EMBO/sd-character-level-ner huggingface dataset:
Figure 4A,B EDN target phosphopeptides for (A) UACC257 and (B) A2058 were segregated into six clusters each, using c-means clustering (MFuzz). Each line represents one phosphopeptide and cluster membership is indicated by colour. Most likely kinases for each phosphosite were predicted using NetworKIN and enrichment of kinase substrates was calculated for each cluster. Enriched kinases are indicated above each cluster. Note that substrates of these kinases only account for a fraction of all phosphosites in a cluster. C Average MS1 intensity z-score kinetics for 217 predicted CaMKII-α sites that are shared between both cell lines. For comparison, average (n = 6) calcium abundance kinetics are shown.
Figure 7A. The structural modeling result of RIG-I/dsRNA complex and positions of the lysine residue were presented. RIG-I helicase A and B are shown as green and pale blue ribbons. The dsRNA strands were presented as purple and yellow rods, respectively. Two lysine residues, K907 and K909, on RIG-I that interact with dsRNA are indicated as violet sticks. K907 forms hydrogen bonding with neither 5´-ppp dsRNA nor K907.B, C. Acetylated K909 and K907 on RIG-I are indicated as green sticks. There were hydrogen bonding between NH2 of K907 and oxygen of acetyl-K909.
Figure 6(A) DDL-N but not DDL-C associates with PARP2. FLAG-tagged DDL-N or DDL-C was co-expressed with PARP2-HA in protoplasts for Co-IP assay. Protoplasts were treated without or with 100 nM flg22 for 30 min. (B) in vivo PARylation of DDL-N or DDL-C in protoplasts fed with 32P-NAD+. PARylation was visualized with autoradiography. (C) In vitro PARylation of DDL-N or DDL-C. FLAG-tagged DDL-N or DDL-C was expressed and immunoprecipitated from Arabidopsis protoplasts and subjected to in vitro PARylation by GST-PARP2 with Biotin-NAD+. PARylated proteins were detected by streptavidin-HRP antibody. (D) List of the mass/charge values of b+ ions and y+ ions of the peptide (148AVeEALAAK156). Red mass values correspond to the identified peak in the spectrum. The modified "e" is in bold and lowercase. The mass difference between b2+ and b3+ or y6+ and y7+ indicates that Glu150 is PARylated. (E) MS/MS spectrum of a doubly charged peptide with Glu (the third AA "e" in the peptide, Glu150) PARylation. 2+ indicates doubly charged and 0 indicates loss of H2O. The mass difference between y7+ and y6+ is 144.1, which is about 15 Da bigger than the Glu residue mass of 129.04. (F) DDL12E mutants substantially diminish its PARylation in vivo. In vivo PARylation of DDL12E1 or DDL12E2 was performed using Arabidopsis protoplasts fed with 32P-NAD+. (G) Reduced association of DDL12E1 with PARP2 in protoplasts. Protoplasts were treated with 100 nm flg22 for 30 min before Co-IP assay.
Figure 6The molecular structure of EFV. Intracellular concentrations of cholesterol in GBM cells after treatment with 20 μM EFV or DMSO and normalized to total protein. Data are shown as the mean ± SEM (n = 3). LN229: *P = 0.013; GBM#P3: **P = 0.0023. Filipin staining of GBM cells after treatment with EFV (0 - 20 μM). Scale bar = 30 μm. IC50 curves for EFV in GBM cells determined using OD readings (450nm) from the CCK-8 assay. 24OHC levels in spent media from LN229 (left) or GBM#P3 (right) cells treated with DMSO or 20 μM EFV for 72 h quantified using targeted LC-MS/MS. Images and quantification of colonies formed by GBM cell lines after treatment with different concentrations of EFV (0 - 20 μM). Data are shown as the mean ± SEM (n = 3). LN229: **P = 0.0056, ***P < 0.0001; LN18: ***P < 0.0001. Flow cytometry to detect annexin V-FITC and PI staining to determine the percentage of GBM cells undergoing apoptosis after treatment with EFV (0 - 20 μM) for 72 h. Data are shown as the mean ± SEM (n = 3). LN229: ***P = 0.0006, ***P < 0.0001; GBM#P3: ***P < 0.0001. Tumoursphere formation assays for GSCs treated with different concentrations of EFV (0 - 20 μM). Scale bar = 100 μm. Graphic representation of the quantification of tumoursphere formation (right). Data are shown as the mean ± SEM (n = 3). GBM#P3: **P = 0.0011, ***P = 0.0003, *** P < 0.0001; GBM#BG7: ***P < 0.0001; GBM#BG5: **P = 0.0037, **P = 0.0015, ***P = 0.0002. Western blot analysis of cholesterol homeostasis related proteins (LDLR, ABCA1, P-SREBP1 and N-SREBP1), c-PARP, PCNA and SOX2 in GBM cells treated with EFV (0 - 20 μM) for 72 h. Schematic diagram of the schedule for implantation and drug treatment in the GBM xenograft model. Seven days after implantation of tumour cells, mice were treated with EFV by gavage (0.1 mg/kg/day). Bioluminescent imaging (BLI) using IVIS was performed at days 4, 7, 14 and 21. The survival curves of tumour-bearing mice implanted with LN229 or GBM#P3 cells after EFV or PBS treatment (n = 5 per group). A log-rank test was used to assess statistical significance. Bioluminescent images and the corresponding quantification of tumours in mice implanted with GBM#P3 cells at days 7 (n = 5 per group, P = 0.58) and 21 (n = 5 per group, P = 0.005). Statistical significance was determined by two-sided Student's t‐test. Body weight of tumour-bearing mice after 3 weeks of EFV or PBS treatment. Data are shown as the mean ± SEM. Statistical significance was determined by two-sided Student's t‐test.
Accumulation of LC3-II, p62 aggregates, and ubiquitin-positive inclusions in Epg5 knockout mice.Figure 3.(A) Levels of LC3 and p62 in brain and spinal cord extracts from Epg5+/− and Epg5−/− mice. SC, spinal cord. (B) Polyubiquitinated proteins in detergent-soluble (S) and -insoluble (I) fractions from brain and spinal cord homogenates of Epg5+/− and Epg5−/− mice. (C) p62 aggregates and ubiquitin-positive aggregates are absent from Epg5+/− mice but dramatically accumulate and are colocalized (arrows) in Epg5−/− mice. (D-F) p62 aggregates are detected in neurons (arrows; D) and oligodendrocytes (arrows; E) but absent from astrocytes (F) in Epg5−/− mice. (G) Cytoplasmic TDP-43 aggregates accumulate in motor neurons in the anterior horn of spinal cord in Epg5−/− mice. Arrows show cytoplasmic TDP-43 aggregates. DSCT, dorsal corticospinal tract. (H) Percentages of cytoplasmic TDP-43 aggregate-positive motor neurons in the fifth layer of cortex and anterior horn of the spinal cord in Epg5+/− and Epg5−/− mice. Means ± SEM of three mice are shown. LC, lateral column. Bars, 10 µm.
Figure 6(A) Effect of flg22 on the NbFLS2-GmLMM1 interaction. The indicated constructs were transiently expressed in N. benthamiana by Agrobacterium-mediated transient expression for 2 days, and the leaves were treated with flg22 (1 μM) or water for 10 min before being subjected to Co-IP assay. Data information: The experiments were performed three times , as biological replicates, with similar results. (B) Regulation of NbBAK1-GmLMM1 interaction by flg22. Data information: The experiments were performed three times as biological replicates, with similar results. (C) Suppression of flg22-induced NbFLS2-NbBAK1 interaction by GmLMM1. NbFLS2 and NbBAK1 were co-expressed in N. benthamiana along with GmLMM1 or GmLMM1L407H, treated with H2O or flg22 and subjected to co-IP assays. Data information: The experiments were performed three times as biological replicates, with similar results. (D Suppression effect of GmLMM1 on the flg22-induced interaction of GmFLS2 with GmBAK1a (D) Data information: The experiments were performed two times (D as biological replicates, with similar results. E) Suppression effect of GmLMM1 on the flg22-induced interaction of GmFLS2 with GmBAK1b (E). Data information: The experiments were performed two times E), as biological replicates, with similar results. (F) A model chart illustrating GmLMM1 mediation of PTI signaling balance in soybean. Flg22 induces the FLS2-BAK1 interaction to trigger downstream immune signaling. GmLMM1 constitutively interacts with BAK1 both before and after immune activation. GmLMM1 can suppress the FLS2-BAK1 interaction to control an appropriate FLS2-BAK1 interaction level. Thus, GmLMM1 works as a molecular switch to regulate moderate immune activation.
