Journal: Aging cell

Article Title: Akt Regulates TPP1 Homodimerization and Telomere Protection

doi: 10.1111/acel.12137

Figure Lengend Snippet: (A) HTC75 cells treated with triciribine or mock treated with DMSO for 3 hours were analyzed by western blotting using the indicated antibodies. Actin was used as loading control. Arrows indicate the two POT1 isoforms. (B) Cells from (A) were examined by telomere chromatin immunoprecipitation (ChIP) using antibodies against TPP1, POT1 and RAP1, followed by dot-blotting with probes against telomere sequences or Alu repeats. Rabbit IgG served as control. (C) Quantification of data from (B) (two independent experiments). ChIP signal intensities were normalized against input DNA. Error bars represent s.d. P-values were determined by the Student t-test.

Article Snippet: The antibodies used for ChIP are: rabbit polyclonal POT1 (N-terminal) ( O'Connor et al., 2006 ), rabbit polyclonal TPP1 (467) ( Liu et al., 2004 ), polyclonal anti-RAP1 (Bethyl Laboratories) and rabbit IgG (Santa Cruz, SC2027).

Techniques: Western Blot, Control, Chromatin Immunoprecipitation

Journal: Aging cell

Article Title: Akt Regulates TPP1 Homodimerization and Telomere Protection

doi: 10.1111/acel.12137

Figure Lengend Snippet: (A) HTC75 cells co-expressing YFPn-tagged TPP1 with YFPc-tagged TPP1 or TIN2 were examined by BiFC assays. The percentage of cells displaying fluorescence complementation was quantitated by flow cytometry. Error bars indicate standard error (n=3). (B) 293T cells transiently co-expressing Myc-tagged TPP1 with Flag-tagged TPP1 or TIN2 were immunoprecipitated with anti-Flag antibodies. The immunoprecipitates were western blotted as indicated. (C) 293T cells transiently co-expressing Myc-tagged TPP1 with Flag-tagged TPP1 or TPP1 OB-fold deletion mutant (TPP1ΔOB) were immunoprecipitated with anti-Flag antibodies. The immunoprecipitates were western blotted as indicated. (D) Bacterially purified GST-tagged TPP1 OB fold only mutant (TPP1 OB) was incubated with MBP-tagged TPP1 OB for GST pull-down assays. The precipitates were resolved by SDS-PAGE and visualized by Coomassie staining or western blotting. GST-tagged Raf-1 Ras-binding domain (RBD) and MBP-tagged FAM118B were used as controls. (E) HTC75 cells were transfected with siRNA oligos against Akt1 (siAKT1-1) in combination with Flag-tagged wildtype TPP1 or TPP1 OB fold deletion mutant (TPP1ΔOB), and then analyzed by Western blotting using the indicated antibodies. Actin was used as loading control. (F) Cells from (E) were examined by immunostaining using anti-53BP1 (red) and TRF2 (green) antibodies. Arrows indicate overlapping foci. (G) Quantification of data from (F). Only cells with >4 co-localized foci were scored. Error bars indicate s.e.m. (n=3). P-values were determined by the Student t-test.

Article Snippet: The antibodies used for ChIP are: rabbit polyclonal POT1 (N-terminal) ( O'Connor et al., 2006 ), rabbit polyclonal TPP1 (467) ( Liu et al., 2004 ), polyclonal anti-RAP1 (Bethyl Laboratories) and rabbit IgG (Santa Cruz, SC2027).

