Journal: Molecular Medicine Reports

Article Title: Alterations in the long non-coding RNA transcriptome in mesangial cells treated with aldosterone in vitro

doi: 10.3892/mmr.2017.7313

Figure Lengend Snippet: Reverse transcription-quantitative polymerase chain reaction quantification of microarray hybridization. (A) The relative expression level of lncRNAs BC168211, BC088254, AF336872, AY325162, BC168687, AF230638 and BC167085. (B) The relative expression levels of upregulated mRNAs, NM_001108598, NM_001109190 and NM_001101018, and (C) downregulated mRNAs, NM_019347, NM_177,962 and NM_0,011,08823. *P<0.05 vs. the control group. ALD, aldosterone; lncRNAs, long non-coding RNAs; MCs, mesangial cells.

Article Snippet: The cells were stored at −80°C and sent to the LncRNA Expression Microarray (Arraystar, Rockville, MD, USA).

Techniques: Reverse Transcription, Real-time Polymerase Chain Reaction, Microarray, Hybridization, Expressing, Control

Journal: Journal of Assisted Reproduction and Genetics

Article Title: Co-localization of NANOG and OCT4 in human pre-implantation embryos and in human embryonic stem cells

doi: 10.1007/s10815-012-9824-9

Figure Lengend Snippet: Representative pictures show double staining of OCT4 and NANOG in the human morula and blastocyst. a Immunostaining of OCT4 in the morula. b Immunostaining of NANOG in the same morula. c DAPI staining in the morula. d Staining of OCT4 in the blastocyst. Immunostaining is seen both in the inner cell mass and the trophoblast; arrows. e Staining of NANOG in the same blastocyst. Staining is seen only in the inner cell mass; arrow. f DAPI staining in the same blastocyst. g Exclusion of OCT4 antibody. h Exclusion of NANOG antibody. i DAPI staining of the embryo without primary antibodies present

Article Snippet: Primary antibodies—monoclonal mouse antibodies for OCT4 (sc-5279, Santa Cruz Biotechnologies, Santa Cruz, USA), diluted 1:80, and monoclonal goat antibody for NANOG (MAB1997, R&D systems, Minneapolis, MN USA), diluted 1:200—were added in 5 % blocking buffer overnight at 4 °C and washed three times with PBS to remove any unbound primary antibodies.

Techniques: Double Staining, Immunostaining, Staining

Journal: Journal of Assisted Reproduction and Genetics

Article Title: Co-localization of NANOG and OCT4 in human pre-implantation embryos and in human embryonic stem cells

doi: 10.1007/s10815-012-9824-9

Figure Lengend Snippet: In situ hybridization of NANOG in the human pre-implantation embryo. a 2-cell embryo, day 1. b 4-cell embryo, day 2. c 8-cell embryo, day 3. d morula, day 4. e late day 4 embryo. f blastocyst, day 5. g mouse ESCs hybridized with human NANOG primer

Article Snippet: Primary antibodies—monoclonal mouse antibodies for OCT4 (sc-5279, Santa Cruz Biotechnologies, Santa Cruz, USA), diluted 1:80, and monoclonal goat antibody for NANOG (MAB1997, R&D systems, Minneapolis, MN USA), diluted 1:200—were added in 5 % blocking buffer overnight at 4 °C and washed three times with PBS to remove any unbound primary antibodies.

Techniques: In Situ Hybridization

Journal: Journal of Assisted Reproduction and Genetics

Article Title: Co-localization of NANOG and OCT4 in human pre-implantation embryos and in human embryonic stem cells

doi: 10.1007/s10815-012-9824-9

Figure Lengend Snippet: Immunostaining of OCT4 and NANOG in human embryonic stem cells, and in situ hybridization of NANOG in embryonic stem cells. a Light microscopy of human embryonic stem cells. b NANOG immunostaining in embryonic stem cells. The feeder cells do not show staining for NANOG (red arrow). c Nuclear staining of embryonic stem cells and fibroblast cells. DAPI staining is seen in feeder cells (blue arrow). d & e In situ hybridization showing NANOG mRNA in embryonic stem cells. NANOG f Mouse embryonic stem cells hybridized with human NANOG primer. g OCT4 immunostaining of stem cell line HS426 (green). Blue colour shows nuclear DAPI staining

Article Snippet: Primary antibodies—monoclonal mouse antibodies for OCT4 (sc-5279, Santa Cruz Biotechnologies, Santa Cruz, USA), diluted 1:80, and monoclonal goat antibody for NANOG (MAB1997, R&D systems, Minneapolis, MN USA), diluted 1:200—were added in 5 % blocking buffer overnight at 4 °C and washed three times with PBS to remove any unbound primary antibodies.

