Scatter-plot of the observed haemoglobin and observed haematocrit relations

<p><b>Copyright information:</b></p><p>Taken from "Haemoglobin and haematocrit: the threefold conversion is also non valid for assessing anaemia in malaria-endemic settings"</p><p>http://www.malariajournal.com/content/6/1/166</p><p>Malaria Journal 2007;6():166-166.</p><p>Published online 17 Dec 2007</p><p>PMCID:PMC2222606.</p><p></p> The line of best fit (red) indicates a trend towards higher haemoglobin and haematocrit values, giving the following Equation 1: Estimated Haemoglobin = (Haematocrit + 0.6215)/3.1906

Additional file 1: Figure S1. of Identification of repressive and active epigenetic marks and nuclear bodies in Entamoeba histolytica

The N-terminal region of histone H4 of E. histolytica is conserved among different eukaryotic cells. A) Alignment of the N-terminal region of histone H4 from E. histolytica (Eh), P. falciparum (Pf), Saccharomyces cerevisiae (Sc), Drosophila melanogaster (Dm), and Homo sapiens (Hs). (*). Identical residues; (:) compensatory changes. Three insertions present in the N-terminal region of histone H4 of E. histolytica are indicated in red letters. B) Elimination of the three insertions present in the N-terminal region of histone H4 of Eh (indicated by lines) generates a highly conserved N-terminal region among Eh, Pf, Sc, Dm and Hs. Alignment of the H4 N-terminal region of Eh, Pf, Sc, Dm and Hs with H4 pan-acetylated peptide from Tetrahymena thermophiles and a H4 arginine 3 mono-methylated peptide are shown. Rectangle indicated the most conserved residues identified for both peptides. Lysines and arginine amino acids susceptible to be acetylated are indicated in bold. (DOCX 27 kb

Immunological and Functional Characterization of RhoGDI3 and Its Molecular Targets RhoG and RhoB in Human Pancreatic Cancerous and Normal Cells

<div><p>RhoGDI proteins have been implicated in several human cancers; changes in their expression levels have shown pro- or anti-tumorigenic effects. Pancreatic Ductal Adenocarcinoma (PDAC) is a complex pathology, with poor prognosis, and most patients die shortly after diagnosis. Efforts have been focused on understanding the role of RhoGDI's in PDAC, specially, RhoGDI1 and RhoGDI2. However, the role of RhoGDI3 has not been studied in relation to cancer or to PDAC. Here, we characterized the expression and functionality of RhoGDI3 and its target GTPases, RhoG and RhoB in pancreatic cell lines from both normal pancreatic tissue and tissue in late stages of PDAC, and compared them to human biopsies. Through immunofluorescences, pulldown assays and subcellular fractionation, we found a reduction in RhoGDI3 expression in the late stages of PDAC, and this reduction correlates with tumor progression and aggressiveness. Despite the reduction in the expression of RhoGDI3 in PDAC, we found that RhoB was underexpressed while RhoG was overexpressed, suggesting that cancerous cells preserve their capacity to activate this pathway, thus these cells may be more eager to response to the stimuli needed to proliferate and become invasive unlike normal cells. Surprisingly, we found nuclear localization of RhoGDI3 in non-cancerous pancreatic cell line and normal pancreatic tissue biopsies, which could open the possibility of novel nuclear functions for this protein, impacting gene expression regulation and cellular homeostasis.</p></div

Cancerous and non-cancerous pancreatic cell lines show different expression patterns of RhoGDI3 protein.

<p>(A) Immunofluorescence microscopy analysis of RhoGDI3 protein (green); 58 kDa protein, Golgi apparatus marker (Red) and Nuclei (DAPI, blue) was performed on hTERT-HPNE (upper panel), BxPC3 (middle panel) and PANC-1 (bottom panel) cells lines. (B) Representative Immunoblot using antibodies anti-RhoGDI3, anti-RhoGDI2 and anti-GAPDH were used as loading control. Total lysates from hTERT-HPNE, BxPC3 and PANC-1 cell lines were analyzed. (C) Densitometric analysis of the bands detected in the Western blots of RhoGDI3 (n = 3) of protein extracts from all three cell lines, the data was normalized to GAPDH. Densitometric analysis was determined with Image Lab software. Values are means ± SEM, **P<0.005 (Anova-test). Scale bar 20μm.</p

Nuclear localization of RhoGDI3 in rhEGF treated hTERT-HPNE cells.