Techniques: Expressing, Fluorescence, Flow Cytometry, Immunoprecipitation, Western Blot, Mutagenesis, Purification, Incubation, SDS Page, Staining, Binding Assay, Transfection, Control, Immunostaining

Journal: Aging cell

Article Title: Akt Regulates TPP1 Homodimerization and Telomere Protection

doi: 10.1111/acel.12137

Figure Lengend Snippet: (A) HTC75 cells co-expressing YFPn-tagged TPP1 with YFPc-tagged TPP1 or TIN2 were either serum starved (0.01% FBS) or treated with triciribine, and then examined by BiFC assays. Error bars indicate standard error (n=3). (B) 293T cells transiently co-expressing TPP1-Flag with Myc-TPP1 were treated with DMSO or triciribine. Cells co-expressing SFB-Akt1 and Myc-TPP1 were also examined. Cell lysates were immunoprecipitated with anti-Flag antibodies and blotted as indicated. (C) HTC75 cells co-expressing YFPn-TPP1 with YFPc-TPP1 were transiently transfected with siRNA oligos against Akt1 and analyzed in BiFC assays. Error bars indicate standard errors (n=3). (D) Cells from (C) were examined by western blotting for Akt1 knockdown efficiency. Actin was used as loading control. (E) HTC75 cells co-expressing YFPn-TPP1 and YFPc-TPP1 were transiently co-transfected with control oligos or siRNA oligos against Akt1 in combination with siRNA-resistant SFB-tagged wildtype (WT) Akt1 or kinase-dead Akt1 (Akt1 K179M). The cells were then examined by western blotting using the indicated antibodies. Actin was used as a loading control. (F) Cells from (E) were analyzed in BiFC assays. P-values were determined by the Student t-test.

Article Snippet: The antibodies used for ChIP are: rabbit polyclonal POT1 (N-terminal) ( O'Connor et al., 2006 ), rabbit polyclonal TPP1 (467) ( Liu et al., 2004 ), polyclonal anti-RAP1 (Bethyl Laboratories) and rabbit IgG (Santa Cruz, SC2027).

Techniques: Expressing, Immunoprecipitation, Transfection, Western Blot, Knockdown, Control

Journal: Biochimie

Article Title: The OB-fold domain 1 of human POT1 recognizes both telomeric and non-telomeric DNA motifs.

doi: 10.1016/j.biochi.2015.04.015

Figure Lengend Snippet: Fig. 6. The NT motif is recognized by purified POT1 and POT1/TPP1 dimer. A) Purity of the isolated POT1 (399 ng) and TPP1 proteins (163 ng), as determined by SDS-PAGE and silver staining. B) EMSA with the purified human POT1 protein. The purified protein (0.5 pmol) was incubated with the indicated radiolabeled probes and the complexes formed (arrowhead) were resolved by electrophoresis. C) Titration of the TPP1 protein for the formation of POT1/TPP1 dimers. EMSA was performed with (þ) and without () POT1 (0.5 pmol) in the presence of an increasing molar excess of TPP1 (1, 2, 4, 8). Probe used was the canonical OB1/OB2 binding site. Arrowheads show the location of the POT1/ DNA and POT1/TPP1/DNA complexes. D) Sequence specificity of the POT1/TPP1 dimer and its interaction with the NT motif. EMSA was performed with the indicated probes in the presence of either no protein, POT1 alone (0.5 pmol), or POT1 (0.5 pmol) plus TPP1 (2 pmol). Arrowheads show the location of the POT1/DNA and POT1/TPP1/DNA complexes.

Article Snippet: A human TPP1 protein purified from transfected human cells was purchased (OriGene Technologies Inc., Rockville, MD).

Techniques: Isolation, SDS Page, Silver Staining, Incubation, Electrophoresis, Titration, Binding Assay, Sequencing

