EMBO/sd-character-level-ner huggingface dataset:
Figure 1A) oxLDL plasma concentration in ACM patients and HC (n=36; Mann-Whitney test). B) oxLDL plasma concentration in mutated ACM (n=7) and NON ACM relatives, carriers of the same causative mutation (n=9; Mann-Whitney test). C) oxLDL plasma concentration in ACM patients carriers of a PKP2 mutation and ACM patients carriers of other desmosomal or non desmosomal mutations, or gene elusive (n=10 vs. n=26; One-Way ANOVA). Data information: mean ± SEM. * p<0.05; ** p<0.01; *** p<0.001. D) Representative images of MDA immunostaining (green) on ACM and HC ventricular tissue sections and relative quantification (n=4 biological replicates; Two-tailed Student's t-test). Nuclei are counterstained with Hoechst33342 (blue). E) Representative images of CD36 immunostaining on ACM and HC ventricular tissue sections and relative quantification (n=4 biological replicates; Two-tailed Student's t-test). Nuclei are counterstained with Hoechst33342 (blue). Data information: mean ± SEM. * p<0.05; ** p<0.01; *** p<0.001.
Figure 1(a) C57Bl/6 mice were infected (n=9) or not (n=8) intranasally with 150 pfu of influenza A virus (IAV; A/Scotland/20/74 (H3N2)). Bronchoalveolar lavage fluids (BALs) were collected 4 days post-infection. Samples were further processed for metabolomic analysis by NMR. Left panel shows a volcano-plot representing the comparison of "IAV-infected" versus "control, non-infected" animals. The graph plots the -log (p-value) against the fold change (log2 (ratio)) for individual metabolites. Vertical dashed blue lines mark the two-fold change and horizontal dashed blue line marks the cut-off p-value of 0.0001. Metabolites in the upper right square in red are significantly more abundant in "IAV-infected" compared to "control, non-infected" samples. Succinate is one of these compounds with a FC=2.2 and a FDR-adjusted p-value=8.10-7. The boxplots whiskers correspond to the minimum and maximum values, while the central band is the mean. The first and third quartiles (boxes) are merely the average values between the median and the extrema. Data information: Data are mean ± SEM and statistical analysis was performed using the Mann-Whitney U-test. (b) shows succinate quantification by NMR from tracheal aspirates collected in mechanically-ventilated patients with (pink bar, n=9) or without (orange bar, n=7) diagnostic of IAV pneumonia. *P < 0.05. (c) IL-6 and IL-8 measurements by ELISA from these same tracheal aspirates. Data information: Data are mean ± SEM and statistical analysis was performed using the Mann-Whitney U-test. (d) Main baseline characteristics of the patients included in the study.
Figure 5(A) Auxin levels in root tips, as indicated by the ProDR5:GFP reporter line. Seeds were germinated on HK medium for 4 d, then the seedlings were transferred to LK or HK medium for the indicated times. Scale bar, 10 μm. (B) Quantification of GFP fluorescence in the QC cells shown in (A). Data are means ± SE (n = 20, individual plants). Student's t-test (**P < 0.01) was used to analyze statistical significance.
a) HeLa.figf3(a) HeLa.Vec and HeLa.UVRAG cells transfected with HA-Vps16 and GFP-LC3 were treated with 2 μM rapamycin for 2 h, and processed for confocal microscopy (left panel; scale bars, 5 μm). The percentage of GFP-LC3 punctae positive for HA-Vps16 staining was quantified (right panel; data are mean ± s.e.m., n = 60, *P 0.01). (b) Effects of Vps16 on UVRAG-mediated autophagosome formation. Light microscopic quantification of autophagosomes in HeLa.Vec and HeLa.UVRAG cells (upper left panel) or in HCT116.Vec and HCT116.UVRAG cells (bottom left panel) when transfected with GFP-LC3 together with control siRNA or Vps16 siRNA (data are mean ± s.e.m., n = 60 for HeLa cells; n = 450 for HCT116 cells, *P 0.01; **P 0.001). Immunoblotting of Vps16 and tubulin are shown in the right panel.