Figure 5(A) LPS/ATP treated wild type microglia were stained for NLRP3 (green) and CALCOCO2 (red) and analysed by confocal microscopy. Colocalizations of NLRP3 aggregates (arrows) and CALCOCO2 (arrowheads) are apparent; scale bar: 10 µm. (B) LPS/ATP treated wild type microglia were stained for NLRP3 (green) and CALCOCO2 (red) and analysed by super resolution microscopy (SIM). NLRP3 aggregates coclustering with CALCOCO2-positive signals are visible. 3D volume rendering of NLRP3 aggregates from the magnified ROIs showed close contact of NLRP3 with the autophagic receptor CALCOCO2; scale bar: 10 µm; magnified ROIs: 1 µm. (C) 3D Radial intensity profiles of NLRP3 and CALCOCO2 signals derived from SIM images in wild type microglia, centered on the maxima of NLRP3 clusters. The radial profiles confirm that both proteins colocalize; mean ± SEM, n = 3. (D) In SIM images from wild type microglia the sub-autophagosomal localization of NLRP3 was analysed in relation to LC3 and CALCOCO2. LC3 and CALCOCO2 intensities from the center of mass of NLRP3 aggregates were normalised to their maximum within the cluster. While the center of NLRP3 aggregates shows near maximal CALCOCO2 intensity, the relative LC3 intensity there was significantly lower; mean ± SEM, LC3: n = 4, CALCOCO2: n = 3; two-tailed t-test **: p < 0.01. (E) Microglia from Becn1+/- mice were transfected with scrambled siRNA or with siRNA for CALCOCO2. 6 d after transfection cells were stimulated with LPS/ATP. The expression of CALCOCO2 protein was determined by Western blot. Application of CALCOCO2 siRNA resulted in significantly reduced expression of CALCOCO2; mean ± SEM, n = 3; two-tailed t-test *: p < 0.05. (F) The amount of IL-1beta in the supernatant released from the cells described in (E) was determined by ELISA. Microglia with a CALCOCO2 knockdown released significantly more IL-1beta; mean ± SEM, n = 3; two-tailed t-test *: p < 0.05.
Figure 1(A) Schematic representation of the features implemented in the XRep cell line. A hybrid background in which cells carry one X chromosome from M.m musculus (Xmus) and one from M.m castaneus (Xcas). The cell line carries an rtTA under the control of the Rosa26 locus and piggyBac transposon-based vectors with doxycycline (Dox)-responsive promoters driving the expression of Prdm14, Blimp1 and Tfap2c. The Xmus carries a GFP reporter and a truncation of the Tsix transcript while the Xcas carries a tdTomato reporter. (B) Overview of the adapted PGCLC differentiation timeline. Stages of the culture system are shown. (C) Representative FACS data of primordial germ cells specific surface markers CD61 and SSEA1 in ESCs, EpiLCs d4 and PGCLCs d5. Numbers indicate the percentages of SSEA1+/CD61+ gated cells over time. Shown are contour plots gated on live cells. (D) Immunostaining of PGCLCs d5 cryosections for SOX2 (magenta) and TFAP2C (cyan). Barplot indicates the quantification of SOX2+ cells, TFAP2C+ cells, and SOX2+/TFAP2C+. n=1150 cells, from n=3 separate inductions, using two biological clones. The white squares represent the position of the magnified region at the bottom. Scale bar, 50 µm and 10 µm for the magnified region. (E) Representative culture showing the X-activity reporter during PGCLC induction. Images for bright field (BF), XGFP, and XTomato were taken for ESCs, EpiLC d4 and PGCLC d5. Scale bar, 50 μm. (F) Representative FACS data showing XGFP and XTomato distribution during PGCLC induction. Numbers indicate the percentage of cells gated according to the XGFP and XTomato status (grey = X-inactive, green/red = X-active). Dashed line indicates the transition from X-active to X-inactive according to XGFP levels. XGFP and XTomato percentages in ESCs, EpiLCs and d1 full bodies are calculated from the entire cell population, while in PGCLC d2 to PGCLC d5 are calculated from SSEA1+/CD61+ PGCLCs as indicated. Shown are histograms gated on live cells.