Journal: American Journal of Respiratory Cell and Molecular Biology

Article Title: Shelterin Telomere Protection Protein 1 Reduction Causes Telomere Attrition and Cellular Senescence via Sirtuin 1 Deacetylase in Chronic Obstructive Pulmonary Disease

doi: 10.1165/rcmb.2016-0198OC

Figure Lengend Snippet: Small airway epithelial cells (SAECs) from patients with chronic obstructive pulmonary disease (COPD) show increased senescence and telomere dysfunction, together with telomere protection protein 1 (TPP1) reduction, which is augmented by cigarette smoke extract (CSE) treatment. SAECs from normal subjects and from patients with COPD were treated with CSE (0.2%) for 10 days. (A) Representative images showing senescence-associated β-galactosidase activity in human SAECs. (B) Representative images of cells stained with TPP1 (red) and Dapi (4′6-diamidino-2-phenylindole, blue), showing TPP1 expression in SAECs from normal subjects. Scale bar: 100 μm. The average fluorescent intensity of TPP1 was calculated in SAECs treated with CSE using MetaMorph software. (C) Southern blot–based telomere length assay was performed in SAECs treated with CSE. (D) Representative images of CSE-treated SAECs from normal subjects and from patients with COPD were stained with γH2AX (green) and teloprobe (Telo, red), together with Dapi (blue) for telomere dysfunction–induced foci. Scale bar: 100 µm. Data are shown as mean ± SEM (n = 3–5). ***P < 0.001 versus control (Con). A.U., arbitrary units. Con, untreated control.

Article Snippet: Constructs and Transfections Myc-tagged TPP1 vectors were obtained from Origene (Rockville, MD).

Techniques: Activity Assay, Staining, Expressing, Software, Southern Blot

Journal: American Journal of Respiratory Cell and Molecular Biology

Article Title: Shelterin Telomere Protection Protein 1 Reduction Causes Telomere Attrition and Cellular Senescence via Sirtuin 1 Deacetylase in Chronic Obstructive Pulmonary Disease

doi: 10.1165/rcmb.2016-0198OC

Figure Lengend Snippet: TPP1 reduction is observed in primary airway epithelial cells, mouse lung cells, and lungs of patients with COPD. (A) SAECs showing reduced TPP1 levels after being treated with CSE (0.2%) for 15 days. Densitometry of the corresponding bands was normalized to β-actin loading control. (B) Southern blot–based telomere length assay was performed in lungs from nonsmokers (NS), smokers, and patients with COPD. Densitometry of bands for telomere length is presented. (C) Western blot of TPP1 in lung homogenates from NS, smokers, and patients with COPD was performed, and representative glyceraldehyde phosphate dehydrogenase (GAPDH) was used as a loading control. Densitometry of the corresponding bands was normalized to GADPH. (D) Representative images of bronchial staining of TPP1 in lungs from NS, smokers, and patients with COPD are presented; histogram shows the average intensity of TPP1 staining using MetaMorph software. Representative images of the alveolar region of TPP1 staining in lungs from NS, smokers, and patients with COPD are presented. Average intensity of TPP1 staining was calculated using MetaMorph software. Scale bars: 100 μm. Data are shown as mean ± SEM (n = 3–4). *P < 0.05, **P < 0.01 versus NS; ***P < 0.001 versus control.

Article Snippet: Constructs and Transfections Myc-tagged TPP1 vectors were obtained from Origene (Rockville, MD).

Techniques: Southern Blot, Western Blot, Staining, Software

Journal: American Journal of Respiratory Cell and Molecular Biology

Article Title: Shelterin Telomere Protection Protein 1 Reduction Causes Telomere Attrition and Cellular Senescence via Sirtuin 1 Deacetylase in Chronic Obstructive Pulmonary Disease

doi: 10.1165/rcmb.2016-0198OC

Figure Lengend Snippet: TPP1 reduction in airway epithelium, alveolar type II cells, and lung fibroblasts by chronic cigarette smoke (CS) exposure in emphysematous mice. (A) TPP1 expression in CCSP+ (Club/Clara Cell Secretory Protein, CC16 in the small airways [CC16/CC10]) cells, (B) proSPC+ (pro-surfactant protein C) alveolar type II cells, and (C) α smooth muscle actin (αSMA)+ lung fibroblasts was reduced by chronic CS exposure in mice. Immunofluorescence staining scores for TPP1 expression in CCSP+ CC10 airway, SPC+ alveolar epithelial cells, and αSMA+ lung fibroblasts were determined semiquantitatively in a blinded fashion and are presented as staining score percentage (%). Data are shown as mean ± SEM (n = 3–4/group). *P < 0.05, **P < 0.01 versus air group.