Techniques: Immunostaining, In Situ Hybridization, Light Microscopy, Staining

Journal: Journal of Assisted Reproduction and Genetics

Article Title: Co-localization of NANOG and OCT4 in human pre-implantation embryos and in human embryonic stem cells

doi: 10.1007/s10815-012-9824-9

Figure Lengend Snippet: Data from microarray analysis of human embryos and embryonic stem cells and fibroblasts. For Nanog, the average signal intensity from one probe set and for Oct4 the average number of three probe sets is shown

Article Snippet: Primary antibodies—monoclonal mouse antibodies for OCT4 (sc-5279, Santa Cruz Biotechnologies, Santa Cruz, USA), diluted 1:80, and monoclonal goat antibody for NANOG (MAB1997, R&D systems, Minneapolis, MN USA), diluted 1:200—were added in 5 % blocking buffer overnight at 4 °C and washed three times with PBS to remove any unbound primary antibodies.

Techniques: Microarray

Journal: OncoTargets and therapy

Article Title: Overexpression of UHRF2 in intrahepatic cholangiocarcinoma and its clinical significance

doi: 10.2147/OTT.S149361

Figure Lengend Snippet: The expression of UHRF2 in human cancers and ICC patients. Notes: ( A ) Microarray data analysis of UHRF2 mRNA in gastric cancer, colon cancer, and hepatocellular carcinoma from the publicly available Oncomine database. ( B ) UHRF2 protein expression in randomly selected ICC tissues and their matched peritumor tissues by Western blot. ( C ) Relative expression of UHRF2 among tumor and peritumor tissues using qRT-PCR. ( D ) Positive UHRF2 was observed primarily in the cytoplasm. Representative UHRF2 and H

Article Snippet: Rabbit anti-human UHRF2 monoclonal antibody (1:1,000; Wuhan Boster Biological Technology, Ltd., Wuhan, People’s Republic of China) and rabbit anti-human E-cadherin antibody (1:100; Cell Signaling Technology, Danvers, MA, USA) were utilized to detect the levels of UHRF2 and E-cadherin.

Techniques: Expressing, Microarray, Western Blot, Quantitative RT-PCR, Staining, Real-time Polymerase Chain Reaction, Ubiquitin Proteomics, Immunohistochemistry

Journal: OncoTargets and therapy

Article Title: Overexpression of UHRF2 in intrahepatic cholangiocarcinoma and its clinical significance

doi: 10.2147/OTT.S149361

Figure Lengend Snippet: UHRF2 expression resulted in ICC cell proliferation, invasion, migration, and antiapoptosis. Notes: ( A ) UHRF2 expression was interfered by siRNA and confirmed by Western blot and qRT-PCR. GAPDH was used as internal control. ( B ) Cell counting kit-8 assay was used to assess the ability of cell proliferation. ( C ) Transwell assay was used to show the invasion of ICC cells with different UHFR2 expression in 48 hours. Scale bar =100 μm. ( D ) Wound healing assays showed that inhibition of UHRF2 decreased wound healing compared with control cells. Scale bar =100 μm. ( E ) FCM results indicated that anti-UHRF2 caused acceleration of cell apoptosis. The results are mean ± SD of triplicated independent experiments. * P <;0.05, ** P <;0.01, *** P <;0.001. Abbreviations: FCM, flow cytometry; FITC, fluorescein isothiocyanate; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; ICC, intrahepatic cholangiocarcinoma; qRT-PCR, quantitative real-time polymerase chain reaction; UHRF2, ubiquitin-like with PHD and ring finger domains 2.

Article Snippet: Rabbit anti-human UHRF2 monoclonal antibody (1:1,000; Wuhan Boster Biological Technology, Ltd., Wuhan, People’s Republic of China) and rabbit anti-human E-cadherin antibody (1:100; Cell Signaling Technology, Danvers, MA, USA) were utilized to detect the levels of UHRF2 and E-cadherin.