Figure 1A Protein and genetic structure including the different gephyrin domains and genomic location of the gene with an indication of the different exons. The position of the mutation identified in this study is indicated on the protein level, affecting the E-domain and on the genomic level, affecting exon 12. B Multiple alignment of gephyrin and homologous proteins from various species from different kingdoms of life (aligned using Clustal Omega). C Co-crystal structure of gephyrin E-domain (blue) and bound GlyR ß-loop (red - PDB code 2FTS (Kim et al., 2006)). The mutated gephyrin Gly375 residue is shown in orange.
HBV induces mitochondrial translocation of Parkin and stimulates mitophagy-related genes.Figure 2(A) Huh7 cells transfected with HBV construct were prestained with MitoTracker (Mito, red) and subsequently, immunostained with anti-Parkin (green) and anti-HBsAg (white) antibodies. Nuclei are demarcated with white dots. Transfected (+) and untransfected (−) cells are marked. In the zoomed images, yellow color indicates the accumulation of endogenous Parkin recruited to the mitochondria. Quantification of fluorescence intensity of Parkin aggregates on the mitochondria in HBV-expressing cells. (B) The cytosolic (Cyto) and mitochondrial (Mito) fractions were isolated from HepAD38 cells grown in the absence or presence of tetracycline for 48 h. Cellular fractions were analyzed by Western blotting with antibodies specific for the indicated proteins. Fractions: WCL, whole cell lysates; Cyto, purified cytoplasm; Mito, purified mitochondria. Organelle markers: VDAC1, mitochondria; GAPDH, cytoplasm. (C) Parkin protein in whole cell lysates of HBV-expressing cells was immunoprecipitated by anti-Parkin antibody, followed by immunoblotting (IB) with anti-ubiquitin (Ub) antibody. Normal rabbit IgG was used as a control for immunoprecipitation (IP). Western blot analysis for β-actin indicates equivalent amount of cell lysates for IP (C and D). (D) Mfn2 protein in whole cell lysates extracted from HepAD38 cells transfected with non-targeting (NT) and Parkin (P) siRNA, respectively, for 48 h was immunoprecipitated by anti-Mfn2 antibody, followed by immunoblotting (IB) with anti-ubiquitin (Ub) antibody. Normal mouse IgG was used as a control for immunoprecipitation (IP). The protein expression was analyzed by Western blotting with antibodies specific to Mfn2, Parkin, and β-actin proteins. ( (E) Intracellular mRNA levels of Parkin, PINK1, and LC3B were analyzed by real-time qRT-PCR. GAPDH was used to normalize changes in Parkin, PINK1, and LC3B gene expression. (F) The protein expression was analyzed by Western blotting with antibodies specific for the indicated proteins. β-actin was used as a loading cont(F) The protein expression was analyzed by Western blotting with antibodies specific for the indicated proteins. β-actin was used as a loading control (F and G). The relative intensity of ATF4, Parkin, PINK1, and LC3B protein expression normalized to β-actin was analyzed by ImageJ software. . (G) Whole cell lysates extracted from Huh7 cells with empty vector (Mock) and those with HBV construct for 48 h were analyzed by Western blotting with antibodies specific to the indicated proteins.
Figure 8A. HeLa cells were transfected with two different siRNAs against NSUN3 (siNSUN3_1, siNSUN3_2), ABH1 (siABH1_1, siABH1_2) or with nontarget (siNT) siRNA and the knockdown efficiency was analysed by quantitative PCR. The relative abundance of the NSUN3 or ABH1 mRNA was normalized to GAPDH levels. Data are presented as mean ± SD. B. Chemically synthesised f5C modified mt-tRNAMet and total RNA from wild-type (WT) cells or those transfected with siRNAs as in (A) were treated with TMI to specifically label f5C residues. Primer extension, using a radiolabelled antisense primer, was performed under limited dNTP conditions. Products were separated on a denaturing polyacrylamide gel alongside a sequencing ladder and RNAs were detected using a phosphorimager. C. Primer extension reactions were performed on total RNA from cells transfected with siRNAs as described in B. Stops corresponding to position C34 in mt-tRNAMet were quantified in three independent experiments and results are shown graphically as mean ± SD. D. RNA from wild-type HeLa cells and cells treated with siRNAs against NSUN3 or ABH1 (as in A) was either first reduced with NaBH4 or directly treated with bisulfite. After deamination and desulfonation, mt-tRNAMetRNAs were reverse transcribed, amplified, cloned and sequenced. The proportions of thymine (grey) generated by bisulfite-conversion or non-converted cytosine (black) at position 34 of mt-tRNAMet are shown. Note that for sequences from non-reduced samples, thymine can originate from unmodified or f5C-containing mt-tRNAMet, while in reduced samples it originates from unmodified cytosine.