Article Snippet: Constructs and Transfections Myc-tagged TPP1 vectors were obtained from Origene (Rockville, MD).

Techniques: Expressing, Immunofluorescence, Staining

Journal: American Journal of Respiratory Cell and Molecular Biology

Article Title: Shelterin Telomere Protection Protein 1 Reduction Causes Telomere Attrition and Cellular Senescence via Sirtuin 1 Deacetylase in Chronic Obstructive Pulmonary Disease

doi: 10.1165/rcmb.2016-0198OC

Figure Lengend Snippet: CSE causes TPP1 acetylation and disrupts its interaction with Sirtuin 1 (Sirt1) in HFL1 cells. HFL1 cells were treated with CSE (0.5%) for (A) 24 hours or (B–D) 15 days. (A) Cell lysates were immunoprecipitated by TPP1 antibody, followed by probing with anti–acetyl lysine antibody. (B) Immunoprecipitated TPP1 immunocomplex was used for Western blot, which was probed with Sirt1 and TPP1 antibodies. (C) Western blot images of immunoprecipitation of Sirt1, followed by its probing with TPP1. Representative GAPDH was used as a loading control, which was run separately from the same samples without IP. Densitometry of the corresponding blots is presented. (D) Representative images of TPP1 (green) and Sirt1 (red) showing the colocalization in the nucleus, which was stained with Dapi (blue). Scale bar: 100 µm. Line scan data of corresponding images show the degree of colocalization between TPP1 and Sirt1. Data are shown as mean ± SEM (n = 3–4). *P < 0.05 versus control. IP, immunoprecipitation.

Article Snippet: Constructs and Transfections Myc-tagged TPP1 vectors were obtained from Origene (Rockville, MD).

Techniques: Immunoprecipitation, Western Blot, Staining

Journal: American Journal of Respiratory Cell and Molecular Biology

Article Title: Shelterin Telomere Protection Protein 1 Reduction Causes Telomere Attrition and Cellular Senescence via Sirtuin 1 Deacetylase in Chronic Obstructive Pulmonary Disease

doi: 10.1165/rcmb.2016-0198OC

Figure Lengend Snippet: TPP1 reduction is observed in mouse lungs with emphysema. Sirt1+/−, Sirt1 Tg, and WT mice were exposed to CS for 6 months. (A) Telomere length assay was performed in mouse lungs on the basis of a Southern blot–based technique. (B) Western blot of TPP1 in lung homogenates was performed, and representative GAPDH was used as a loading control. Densitometry of the corresponding bands was normalized to GADPH. Representative images of TPP1 expression in (C) the bronchial airways (insets showing magnified image of stained bronchial epithelial cells) and (D) the alveolar region in mouse lungs and histogram show the average intensity of TPP1 staining in the bronchial and alveolar regions calculated using MetaMorph software. Arrows indicate dark brown staining for TPP1 expression in bronchial airway and alveolar epithelial cells shown in (C) and (D). Data are shown as mean ± SEM (n = 3–4). *P < 0.05, ***P < 0.001 versus WT-Air; +P < 0.05, ++P < 0.01 versus WT-CS. Sirt1+/−, Sirt1 heterozygous knockout; Sirt1 Tg, Sirt1 transgenic; WT, wild type.

Article Snippet: Constructs and Transfections Myc-tagged TPP1 vectors were obtained from Origene (Rockville, MD).