Techniques: Expressing, Migration, Western Blot, Quantitative RT-PCR, Control, Cell Counting, Transwell Assay, Inhibition, Flow Cytometry, Real-time Polymerase Chain Reaction, Ubiquitin Proteomics

Journal: OncoTargets and therapy

Article Title: Overexpression of UHRF2 in intrahepatic cholangiocarcinoma and its clinical significance

doi: 10.2147/OTT.S149361

Figure Lengend Snippet: UHRF2 was negatively associated with E-cadherin expression in ICC. Notes: ( A ) High expression of UHRF2 and low expression of E-cadherin, and low expression of UHRF2 and high expression of E-cadherin are shown. Scale bar: 100, 200 μm. ( B ) The relationship between UHRF2 and E-cadherin in ICC patients; the protein expression of UHRF2 was significantly negatively correlated with E-cadherin expression. ( C ) UHRF2 and E-cadherin protein were detected by Western blot analysis. ( D ) Representative immunofluorescent images of UHRF2 and E-cadherin in ICC cell lines are shown. Scale bars =100 μm. * P <;0.05, ** P <;0.01, *** P <;0.001. Abbreviations: H

Article Snippet: Rabbit anti-human UHRF2 monoclonal antibody (1:1,000; Wuhan Boster Biological Technology, Ltd., Wuhan, People’s Republic of China) and rabbit anti-human E-cadherin antibody (1:100; Cell Signaling Technology, Danvers, MA, USA) were utilized to detect the levels of UHRF2 and E-cadherin.

Techniques: Expressing, Western Blot, Ubiquitin Proteomics

Journal: OncoTargets and therapy

Article Title: Overexpression of UHRF2 in intrahepatic cholangiocarcinoma and its clinical significance

doi: 10.2147/OTT.S149361

Figure Lengend Snippet: Prognostic significance was assessed by log-rank tests and Kaplan–Meier analysis. Notes: ( A – D ) Representative graphs of H

Article Snippet: Rabbit anti-human UHRF2 monoclonal antibody (1:1,000; Wuhan Boster Biological Technology, Ltd., Wuhan, People’s Republic of China) and rabbit anti-human E-cadherin antibody (1:100; Cell Signaling Technology, Danvers, MA, USA) were utilized to detect the levels of UHRF2 and E-cadherin.

Techniques: Immunohistochemical staining, Staining, Expressing, Ubiquitin Proteomics

Journal: OncoTargets and therapy

Article Title: Overexpression of UHRF2 in intrahepatic cholangiocarcinoma and its clinical significance

doi: 10.2147/OTT.S149361

Figure Lengend Snippet: Correlations between UHRF2 with clinicopathologic features in 139 ICC patients

Article Snippet: Rabbit anti-human UHRF2 monoclonal antibody (1:1,000; Wuhan Boster Biological Technology, Ltd., Wuhan, People’s Republic of China) and rabbit anti-human E-cadherin antibody (1:100; Cell Signaling Technology, Danvers, MA, USA) were utilized to detect the levels of UHRF2 and E-cadherin.

Techniques:

Journal: OncoTargets and therapy

Article Title: Overexpression of UHRF2 in intrahepatic cholangiocarcinoma and its clinical significance

doi: 10.2147/OTT.S149361

Figure Lengend Snippet: Univariate and multivariate analyses of factors associated with recurrence and survival

Article Snippet: Rabbit anti-human UHRF2 monoclonal antibody (1:1,000; Wuhan Boster Biological Technology, Ltd., Wuhan, People’s Republic of China) and rabbit anti-human E-cadherin antibody (1:100; Cell Signaling Technology, Danvers, MA, USA) were utilized to detect the levels of UHRF2 and E-cadherin.