a,b) Constructs identical to those described in a, but lacking mRFP (representations of S, HS, Q, HQ are shown in Supplementary Fig.figf4(a,b) Constructs identical to those described in Figure 2a, but lacking mRFP (representations of S, HS, Q, HQ are shown in Supplementary Fig. 3a) were co-transfected with tNHTT-16Q-EGFP (a) and tNHTT-150Q-EGFP (b) constructs into Neuro2a cells (bar, 4 μm). (c) Dot blot analysis of the lumenal fraction of purified lysosomes from Neuro2a cells co-transfected with tNHTT-60Q-EGFP and R, RS, RHS, RQ or RHQ. PNS, post-nuclear supernatant. (d) Quantification of HTT translocation into lysosomes. Levels of HTT detected by EM48 (anti-huntingtin antibody) were normalized to levels of β-tubulin (in PNS) and levels of cathepsin D (in lysosomes). (e) Compound images generated by ArrayScan illustrating inclusion formation in 150Q Neuro2a cells transfected with R, RS, RHS, RQ and RHQ and shRNA specific for HSC70 and/or Lamp2a. (blue, Hoechst 33258; green, tNHTT-150Q-EGFP; red, tested molecule). (f) RNAi of HSC70, Lamp2a or both alleviated the inhibitory effect of RHQ on the formation of inclusions containing tNHTT-150Q-EGFP inclusions. (g) Western blot analysis of soluble polyQ protein in 150Q Neuro2a cells transfected with tested constructs and shRNA for HSC70 and/or Lamp2a. Full-length western blots are presented in Supplementary Figure 13. (h) Silencing of HSC70, Lamp2a or both alleviated the inhibitory effect of RHQ on levels of soluble tNHTT-150Q-EGFP. Bars in d and f represent mean values ± s.e.m. from three independent experiments. Bars in h represent relative mean values ± s.e.m. from three independent experiments. For h, values for the R construct were arbitrarily set to 1.
KSHV dysregulates inhibitory miRNA(A) Heat map of differentially expressed miRNAs during KSHV replication at 0, 16 and 24 hour identified through miR-Seq.Figure 1(A) Heat map of differentially expressed miRNAs during KSHV replication at 0, 16 and 24 hour identified through miR-Seq. miR-29b and miR-30c are highlighted in red, with T test analysis. (B) qPCR analysis of miR-29b and miR-30c levels in cells at 0 and 24 hours post-lytic induction. SNORD68 was used as a housekeeper (n=3). Data information: data are presented as mean ± SD. **P<0.01, ***P<0.001 (Unpaired Student's t-test). All repeats are biological. (C, D) qPCR analysis of TREx 0 and 24 post-induction (n=3) using GAPDH as a housekeeper, analysis is of levels of pri-miR-29b/pri-miR-30c (C) and pri+pre-miR-29b/pri+pre-miR-30c (D). Data information: data are presented as mean ± SD. **P<0.01, ***P<0.001 (Unpaired Student's t-test). All repeats are biological. (E) Representative western blot of scr or miR-30c transfected TREx cells analysed for ORF65 expression with GAPDH as a loading control. Densitometry analysis performed on n=3. Data information: data are presented as mean ± SD. **P<0.01, ***P<0.001 (Unpaired Student's t-test). All repeats are biological. (F) Representative western blot of scr or miR-29b transfected TREx cells analysed for ORF65 expression with GAPDH as a loading control. Densitometry analysis performed on n=3. Data information: data are presented as mean ± SD. **P<0.01, ***P<0.001 (Unpaired Student's t-test). All repeats are biological. (G) qPCR analysis of ORF57 DNA levels for viral load at 72 hours post induction, with scr and miR-30c transfected TREx cells with uninduced cells as a control and GAPDH as a housekeeper (n=3). Data information: data are presented as mean ± SD. **P<0.01, ***P<0.001 (Unpaired Student's t-test). All repeats are biological. (H) qPCR analysis of ORF57 in HEK-293T cells for reinfection assay. Data information: data are presented as mean ± SD. **P<0.01, ***P<0.001 (Unpaired Student's t-test). All repeats are biological.