Detection of ctDNA in CSF, plasma and urine from glioma patients using patient-specific sequencing panels and INVAR analysis.Figure 2A: Schematic of ctDNA detection in matched CSF, plasma and urine samples from glioma patients using INVAR (INtegration of VAriants Reads). Depth of sequencing indicated is the mean across the samples analysed. S1 to S4 indicate tumor subparts. B: Number of tumor tissue DNA mutations passing INVAR filters for the 8 patients included. C: Estimated ctDNA fractions for the plasma (10/12 detected), CSF (7/8 detected) and urine samples (10/16 detected) collected from 7 patients with primary glioblastoma and one patient (GB12) with anaplastic oligodendroglioma. The ctDNA fraction is expressed as IMAF (Integrated Mutant Allele Fraction). Detected cases are indicated by full circle and non-detected cases as an open circle. ND: non detected. D: Estimated tumor DNA DNA fraction (IMAF) in CSF, plasma and urine depending on the number of mutant reads detected for each samples included and number of informative reads supporting the observation. Detected cases are indicated by full circle and non-detected cases as an open circle. ND: non detected. E: Estimated tumor DNA fractions (IMAF) in CSF, plasma and urine for the matched samples collected at baseline pre-surgery. Detected cases are indicated by full circle and non-detected cases as an open circle. ND: non detected. F: IMAF for the CSF, plasma and urine of the patients with samples collected at 6 months follow-up (n=3). Matched MRI scans are added for annotation.
Figure 8A Two independently derived CRISPR/Cas9 PC3-LN4 EDC3 S161A mutants (EDC3 SA#1, #2) and wild type cells were injected subcutaneously in the flank (5X106) of SCID mice (n=8/group), and tumor growth was monitored until the control group reached 1000 mm3. The subcutaneous tumor volume measurement (mm3) was done as outlined in Materials and Methods. Data are presented as mean ± SEM, (n = 8/group). B Phosphorylated EDC3 levels were examined in normal immortalized breast cell lines, MCF10A and MCF12A, human breast cancer cell lines HCC38, HCC1954, MDA-MB-231, MDA-MD-468, BT-549, MRKNU1, and mouse 4T1 by Western blotting with the indicated Abs. C Dual immunohistochemistry staining of human breast tumor with p-EDC3 Ab (Fredriksson et al.) and total EDC3 (brown). Scale bar: 50µm. D Dual immunohistochemistry staining of human normal resected breast tissue sections with p-EDC3 Ab (Fredriksson et al.) and total EDC3 (brown). Scale bar: 50µm.
Loss of TREM2 does not prevent synapse loss and activated microglia morphologyA.Figure 8A. Immunohistochemical analysis of synaptophysin (SPH, pink) and VGAT (yellow) in coronal brain sections. Representative images of thalamus are shown. Scalebars = 50 μm. B. Quantification of SPH positive area. Three images per mouse were taken, means were normalized to WT samples (n=3 per genotype, female). C. Quantification of VGAT positive area. Three images per mouse were taken, means were normalized to WT samples (n=3 per genotype, female). D Data information: Data represent mean ± SEM. For statistical analysis one-way ANOVA with Tukey's post hoc test of Grn-/-, Trem2-/- and Double-/- was used. Statistical significance was set at **, p < 0.01; ***, p < 0.001; ****, p < 0.0001, and ns, not significant. D. Western blot of SPH in RIPA lysates from 14-month-old female WT, Grn-/-, Trem2-/-, and Double-/- mice. Actin was used as loading control. E. Quantification of SPH protein levels in D normalized to WT (n=3). F Data information: Data represent mean ± SEM. For statistical analysis one-way ANOVA with Tukey's post hoc test of Grn-/-, Trem2-/- and Double-/- was used. Statistical significance was set at **, p < 0.01; ***, p < 0.001; ****, p < 0.0001, and ns, not significant. F. Morphological analysis of cortical microglia. Representative maximum intensity projections of confocal z-stack images showing IBA1+ microglial cells of female WT, Grn-/-, Trem2-/- and Double-/- mice (scalebar = 50 µm). Arrows point to individual microglia, which are shown as three-dimensional reconstruction, scalebar = 10 µm. G. Morphological differences of cortical microglia from WT, Grn-/-, Trem2-/- and Double-/- mice shown by branch volume, sphericity score, branch length and the number of branch nodes. Statistical analysis of group difference for the morphological scores "Branch volume" (auc = 0.72), "Sphericity score" (auc = 0.82), "Branch length" (auc = 0.69) and "Number of branch nodes" (auc = 0.80) was performed using the Wilcoxon rank sum test with continuity correction and Bonferroni post-hoc correction for multiple testing in R (version 4.0.3). Two images per mouse (n=3 per genotype, female) were analyzed, each data point represents one microglia cell. Median and interquartile range are displayed.