Techniques:

Journal: Frontiers in Molecular Neuroscience

Article Title: Whole Genome Expression Analysis in a Mouse Model of Tauopathy Identifies MECP2 as a Possible Regulator of Tau Pathology

doi: 10.3389/fnmol.2017.00069

Figure Lengend Snippet: Microarray analysis of the hippocampi of 6-month-old WT and hTau MaptKO ( Duke ) mice. (A) Total RNA from hippocampi of 6-month-old WT and hTau MaptKO ( Duke ) mice was hybridized to MouseWG-6 v2.0 Expression BeadChip (Illumina ® ). Expression of 64 genes that were significantly altered in hTau MaptKO ( Duke ) mice compared to WT is shown. Values displayed as fold change in expression level: up-regulated (red) and down-regulated (blue) genes. Data represents mean fold change from three mice per genotype. (B) Gene interaction network analysis (using Metacore analytical suite) for the 64 significantly altered genes in hTau MaptKO ( Duke ) mice compared to WT mice. The genes with red (or blue) next to their graphic key is either up- (or down-) regulated. As expected, endogenous mouse tau (MAPT) is one of the most down-regulated genes. Other genes relevant to neuronal function or neurological disease include Prkca, Mecp2, Strn4, Slc40a1, Pold2, Pcsk2 (up-regulated) and Krt12, Lass1, Plat and Nrxn1 (down-regulated). Many of the altered genes are regulated by the transcription factor SP1. (C) Venn diagrams showing GO’s biological processes when all 64 genes are categorized for top biological processes segregate into three distinct Venn diagrams: Group 1: anatomical structure development – 17 altered genes out of total 47, i.e., 17/47; cellular nitrogen compound metabolic process – 15/47; biosynthetic process 13/47. Group 2: cell death – 9/47; signal transduction 8/47; response to stress 8/47. Group 3: neurological system 5/47, cell cycle 2/47 and immune system processes 4/47. Note the genes written in red (or blue) are significantly up- (or down-) regulated.

Article Snippet: Other antibodies: GAPDH (rabbit polyclonal antibody; EMD Millipore; #ABS16), T-MECP2 (D4F3; rabbit monoclonal antibody; Cell Signaling #3456); phospho-MECP2 (pSer80; rabbit polyclonal antibody; PhosphoSolutions #p1205-80).

Techniques: Microarray, Expressing, Transduction

Journal: Frontiers in Molecular Neuroscience

Article Title: Whole Genome Expression Analysis in a Mouse Model of Tauopathy Identifies MECP2 as a Possible Regulator of Tau Pathology

doi: 10.3389/fnmol.2017.00069

Figure Lengend Snippet: MECP2 expression and phosphorylation is up-regulated in 6-month-old hTau MaptKO ( Duke ) mice. (A) qRT-PCR analysis showing statistically significant ( ∗ p < 0.05; unpaired t- test; n = 4 WT, and n = 4 for hTau MaptKO ( Duke ) mice; mean + SEM) up-regulation of MECP2 in the hemi-brains of 6-month-old hTau MaptKO ( Duke ) vs. WT mice. (B) Note the regional differences in the expression of MECP2 in the hippocampus (HIP), cortex (CX), and rest of the brain (ROB) that are devoid of CX and HP. (C) Another gene ( Vat1l ) that was increased in our whole genome microarray analysis also showed modest up-regulation in its mRNA levels in the brains of hTau MaptKO ( Duke ) mice compared to age-matched WT mice. (D,E) Western blot analysis showing a trend toward increased levels for phospho(p)-Ser80 MECP2/total (t) MECP2 and tMECP2/GAPDH [ p = 0.08; unpaired t- test; n = 3, all females for WT and n = 10, three females and six males for hTau MaptKO ( Duke ) ] in 6-month-old hTau MaptKO ( Duke ) versus WT mice; mean + SEM). (F) Double IF and confocal microscopy analysis revealing a modest increase in pMECP2 in the CA3 region of HP in 6-month-old hTau MaptKO ( Duke ) mice compared to age-matched WT controls. Scale bar 25 μm.

Article Snippet: Other antibodies: GAPDH (rabbit polyclonal antibody; EMD Millipore; #ABS16), T-MECP2 (D4F3; rabbit monoclonal antibody; Cell Signaling #3456); phospho-MECP2 (pSer80; rabbit polyclonal antibody; PhosphoSolutions #p1205-80).