Techniques: Southern Blot, Western Blot, Expressing, Staining, Software, Knock-Out, Transgenic Assay

Journal: American Journal of Respiratory Cell and Molecular Biology

Article Title: Shelterin Telomere Protection Protein 1 Reduction Causes Telomere Attrition and Cellular Senescence via Sirtuin 1 Deacetylase in Chronic Obstructive Pulmonary Disease

doi: 10.1165/rcmb.2016-0198OC

Figure Lengend Snippet: Sirt1 regulates CSE-induced TPP1 reduction and TIF formation in lung fibroblasts. (A–D) Mouse lung fibroblasts isolated from Sirt1+/−, Sirt1 Tg, and WT mice were treated with CSE (0.25% for 15 d). (A) Sirt1 levels determined by FACS in CSE-treated lung fibroblasts from WT, Sirt1+/−, and Sirt1 Tg mice. (B) Representative images of SA-β-gal staining in mouse lung fibroblasts treated with CSE. Scale bar: 100 µm. (C) Western blot of TPP1 in whole cell extracts of lung fibroblasts obtained from WT, Sirt1+/−, and Sirt1 Tg mice. Representative GAPDH was used as a loading control. Densitometry of the corresponding TPP1 bands normalized to GAPDH. (D) Lung fibroblasts were stained with Dapi (blue), γH2AX (green), and Telo (red) for TIF formation. Scale bar: 100 µm. (E–G) HFL1 cells were treated with CSE (0.5%) for 15 days in the presence of SIRT1 inhibitor (sirtinol, 1 μM) or activator (SRT1720, 10 μM). (E) TPP1 levels were determined by the FACS in CSE-treated HFL1 cells in the presence of sirtinol or SRT1720. (F) HFL1 cells were stained with γ-H2AX (green) and Telo (red) for TIF formation, together with Dapi (blue). Scale bar: 100 µm. (G) C12FDG fluorescence in CSE-treated HFL1 cells measured by the FACS was plotted as FC12FDG in cells, which showed that the degree of cellular senescence was arbitrary. Data are shown as mean ± SEM (n = 3–4). *P < 0.05, **P < 0.01 versus Con; +P < 0.05, ++P < 0.01 versus CSE group. FC12FDG, fluorescent intensity of 5-dodecanoylaminofluorescein di-β-d-galactopyranoside; FSirt1, fluorescent intensity of Sirt1; FTPP1, fluorescent intensity of TPP1; Veh, vehicle.

Article Snippet: Constructs and Transfections Myc-tagged TPP1 vectors were obtained from Origene (Rockville, MD).

Techniques: Isolation, Staining, Western Blot, Fluorescence

Journal: Oncology reports

Article Title: The transcription factor RFX5 is a transcriptional activator of the TPP1 gene in hepatocellular carcinoma.

doi: 10.3892/or.2016.5240

Figure Lengend Snippet: Figure 2. RFX5 binds to the TPP1 promoter region and activates transcription of the TPP1 promoter. (A) The sketch of the TPP1 promoter region shows the RFX5 ChIP-seq binding peak in HepG2 cell and designed primers. Upper, RFX5 motif consensus sequence from ENCODE is shown, the S-Y motifs from HLA-DRA, HLA-DMA, HLA-DMB and TPP1 are boxed. Lower, (blue bold lines) the design of two PCR amplicons for the luciferase reporter gene, (Box) annotation of the TPP1 promoter based on the ENCODE Histone Modification Tracks, (light blue bold lines) the two ChIP-PCR amplicons and (purple peak) the RFX5 ChIP-seq binding peak in the TPP1 promoter. (B) ChIP-PCR assays showing RFX5 binding to the TPP1 promoter in HCC cells. Immunoprecipitated DNA fragments were analyzed by PCR using two independent primers mapped to the RFX5 ChIP-seq binding peak. (C) The transcriptional activity of two TPP1 promoter reporter constructs. Relative luciferase activity (RLA) was calculated as the ratio of firefly to Renilla luciferase activities to represent the promoter activity and performed in triplicate. The results represent one of three experiments. (D) Western blotting showing RFX5 protein with or without overexpression in 293T cells. (E) Luciferase assay to analyze the transcriptional impact of RFX5 on the TPP1 promoter in HCC cells. *P<0.05, ****P<0.0001.

