples analyzed 30% of tumors showed lack of or barely detectable WWOX expression. The remaining ovarian carcinomas (70%) stained moderately to strongly positive for this protein. The two histotypes showing significant loss of WWOX expression were of the Mucinous (70%) and Clear Cell (42%) types. Reduced WWOX expression demonstrated a significant association with clinical Stage IV (FIGO) (p = 0.007), negative Progesterone Receptor (PR) status (p = 0.008) and shorter overall survival (p = 0.03). CONCLUSION: These data indicate that WWOX protein expression is highly variable among ovarian carcinoma histotypes. It was also observed that subsets of ovarian tumors demonstrated loss of WWOX expression and is potentially associated with patient outcome

CXCR4 identifies transitional bone marrow premonocytes that replenish the mature monocyte pool for peripheral responses

It is well established that Ly6C(hi) monocytes develop from common monocyte progenitors (cMoPs) and reside in the bone marrow (BM) until they are mobilized into the circulation. In our study, we found that BM Ly6C(hi) monocytes are not a homogenous population, as current data would suggest. Using computational analysis approaches to interpret multidimensional datasets, we demonstrate that BM Ly6C(hi) monocytes consist of two distinct subpopulations (CXCR4(hi) and CXCR4(lo) subpopulations) in both mice and humans. Transcriptome studies and in vivo assays revealed functional differences between the two subpopulations. Notably, the CXCR4(hi) subset proliferates and is immobilized in the BM for the replenishment of functionally mature CXCR4(lo) monocytes. We propose that the CXCR4(hi) subset represents a transitional premonocyte population, and that this sequential step of maturation from cMoPs serves to maintain a stable pool of BM monocytes. Additionally, reduced CXCR4 expression on monocytes, upon their exit into the circulation, does not reflect its diminished role in monocyte biology. Specifically, CXCR4 regulates monocyte peripheral cellular activities by governing their circadian oscillations and pulmonary margination, which contributes toward lung injury and sepsis mortality. Together, our study demonstrates the multifaceted role of CXCR4 in defining BM monocyte heterogeneity and in regulating their function in peripheral tissues

Evidence that substrate-specific effects of C5 protein lead to uniformity in binding and catalysis by RNase P

The ribonucleoprotein enzyme RNase P processes all pre-tRNAs, yet some substrates apparently lack consensus elements for recognition. Here, we compare binding affinities and cleavage rates of Escherichia coli pre-tRNAs that exhibit the largest variation from consensus recognition sequences. These results reveal that the affinities of both consensus and nonconsensus substrates for the RNase P holoenzyme are essentially uniform. Comparative analyses of pre-tRNA and tRNA binding to the RNase P holoenzyme and P RNA alone reveal differential contributions of the protein subunit to 5′ leader and tRNA affinity. Additionally, these studies reveal that uniform binding results from variations in the energetic contribution of the 5′ leader, which serve to compensate for weaker tRNA interactions. Furthermore, kinetic analyses reveal uniformity in the rates of substrate cleavage that result from dramatic (>900-fold) contributions of the protein subunit to catalysis for some nonconsensus pre-tRNAs. Together, these data suggest that an important biological function of RNase P protein is to offset differences in pre-tRNA structure such that binding and catalysis are uniform