Techniques: Expressing, Phospho-proteomics, Quantitative RT-PCR, Microarray, Western Blot, Confocal Microscopy

Journal: Frontiers in Molecular Neuroscience

Article Title: Whole Genome Expression Analysis in a Mouse Model of Tauopathy Identifies MECP2 as a Possible Regulator of Tau Pathology

doi: 10.3389/fnmol.2017.00069

Figure Lengend Snippet: MECP2 phosphorylation is up-regulated in 12-month-old hTau MaptKO ( Duke ) mice and in human AD brain. (A,B) Western blot analysis showing pMECP2/GAPDH ratio significantly higher [ ∗ p < 0.05; unpaired t- test; n = 3 for hTau MaptKO ( Duke ) versus WT mice; all three males for WT; two males and one female for hTau MaptKO ( Duke ) ; mean + SEM] in the hippocampus of 12-month-old hTau MaptKO ( Duke ) mice compared to WT controls. No alteration in the tMECP2/GAPDH ratio in the hippocampus of 12-month-old hTau MaptKO ( Duke ) mice compared to controls. (C) Significantly elevated pMECP2 immunoreactive specks within the nucleus of CA3 hippocampal neurons of 12-month-old hTau MaptKO ( Duke ) mice compared to age-matched WT controls. (D) Double IF and confocal microscopy analysis shows a modest increase in the pMECP2 in the CA3 neuronal layer of 12-month-old hTau MaptKO ( Duke ) mice compared to age-matched WT controls. (E–G) Confocal projections (in E,G ) and bright field images show elevated pMECP2 immunoreactivity and co-localization with nuclear stain DAPI (in E,G ) or peri-vascular labeling (in F ) specifically in the Layer III of temporal cortex of human AD brain autopsy sections compared to the to non-AD healthy control subject. Orthogonal view in (G) shows pMECP2 labeling to be nuclear or peri-nuclear in the human AD cortex. Scale bar 10 μm (in C,F ); 25 μm (in D,G ); 100 μm (in E ).

Article Snippet: Other antibodies: GAPDH (rabbit polyclonal antibody; EMD Millipore; #ABS16), T-MECP2 (D4F3; rabbit monoclonal antibody; Cell Signaling #3456); phospho-MECP2 (pSer80; rabbit polyclonal antibody; PhosphoSolutions #p1205-80).

Techniques: Phospho-proteomics, Western Blot, Confocal Microscopy, Staining, Labeling, Control

Journal: Frontiers in Molecular Neuroscience

Article Title: Whole Genome Expression Analysis in a Mouse Model of Tauopathy Identifies MECP2 as a Possible Regulator of Tau Pathology

doi: 10.3389/fnmol.2017.00069

Figure Lengend Snippet: MECP2 regulates tau pathology in vitro . N2a cells transiently transfected with human tau 0N3R isoform (‘+Tau’) or a control plasmid (‘-Tau’) were nucleofected with siRNA [scramble siRNA (siScr) or MECP2 siRNA]. After 24 h of siRNA nucleofection, the cells were harvested and probed for AT180, PHF-1, Tau5, Tau12, and T-MECP2. (A,B) Note that siMECP2 significantly reduced levels of MECP2 in both ‘-Tau’ and ‘+Tau’ N2a cells ( ∗ p < 0.01; unpaired t- test; n = 3 replicates; mean + SEM). (C,D) siMECP2 treatment also significantly ( ∗ p < 0.01; unpaired t- test; n = 3 replicates; mean + SEM) reduced the levels of both total tau (Tau5/GAPDH) and human tau (Tau12/GAPDH) ratios in the ‘+Tau’ N2a cells compared to scramble siRNA treated conditions. (E–H) siMECP2 knockdown resulted in statistically significant ( ∗ p < 0.01; unpaired t- test; n = 3 replicates; mean + SEM) increase and decrease in AT180/Tau5 and PHF1/Tau5 ratios, respectively. Note that the ratio for β-actin/GAPDH was not altered either in ‘-Tau’/‘+Tau’ conditions or with/without siMECP2 conditions.

Article Snippet: Other antibodies: GAPDH (rabbit polyclonal antibody; EMD Millipore; #ABS16), T-MECP2 (D4F3; rabbit monoclonal antibody; Cell Signaling #3456); phospho-MECP2 (pSer80; rabbit polyclonal antibody; PhosphoSolutions #p1205-80).

Techniques: In Vitro, Transfection, Control, Plasmid Preparation, Knockdown