Figure 4A Heavy membrane (HM) fractions of CHLA15 were rendered MAM-depleted by immunomagnetic separation to derive purified mitochondria (pMito), and cytochrome c release measured in response to BimBH3 peptide. Data information: data points are mean of duplicate wells (SD<0.05 at all points) in a representative experiment from at least two biological replicates B, C, D TEM analyses were used to quantify mitochondrial size, circularity and roundness in CHLA15 cells transfected with a sh-control (Ctrl), shMFN2 or shPACS2 constructs (mean +/- SD shown). Data information: n=214-246 mitochondria per cell line. statistical analyses were performed using a two-tailed Mann-Whitney U test, with significance p<0.05. E Immunoblot assessment of Mfn2 and Pacs2 protein knockdown. F Proportion of mitochondria with 0 to 5 MAMs are shown for each. Data information: n=214-246 mitochondria per cell line. G Percentage of mitochondria perimeter with an apposed ER within defined gap-widths; dotted-lines denote +10% and -10% change. Data information: , n=214-246 mitochondria per cell line. H In vitro viability of CHLA15-ctrl, CHLA15-shMFN2, CHLA15-shPACS2, and CHLA20 cells following 72hr exposure to ABT-737; dotted-line represents 50% viability. Data information: data points are mean and SD from triplicate wells, experiments are representative of at least two biological replicates. I Mitochondrial cytochrome c release in response to tBid and BimBH3 peptide in CHLA15-Ctrl, CHLA15-shMFN2 and CHLA15-shPACS2 cells. Data information: , data points are mean of duplicate wells (SD<0.05 at all points) in a representative experiment from at least two biological replicates
Fig.Figure 1A) Visualisation of the pathology load at 9M of age of TAUwt, TAUtg and APPtg mice at 9M of age. Immunofluorescent staining for X34 (a fluorescent derivative of Congo Red), Iba1 (microglia) and TO-PRO-3 (nuclei) has been pseudocolored. Scale bar = 100μm. B) Experimental design for mRNA sequencing using n=12 per experimental group. C) Explanation of the 2x2 linear model, where those cells labeled with 1 are compared to the cells labeled with 0. In the age comparison, mRNA expression in all 10-month-old (10M) mice is compared to all 4-month-old (4M) mice. In the genotype comparison, mRNA expression in all transgenic (TG) mice is compared to all wild-type (WT) mice. In the age*genotype comparison, we assess which transcripts are differentially expressed in the 10M TG mice compared to all other groups. D) Based on Marioni et al. (Marioni et al, 2018) various sets of AD GWAS risk genes were created using different cut-off p-values indicated on the x-axis (number of genes within each set is written in grey). Enrichment for AD risk genes was assessed among the different statistical comparisons for APPtg and TAUtg mice (Int: age*genotype, Gen: genotype), using gene set enrichment analysis (GSEA (Subramanian et al, 2005)). Colors represent -log10 (Benjamini-Yuketieli adjusted p-value) for the enrichment; blank means no significant enrichment.