Article Snippet: Paraffin-embedded tissue slides were heatwd with EDTA (pH 9.0) (ZSGB-BIO, Beijing, China) for antigen retrieval and incubated with rabbit anti-human TPP1 antibody (polyclonal, #12479-1-AP, Proteintech), followed by EnVision detection system (Dako, Denmark).

Techniques: ChIP-sequencing, Binding Assay, Sequencing, Luciferase, Modification, Immunoprecipitation, Activity Assay, Construct, Western Blot, Over Expression

Journal: Oncology reports

Article Title: The transcription factor RFX5 is a transcriptional activator of the TPP1 gene in hepatocellular carcinoma.

doi: 10.3892/or.2016.5240

Figure Lengend Snippet: Figure 3. TPP1 expression is linked to RFX5 expression and poor prognoses in HCC. (A) Real-time PCR showing RFX5 and TPP1 expression in HepG2 cells after RFX5 knockdown. (B) Real-time PCR shows RFX5 and TPP1 expression in HepG2 cells after RFX5 overexpression. (C) TPP1 mRNA expression in the TCGA LIHC data determined by RNAseq. (D) Immunohistochemistry staining of TPP1 in HCC and matched adjacent liver tissues by IHC (magnification, x200). (E) Correlation analysis between RFX5 and TPP1 expression from the TCGA LIHC data. (F) Correlation analysis between RFX5 and TPP1 expression using real-time PCR data from the Guilin cohort. (G) Survival analysis from the overall survival time (OST) in HCC patients with relatively lower or higher TPP1 expression in the tumor tissues. The Kaplan-Meier plot was analyzed by the log-rank test. **P<0.01, ***P<0.001, ****P<0.0001.

Article Snippet: Paraffin-embedded tissue slides were heatwd with EDTA (pH 9.0) (ZSGB-BIO, Beijing, China) for antigen retrieval and incubated with rabbit anti-human TPP1 antibody (polyclonal, #12479-1-AP, Proteintech), followed by EnVision detection system (Dako, Denmark).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Knockdown, Over Expression, Immunohistochemistry, Staining

Journal: Cell reports

Article Title: A High-Content Screen Identifies TPP1 and Aurora B as Regulators of Axonal Mitochondrial Transport

doi: 10.1016/j.celrep.2019.08.035

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: pCMV6-Myc-DDK-Tpp1 (human) , OriGene , RC204471.

Techniques: Virus, Recombinant, Drug discovery, Software

Journal: Orphanet journal of rare diseases

Article Title: Neural stem cells for disease modeling and evaluation of therapeutics for infantile (CLN1/PPT1) and late infantile (CLN2/TPP1) neuronal ceroid lipofuscinoses.

doi: 10.1186/s13023-018-0798-2

Figure Lengend Snippet: Fig. 2 Effect of recombinant human PPT1/TPP1 protein on reducing enlarged lysosomes in NCL patient NSCs. The western blot analysis (a, b) showed that there is a PPT1 deficiency in PPT1E8/E1 fibroblasts and NSCs, and also there is no TPP1 expression detected in TPP1E4/E6 and TPP1E4/IVS5 fibroblast and NSCs. The treatment of NCL NSCs with 200 nM rPPT1/rTPP1 significantly reduced the LysoTracker dye staining(c), with an effect nearly 99.9% in the NCL NSC lines treated with ERT (d). The images were taken with 40X objective lens. Data are displayed as mean ± SD. ** P < 0.01

Article Snippet: ERT with recombinant human palmitoyl-protein thioesterase 1 (PPT1) and tripeptidyl-peptidase 1 (TPP1) Human recombinant PPT1 (14703-H08H) was purchased from Sino Biological (Beijing, China), and TPP1 (2237-SE-010) was ordered from R&D Systems (Minneapolis, MN).