Loss of SEZ6 selectively changes the levels of GluK2 and GluK3 at the neuronal surface.Figure 1A) Workflow of the surface analysis performed in SEZ6KO and WT neurons. SEZ6 flox/flox neurons were infected with CRE (SEZ6KO) and GFP (WT) virus at DIV2 and metabolic labeling was started at DIV5. Surface analysis was performed at DIV7. B) Protein classification of the total proteins (3209, left panel) and of the glycoproteins (571, right panel) detected in 3 out of 3 experiments. More than 90% of the glycoproteins detected are classified as membrane proteins, as expected for the surface labeling. C) Changes of the 3209 proteins detected on the cell surface of SEZ6KO neurons compared to WT. The average log2 fold change of SEZ6KO neurons over WT (log2 SEZ6KO/WT) is plotted against the negative log10 of each protein's p-value according to a one-sample t test. Blue labeled dots are considered as hits (including GluK2 and GluK3) and are proteins with a p-value less than 0.05 (neg. log10(0.05)=1.3; horizontal dotted line) and a fold-change to less than -0.5 or more than 0.5 (vertical dotted lines). GluK2 and GluK3 are labeled in pink, GluA1 and GluA2 are indicated with green lines and dots and GluN1 and GluN2B are marked with red lines and dots.
Figure 3A) Left panel: exemplificative FACS analysis of the FITC emission of unstained HC C-MSC (autofluorescence; white), HC C-MSC in GM treated with DCF (grey), ACM C-MSC in GM treated with DCF (red), and HC cells in GM treated with 2mmol/l H2O2 as positive control (blue). Right panel: mean DCF emission of HC and ACM C-MSC in GM (n=5 biological replicates; Two-tailed Student's t-test). Data information: mean ± SEM. * p<0.05, ** p<0.01. B) Left panels: representative images of MDA immunostaining (green) on ACM and HC C-MSC in GM. Nuclei are counterstained with Hoechst33342 (blue). Right panel: image quantification (n=4 biological replicates ACM, n=5 biological replicates HC; Two-tailed Student's t-test). Data information: mean ± SEM. * p<0.05, ** p<0.01. C) GSH/GSSG ratio quantification in HC and ACM C-MSC cultured in GM (n=8 biological replicates). Data information: mean ± SEM. * p<0.05, ** p<0.01. D) Top panel: representative images of Western Blot analysis of proteins extracted from ACM and HC C-MSC cultured in GM, hybridized with anti-CD36 and anti-PPARγ antibodies. Immunostaining of the housekeeping GAPDH is shown for normalization. Bottom panel: d.a. of PPARγ (n=18 biological replicates) and CD36 (n=17 vs. n=16 biological replicates) levels, normalized on GAPDH (Mann-Whitney test). Data information: mean ± SEM. * p<0.05, ** p<0.01. E) Results of a FACS analysis of ACM and HC C-MSC, cultured in GM (time-point 0) or AM for 3, 6 or 10 days (time-points 3, 6 and 10, respectively) and marked with anti-CD36 antibody and Nile red. The mean (n=4 biological replicates ACM, n=5 biological replicates HC) fluorescence of CD36 and Nile Red is shown for each condition, the relative regression line, its equation, R2 and p-value (X-Y correlation). F) Top panels: representative images of internalization of oxLDL (red) in HC and ACM cells, cultured either in GM or in AM, and subjected to 10μg/ml DiI-oxLDL treatment. Nuclei are counterstained with Hoechst33342 (blue). Bottom panel: quantification of the relative mean DiI fluorescence for each sample, measured by FACS analysis (n=3 biological replicates; Two-Way Anova). Data information: mean ± SEM. * p<0.05, ** p<0.01.
Flagellar intermediates in C.Figure 4A-D) Slices through electron cryotomograms (left panels), central slices through STAs (middle panels) and schematic representations (right panels), of C. jejuni ΔflhBC and ΔflhAC illustrating different flagellar intermediates (highlighted by white arrows). Green arrows in (A) and (C) point to the ~50-nm cytoplasmic ring associated with the MS-complex. Scale bars are 50 nm for cryotomogram slices, 20 nm for STAs.
