Journal: Human Reproduction (Oxford, England)

Article Title: Chronic hyperandrogenemia in the presence and absence of a western-style diet impairs ovarian and uterine structure/function in young adult rhesus monkeys

doi: 10.1093/humrep/dex338

Figure Lengend Snippet: Photomicrographs illustrating immunohistochemical staining for estrogen receptor 1 (ESR1), progesterone receptor (PGR) MMP26, TIMP3, Ki-67 and androgen receptor (AR) in the endometrial functionalis zone of the macaque uterus from representative females in each treatment group (C, T, WSD, T+WSD). Brown staining denotes positive expression of proteins. Sections are counterstained with hematoxylin (blue) staining. ESR1, PGR, Ki-67 and AR staining is nuclear, while MMP26 and TIMP3 show cytoplasmic localization. Inset shows a negative control with an irrelevant antibody (Anti-Br(d)U). TIMP3 staining was localized to the predecidual cells around the spiral arteries.

Article Snippet: Antibodies used were against estrogen receptor 1 (ESR1,ER-ID5; Cat#: MS-354-P, Thermo Fisher Scientific), progesterone receptor (PGR, Cat#: Ms-298-P, 1 μg, Lab Vision/NeoMarkers, Fremont, CA, USA), androgen receptor (AR-F39 (Cat#: MU256, 1/50, BioGenex, Fremont, CA, USA), Ki67 (Cat#: MU370-UC, 1/200, BioGenex), MMP26 (Cat# ab57636; Abcam, Cambridge, MA, USA) and TIMP3 (Cat#:MAB973, R&D Systems, Inc. Minneapolis, MN, USA).

Techniques: Immunohistochemical staining, Staining, Expressing, Negative Control

Journal: Human Reproduction (Oxford, England)

Article Title: Chronic hyperandrogenemia in the presence and absence of a western-style diet impairs ovarian and uterine structure/function in young adult rhesus monkeys

doi: 10.1093/humrep/dex338

Figure Lengend Snippet: Expression of ESR1, PGR, MMP26 and TIMP3 mRNAs as detected by qRT-PCR in endometrial biopsies collected from macaques in the mid-luteal phase of the cycle. Significant (P < 0.05) differences between individual treatment groups are denoted by different uppercase letters above columns.

Article Snippet: Antibodies used were against estrogen receptor 1 (ESR1,ER-ID5; Cat#: MS-354-P, Thermo Fisher Scientific), progesterone receptor (PGR, Cat#: Ms-298-P, 1 μg, Lab Vision/NeoMarkers, Fremont, CA, USA), androgen receptor (AR-F39 (Cat#: MU256, 1/50, BioGenex, Fremont, CA, USA), Ki67 (Cat#: MU370-UC, 1/200, BioGenex), MMP26 (Cat# ab57636; Abcam, Cambridge, MA, USA) and TIMP3 (Cat#:MAB973, R&D Systems, Inc. Minneapolis, MN, USA).

Techniques: Expressing, Quantitative RT-PCR

Journal: Oncotarget

Article Title: Long noncoding RNA DANCR promotes invasion of prostate cancer through epigenetically silencing expression of TIMP2/3

doi: 10.18632/oncotarget.9350

Figure Lengend Snippet: ( A ) Effect of DANCR knockdown on the expression of thirty-seven invasion associated genes in C4-2B cells, as detected by RT-qPCR assay. ( B , C ) Knockdown of DANCR increases the expression of TIMP2/3 mRNA and protein in C4-2B and CW22RV1 cells, as detected by RT-qPCR assay and western blot analysis. ( D ) Representation of the TImP2/3 promoter region as mapped by PCR analysis and ChIP assay. The bent arrow represents the transcription start sites (+1). The lines below the TIMP2/3 locus represent the regions amplified by PCR. ( E ) Knockdown of DANCR decreased the binding of EZH2 on the promoter of TIMP2/3. Immuno-precipitated DNA was analyzed by PCR with specific primer sets. Chromatin obtained from C4-2B and CW22RV1 cells were immune-precipitated using antibodies to EZH2, histone H3 (H3) and IgG. Each ChIP experiment was repeated at least three times and a representative experiment is shown. ( F ) Knockdown of DANCR decreased the tri-methyl-histone H3K27 on the promoter of TIMP2/3. Immuno-precipitated DNA was analyzed by PCR with specific primer sets. Chromatin obtained from C4-2B and CW22RV1cells were immune-precipitated using antibodies to tri-methyl-histone H3K27 (3meH3K27), histone H3 (H3) and IgG. Each experiment was repeated at least three times and a representative experiment is shown. ( G ) Knockdown of DANCR decreased the binding of EZH2 on the promoter of TIMP2/3, as detected by oligonucleotides pull down assays. P1: TIMP2-set2; P2: TIMP2-set3; P3: TIMP3-set1; P4: TIMP3-set2; P5: TIMP3-set3. ( H ) DANCR was pulled down by the oligonucleotides with the same sequence of the promoters of TIMP2/3 (P1-P5). Control: The DNA solution TE buffer was used as negative control.

Article Snippet: Then membranes were blocked with 5% skim milk at room temperature for 1 hour and then incubated with primary antibodies against GAPDH (Kangchen, Shanghai, China), TIMP2, TIMP3 and androgen receptor (AR) (Santa Cruz, Dallas, TX, USA) at 4°C overnight, followed by TBST wash and 1 hour incubation with horseradish peroxidase-conjugated secondary antibodies at room temperature.

Techniques: Knockdown, Expressing, Quantitative RT-PCR, Western Blot, Amplification, Binding Assay, Sequencing, Control, Negative Control