<p>Subcellular fractionation was performed after cells were treated with rhEGF (marked above the images as 0, 2 and 10 rhEGF Min). Nuclear (N) and cytosolic (C) fractions from hTERT-HPNE (A), BxPC3 (B) and PANC-1 (C) cell lines were obtained and analyzed by immunoblotting, using anti-RhoGDI3, anti-RhoG, anti-RhoB antibodies. Anti-histone H3 antibody was used as a nuclear control and anti-Aldolase B antibody was used as a cytosol control. 20 μg of cell lysates were loaded.</p

Nuclear localization of RhoGDI3 in normal pancreatic tissue.

<p>Immunofluorescence microscopy staining of RhoGDI3 (green) and RhoG (red) was carried out on human pancreatic normal (A) and moderate (aggressiveness) PDAC biopsies (C). (B) Magnification and lateral view of the immunofluorescence of RhoGDI3 and RhoG, to evidence nuclear localization in human pancreatic normal tissue. Arrowheads denote the localization of RhoGDI3 and RhoG into the nuclei. Scale bar 10 = μm.</p

Activation of RhoG GTPase in hTERT-HPNE, BxPC3 and PANC-1 pancreatic cell lines.

<p>Cells were starved 6 hours and confronted with rhEGF for the period of 0, 2 and 10 minutes (Marked as 0, 2 and 10). Fluorescence microscopic staining of RhoG (green) was carried out in hTERT-HPNE (A), BxPC3 (B) and PANC-1 (C) cells lines. To show the cytoskeleton reorganization, F-Actin was stained with rhodamine phalloidin. Measurement of RhoG activity was performed using RhoG pulldown assay. Immunoblots for RhoG, Rac-1 and GAPDH proteins for hTERT-HPNE (D), BxPC3 (E) and PANC-1 (F) are shown. To quantify the amount of RhoG-GTP and bound-Rac-1 through the temporal course, densitometric analysis was performed using Image Lab software, hTERT-HPNE (G), BxPC3 (H) and PANC-1 (I). For comparison of RhoG activity, the total amount of RhoG in cell lysates was normalized to total RhoG. GAPDH was used as a protein loading control. ELMO1-GST beads coomassie are shown as beads loading control. Arrowheads denote the localization of RhoG into the peripheral membrane; boxes with number represent the number of cells with this phenotype. Scale bar 100 μm.</p

GTPase RhoB shows differential activation in PDAC cell lines.

<p>Cells were starved for 6 hours and treated with rhEGF for a period of 0, 2 and 10 minutes (Marked as 0, 2 and 10 min). An immunofluorescence microscopy analysis of RhoB (green) was carried out on hTERT-HPNE (A), BxPC3 (B) and PANC-1 (C) cells lines. To show the cytoskeleton reorganization, F-Actin was stained with rhodamine phalloidin. Measurement of RhoB activity was performed using RhoB pulldown assay. Immunoblots for RhoB and GAPDH, as loading control for hTERT-HPNE (D), BxPC3 (E) and PANC-1 (F) cell lines are shown. To quantify the amount of RhoB-GTP, densitometric analysis (n = 3) was performed using Image Lab software for samples of hTERT-HPNE (G), BxPC3 (H) and PANC-1 (I) cell lines. For comparison of RhoB activity, GTP-RhoB was normalized to total RhoB. GAPDH was used as a protein loading control. Coomassie of RBD-GST beads are shown as beads loading control. Arrowheads denote the localization of RhoB into the peripheral membrane; boxes with number represent the quantity of cells per field with this phenotype. Scale bar = 100 μm.</p

The expression of RhoG and RhoB proteins is altered in cancerous pancreatic cell lines.

<p>Immunofluorescence microscopy analysis of RhoGDI3 protein (green); RhoG (A) and RhoB (B) (Red) and Nuclei (DAPI, blue) of hTERT-HPNE (upper panel), BxPC3 (middle panel) and PANC-1 (bottom panel) cells lines. Representative Immunoblot using antibodies anti-RhoG (C), anti RhoB (D), GAPDH was used as loading control. Total lysates from hTERT-HPNE, BxPC3 and PANC-1 cell lines were analyzed. Total amount of RhoG (E) and RhoB (F) proteins was normalized to GAPDH (n = 3). Immunoblot densitometric analysis was performed with Image Lab software. Values are means ± SEM, **P<0.005, *P<0.005 (Anova-test). Scale bar 10μm.</p