INQ is located inside the nucleus A–D Cryo‐sections of (A) S.Figure 2A-D Cryo‐sections of (A) S. cerevisiae wt cells expressing GFP‐luciferase‐DM‐NLS, (B) hsp42Δ cells expressing GFP‐VHL. Sections were immunogold‐labeled with GFP‐specific antibodies. Gold particles are marked (black arrows). Electron‐dense regions represent protein aggregates (A). Locations of cytosol (C), nuclear envelope (NE, orange arrows), and nucleus (N) are given. Scale bars, 200 nm.E, F Average numbers of gold particles associated with nuclear and cytosolic protein aggregates or distributed throughout the cytosol or nucleus were determined. A-D Cryo‐sections of S. cerevisiae (C, D) wt or hsp42Δ cells expressing Hsp104‐GFP. Sections were immunogold‐labeled with GFP‐specific antibodies. Gold particles are marked (black arrows). Electron‐dense regions represent protein aggregates (A). Locations of cytosol (C), nuclear envelope (NE, orange arrows), and nucleus (N) are given. Scale bars, 200 nm.E, F Average numbers of gold particles associated with nuclear and cytosolicprotein aggregates or distributed throughout the cytosol or nucleus were determined.
Figure 5A) The SAM binding affinity of FTSJ1 as measured by ITC. (B The binding affinity of FTSJ1 alone or FTSJ1-WDR6 for tRNA analyzed by the gel mobility shift assay. C) For the reaction in figure C, 0.2 μΜ GST, 0.2 μΜ GST-WDR6, 1 μM FTSJ1, the mixture of 1 μM FTSJ1 with 0.2 μΜ GST-WDR6 or 0.2 μΜ GST, 0.5 mM SAM and 250 nM tRNAPhe(GAA) transcript were incubated.
ATP induces gliosis in hEGC A: Schematic workflow on the collection and processing of surgical specimens collected during a pancreaticoduodenectomy.Figure 5A: Schematic workflow on the collection and processing of surgical specimens collected during a pancreaticoduodenectomy. Samples were provided on ice directly from the operation room and the ME was separated from lamina propria mucosae. B: Visual representation of p-value (-log10) against fold enrichment of GO terms associated with enriched genes in patient specimen that underwent IM. C: Heat map of enriched genes connected with GO term gliosis in patient specimens that underwent IM at two time points: early and late; n=3 human patients. D: Venn diagram of gliosis genes expressed in human (red) and murine (blue) ME-tissue. E: IL-6 protein release in hEGC cultures upon stimulation with ATP (200µM) or ATPɣS (100µM) after 6h and 24h treatment (n=4-6, hEGCs). F: IL-6 protein release in hEGC cultures upon stimulation with NTPDase inhibitor ARL67156 at indicated concentrations; n=4-6, hEGCs. G: Immunofluorescence microscopy revealed P2X2 expression (green) in a majority of s100β+ (violet) hEGCs in intact myenteric ganglia of the human colon. White arrows mark double positive glia cells and green arrows mark P2X2 positive neurons. Scale bar, 50µm. H: Effect of the P2X2 antagonism on ATP-induced IL-6 release. ELISA measurement of IL-6 in hEGCs upon treatment with P2X2-antagonist PSB-1011 (20µM) alone or together with ATP (200µM) treated for 24h (n=6, hEGCs). I: PSB-1011 (20μM) treatment inhibited ATP-triggered calcium responses in HEK cells transfected with P2X2. Data are represented as ΔF/F0 +SEM; n=102 HEK cells. J: The P2X2 receptor antagonist PSB-0711 (20μM) nearly abolished the ATP-triggered calcium response in HEK cells transfected with P2X2. n=219 HEK cells. K: Ambroxol blocks ATP-induced IL-6 release in hEGCs; Protein release measurement by ELISA of IL-6 in hEGCs. Cells were treated with ATP (200 µM) alone or together with ambroxol (20 µM) for 24h; n=6, hEGCs. L-N: qPCR analysis of several gliosis and ATP-target genes in the mechanically manipulated surgical specimens incubated in the presence or absence of ambroxol (20µM). (n=7, 4 human patients).