Techniques: Recombinant, Western Blot, Expressing, Staining

Journal: Orphanet journal of rare diseases

Article Title: Neural stem cells for disease modeling and evaluation of therapeutics for infantile (CLN1/PPT1) and late infantile (CLN2/TPP1) neuronal ceroid lipofuscinoses.

doi: 10.1186/s13023-018-0798-2

Figure Lengend Snippet: Fig. 7 Co-localization of subunit c and Lamp1 in NCL NSCs and the expression of subunit c in NCL NSCs. a Co-localization of LAMP-1, a lysosomal marker, with subunit c in PPT1E8/E1, TPP1E4/E6, and wild-type NSCs. The cells were immunostained with antibodies recognizing subunit c (red fluorescence, see white arrows) and Lamp1 (green fluorescence). Minimal overlap of subunit c and Lamp1 immunostaining was observed in wild-type cells (yellow in overlay), but Lamp1 strongly, though not perfectly, overlaps with the accumulated subunit c in PPT1E8/E1 and TPP1E4/E6 NSCs. Treatment of INCL and LINCL NSCs with recombinant PPT1 and TPP1 decreased subunit c accumulation in lysosomes of patient cells, respectively. Similar effects were also observed in cells after treatments with δ-tocopherol and HPBCD. Blue represents Hoechst nuclei stain. Images were captured with 60X objective. b and c Expression of subunit c in NCL fibroblasts analyzed by the Western blot. The expressions of subunit c in PPT1E8/E1 fibroblast were weaker than WT, but the expressions of subunit c increased in TPP1E4/E6 and TPP1E4/IVS5 fibroblast compared to WT (b). It showed that subunit c expression was decreased by 64% in PPT1E8/E1

Article Snippet: ERT with recombinant human palmitoyl-protein thioesterase 1 (PPT1) and tripeptidyl-peptidase 1 (TPP1) Human recombinant PPT1 (14703-H08H) was purchased from Sino Biological (Beijing, China), and TPP1 (2237-SE-010) was ordered from R&D Systems (Minneapolis, MN).

Techniques: Expressing, Marker, Fluorescence, Immunostaining, Recombinant, Staining, Western Blot

Journal: Orphanet journal of rare diseases

Article Title: Neural stem cells for disease modeling and evaluation of therapeutics for infantile (CLN1/PPT1) and late infantile (CLN2/TPP1) neuronal ceroid lipofuscinoses.

doi: 10.1186/s13023-018-0798-2

Figure Lengend Snippet: Fig. 8 Effect of DT, HPBCD and enzyme replacement therapy on the accumulation of subunit c in patient NSCs. Cells were treated with 20 μM DT, 1 mM HPBCD, 20 μM DT plus 125 μM HPBCD, or 200 nM rPPT1/rTPP1 for 3 days. PPT1/TPP1 expression were restored after the PPT1/TPP1 replacement therapy. After the treatment with 20 μM DT plus 125 μM HPBCD, the expression of subunit c in PPT1E8/E1NSCs decreased by 75% (a and b). Moreover, it showed that subunit c expression in TPP1E4/E6 NSCs and TPP1E4/IVS5 NSCs was decreased by 51% and 64% respectively, in TPP1 replacement treatment, and also decreased by 18%, 30% with DT treatment (c, d, e and f). Data are displayed as mean ± SD. * P < 0.05, ** P < 0.01

Article Snippet: ERT with recombinant human palmitoyl-protein thioesterase 1 (PPT1) and tripeptidyl-peptidase 1 (TPP1) Human recombinant PPT1 (14703-H08H) was purchased from Sino Biological (Beijing, China), and TPP1 (2237-SE-010) was ordered from R&D Systems (Minneapolis, MN).

Techniques: Expressing