Figure 8Confocal whole-mount immunofluorescence analysis of phosphorylated Akt (pAkt) (A) and phosphorylated p44/42 MAPK (pErk1/2) (B) in Kit+ cells of wild type (32-36sp) and Sl/Sl (31-35sp) E10.5 dorsal aorta, yolk sac and fetal liver. Images are 2.5μm-thick single longitudinal Z slices; dorsal up (DA). Arrowheads point to Kit+ cells with varying levels of pAkt and pErk1/2 expression. A total of 2 wild type and 2 Sl/Sl embryos for each staining were analyzed. Scale bars: 20μm. Quantification of the immunofluorescence in (A, B), showing mean fluorescence intensity (MFI) ratio of pAkt and p44/42 MAPK (pErk1/2) within wild type (+/+) and Sl/Sl CD31+ Kit+ dorsal aorta (DA), yolk sac (YS) and fetal liver (FL) hematopoietic cells. MFI values were normalized to background in the adjacent ventral mesenchyme for DA, to erythrocytes in the YS or stroma in the FL. Additional quantification was performed on sections from 2 wild type and 2 Sl/Sl embryos of similar age range. MFI measurements were taken using ImageJ. Number of cells measured for pAkt: 100 hematopoietic cells and 73 background (+/+); 79 hematopoietic and 52 background (Sl/Sl) in the AGM; 127 hematopoietic cells and 95 background (+/+); 110 hematopoietic and 82 background (Sl/Sl) in the YS; 102 hematopoietic cells and 78 background (+/+); 78 hematopoietic and 83 background (Sl/Sl) in the FL. Number of cells measured for pErk1/2: 94 hematopoietic cells and 82 background (+/+); 114 hematopoietic and 91 background (Sl/Sl) in the AGM; 174 hematopoietic cells and 109 background (+/+); 123 hematopoietic and 82 background (Sl/Sl) in the YS; 100 hematopoietic cells and 70 background (+/+); 63 hematopoietic and 57 background (Sl/Sl) in the FL. N=4 (+/+), 4 (Sl/Sl) embryos analyzed for DA and YS; N=3 (+/+), 2 (Sl/Sl) for FL. Error bars represent SD. Multiplex Fluidigm qRT-PCR analysis of pre-HSC type I (Ter119- VE-Cadherin+ Kit+ CD41+ CD45-), pre-HSC type II/HSC (Ter119- VE-Cadherin+ Kit+ CD41+ CD45+), YS and FL EMP (Ter119- Kit+ CD41+ CD16/32+), FL Early E (Ter119- Kit+ CD41-) and peripheral blood LMPP (Lin- CD19- B220- CD45+ Kit+ Flt3+ IL7Rα+), isolated from E11.5 (11-17 tail somites) wild type and Sl/Sl embryos. 4 biological replicates for wild type (9 total embryos) and 5 biological replicates (9 total embryos) for Sl/Sl were analyzed. Data are mean (±SD).
Virally expressed KSHV‐TK induces cell contraction and loss of focal adhesionsA schematic illustrating the BAC strategy used to generate a recombinant MuHV‐4 virus (MuHV‐4 [g‐KSHV‐TK]) in which the endogenous MuHV‐4 TK is exchanged for GFP‐KSHV‐TK.Figure 3A A schematic illustrating the BAC strategy used to generate a recombinant MuHV‐4 virus (MuHV‐4 [g‐KSHV‐TK]) in which the endogenous MuHV‐4 TK is exchanged for GFP‐KSHV‐TK. B Immunoblot analysis of BHK cells infected with MuHV‐4 and MuHV‐4 [g‐KSHV‐TK] or uninfected cells using antibodies against phosphotyrosine (p‐Tyr), GFP, MuHV‐4 TK, ORF17 (capsid) and glycoprotein N. C Immunofluorescence microscopy analysis of MuHV‐4‐ and MuHV‐4 [g‐KSHV‐TK]‐infected cells with the indicated antibodies reveals GFP‐KSHV‐TK expressed in the context of lytic infection (MuHV‐4 [g‐KSHV‐TK]) induces the loss of phospho‐FAK Y925 and paxillin Y31 from focal adhesions. White arrowheads indicate examples of infected cells expressing GFP‐KSHV‐TK or GFP (indicating WT MuHV‐4 infection, BAC+ virus). D Immunofluorescence microscopy analysis reveals that GFP‐KSHV‐TK expressed in the context of lytic infection (MuHV‐4 [g‐KSHV‐TK]) does not alter focal adhesion integrity (paxillin) in the presence of the ROCK inhibitor Y‐27632. E Immunofluorescence microscopy analysis of paxillin in the lytically induced rKSHV.219/Vero cell line in the presence or absence of Y‐27632.
Figure 2(A) Isolation of the STC elicited by the last pulse of a high-frequency train of action potentials. Left: superimposed representative traces of the inward current evoked in an astrocyte (held at -90 mV) by extracellular stimulation with a train of 10 pulses (trace a: black) and a train of 9 pulses (trace b: brown) at 50 Hz in a WT cortical slice. The inward current elicited by the 10th pulse was obtained by subtracting the current elicited by 9 pulses from that elicited by 10 pulses (trace a-b). Right: The STC elicited by the 10th pulse (trace a-b-c) was obtained by subtracting the exponential function that simulates the TBOA-insensitive current elicited by a single pulse (trace c, obtained as in Figure 1C) to the 10-9 pulses difference current (trace a-b). The decay of the STC elicited by the 10th pulse was best fitted by a single exponential function with τdecay = 8.04 ms (in red). (B) τdecay of the STC elicited by the 10th pulse of 50 Hz (τ10 (50 Hz), left panel) and 100 Hz (τ10 (100 Hz), right panel) trains in layer 1 astrocytes in acute cortical slices from P22-23 WT (n = 23; N=10 for 50 Hz; n = 18; N=8 for 100 Hz) and KI (n = 21; N=9 for 50 Hz and n = 14; N=7 for 100 Hz) mice. The STC elicited by the 10th pulse was obtained as described in (A). STC τ10 (50 Hz) and τ10 (100 Hz) are 30% and 37% higher in KI compared to WT mice, respectively (unpaired t-test: P < 0.0001 in both cases).
| Comprehensive target methylome analysis identified more hypo-methylated genomic regions in sperm from aged mice.Figure 2A. Strategy for DNA target methylome analyses of sperm from young (3-month-old, n=4) and aged (18-month-old, n=9) mice. B. The average level of CpG methylation at the targeted regions was 33.3%-34.6% and 27.7%-36.2% in sperm datasets from young and aged mice, respectively. The average depth of each sample was 11.3-27.6, which covered 44.2%-62.9% of the target regions at a depth of 10. C. Correlation coefficients were also high (i.e., 0.98-0.99), indicating that the obtained sequence data was sufficient for further analysis of age-specific methylation changes. D. A heat map of the 16 hyper- and 96 hypo-methylated DMRs in sperm from aged mice ordered by their chromosomal positions. M indicates the average. E. A majority of hyper-methylated genomic regions were within the genes (31.9% in exons and 48.8% in introns in count-base analysis and 50.0% in exons and 31.2% in introns in length-base analysis). In contrast, almost half (47.9%) of the hypo-methylated regions were in intergenic regions (51.3% in count-base analysis and 47.9% in length-base analysis); only 4.3% of the regions in count-base analysis and 5.2% in length-base analysis were in exons, and 3.1% in count-base analysis and 11.5% in length-base analysis were in promoter regions (0.5 kb or 2.0 kb upstream of the transcription starting site).
Figure 6A Microarray analysis of IL-33 treated BM-derived eosinophils. Eosinophils from Dusp5+/+ and Dusp5-/- mice were treated for 4 hours with IL-33 (50 ng/ml). Total RNA from independent BM-derived eosinophils cultures derived from 5 mice for each genotype was quantitated by microarray analysis. Genes differentially up- (24 genes) or down-regulated (62 genes) in Dusp5+/+ compared to Dusp5-/- eosinophils by >1.5-fold with a P value <0.05 are highlighted in red or blue, respectively. B Dusp5+/+Rag2-/- or Dusp5-/-Rag2-/- mice were treated with daily intraperitoneal injections with IL-33 (500 ng) for 6 days. Eosinophils were harvested on day 6 one hour following the final dose of IL-33. Total RNA was harvested and Il4, Il13 and TNFa mRNA expression was assayed by Taqman Realtime PCR analysis. Data are represented as fold expression of values using the 2-delta Ct method normalized to Gapdh mRNA levels. Results are representative of three independent experiments. C Dusp5+/+ (open circles) and Dusp5-/- (grey squares) BM-derived eosinophils were unstimulated or stimulated with IL-33 (25 ng/ml) for 24 hours. Culture supernatants were analyzed for IL-4 (top), IL-13 (middle) and TNF-a (bottom) levels by ELISA. Results are representative of three independent experiments. D RT-PCR analysis of Fosl1 expression in Dusp5+/+ and Dusp5-/- BM-derived eosinophils stimulated with IL-33 (50 ng/ml) for 4 hours. Results are representative of three independent experiments. E Cell surface expression of CD11b was assessed on unstimulated or IL-33 (50 ng/ml) treated Dusp5+/+ (red) or Dusp5-/- (blue) BM-derived eosinophils for 20 hours (left). Percentage of CD11bhi surface expression of representative FACS analysis (left) was quantitated (right) from a total of three independent samples.F Cell surface expression of CD69 was assessed on unstimulated or IL-33 (50 ng/ml) treated Dusp5+/+ (red) or Dusp5-/- (blue) BM-derived eosinophils for 20 hours (left). Quantitation of CD69 surface expression of representative FACS analysis (left) was quantitated (right) from three independent samples.