Journal: Molecular Neurobiology

Article Title: A Single Prior Injection of Methamphetamine Enhances Methamphetamine Self-Administration (SA) and Blocks SA-Induced Changes in DNA Methylation and mRNA Expression of Potassium Channels in the Rat Nucleus Accumbens

doi: 10.1007/s12035-019-01830-3

Figure Lengend Snippet: Increased protein levels of ( a ) KCNA1, ( b ) KCNA3, and ( c ) KCNN1 in the NAc of SM rats. Data are presented as means ± SEM of fold changes relative to the controls. Key to statistics: * P < 0.05, ** P < 0.01, *** P < 0.001 in comparison to controls (SS group); # P < 0.05, ## P < 0.01 in comparison to SM rats

Article Snippet: Subsequently, the membranes were incubated overnight at 4 °C with specific antibodies against KCNA1(Abcam, catalog # ab32433), KCNA3 (Santa Cruz, catalog # sc-398,855) and KCNN1 (Abcam, catalog # ab66624).

Techniques: Comparison

Journal: Molecular Neurobiology

Article Title: A Single Prior Injection of Methamphetamine Enhances Methamphetamine Self-Administration (SA) and Blocks SA-Induced Changes in DNA Methylation and mRNA Expression of Potassium Channels in the Rat Nucleus Accumbens

doi: 10.1007/s12035-019-01830-3

Figure Lengend Snippet: METH SA caused decreased DNA methylation in the NAc of SM rats. (a) DNA methylation at CpG-rich sites near the promoter of Kcna1 voltage-gated K + channels in the NAc of SM and MM rats. ( b ) DNA methylation at the Kcna3 promoter showed a decreasing trend ( p = 0.0548) in the NAc of SM rats in comparison to control rats; pre-exposure to METH normalized DNA methylation. ( c ) Decreased DNA methylation of calcium-activated K + channel, Kcnn1, in the NAc of SM rats and this was also reversed in the MM rats. Key to statistics: ***P < 0.001 in comparison to controls (SS group); ## P < 0.01 in comparison to SM rats

Article Snippet: Subsequently, the membranes were incubated overnight at 4 °C with specific antibodies against KCNA1(Abcam, catalog # ab32433), KCNA3 (Santa Cruz, catalog # sc-398,855) and KCNN1 (Abcam, catalog # ab66624).

Techniques: DNA Methylation Assay, Comparison, Control

Journal: Biomedicines

Article Title: Increased ACh-Associated Immunoreactivity in Autonomic Centers in PTZ Kindling Model of Epilepsy

doi: 10.3390/biomedicines8050113

Figure Lengend Snippet: Examples of immunoreactivity for ACh receptors and Kir3.1 channel within the brainstem. Immunoreactivity was significantly increased for both ACh-associated centers in the represented images, the result for male rats was prepared. For female ( A ) and male ( B ) in terms of immunoreactivity at pons; A : Image of M2 receptors for control groups, B : Image of M2 receptors for PTZ-kindled of epilepsy, C : Image of Kir3.1 channels for control groups, D : Image of Kir3.1 channels of PTZ-kindled epilepsy. Pictures were taken at a magnification of ×200. Scale bar: 50 μm. Fold change in immunoreactivity for M2 receptors and Kir3.1 channel within the brainstem ( C ). For male, PTZ-kindled rats, 3.8-fold change in the M2 receptor and 2.7-fold change in Kir3.1 channel were observed when compared to the controls. For female PTZ-kindled rats, there was a 2.1-fold change in M2 receptors and 2.2-fold change in Kir3.1 channel when compared to the controls, * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Immediately, Kir3.1 (mouse monoclonal, 1:200, Alomone Labs, Jerusalem, Israel [APC-005]) and M2 receptor (rabbit polyclonal, 1:200, Alomone Labs, Jerusalem, Israel [APC-002]) primary antibodies were dropped and kept at +4 °C for one night and incubated the next day for 20 min. As a negative control, PBS was used instead of the primary antibody.

Techniques: Control

Journal: Biomedicines

Article Title: Increased ACh-Associated Immunoreactivity in Autonomic Centers in PTZ Kindling Model of Epilepsy

doi: 10.3390/biomedicines8050113

Figure Lengend Snippet: Immunoreactivity results of Kir3.1 channel and M2 receptors in the VN. In both groups, increased immunoreactivity in cervical VN was observed. For female groups; A : Image of cervical VN of control groups, B : Image of thoracic VN of control groups, C : Image of cervical VN of PTZ-kindled epilepsy, D : Image of thoracic VN of PTZ-kindled epilepsy. For male groups; E : Image of cervical VN of control groups, F : Image of thoracic VN of control groups, G : Image of cervical VN of PTZ-kindled epilepsy, H : Image of thoracic VN of PTZ-kindled epilepsy. Pictures were taken at a magnification of ×200. Scale bar: 50 μm ( A ). Fold change in immunoreactivity for M2 receptors and Kir3.1 channel in the VN ( B ). For cervical VN, in terms of the immunoreactivity of the M2 receptor, a 1.6-fold change in male and a 1.8-fold change in female rats compared to controls can be seen; for cervical VN, in terms of the immunoreactivity of Kir3.1, a 2.2-fold change in male and a 2.4-fold change in female rats compared to thoracic VN can be seen, * p < 0.05, ** p < 0.01.

Article Snippet: Immediately, Kir3.1 (mouse monoclonal, 1:200, Alomone Labs, Jerusalem, Israel [APC-005]) and M2 receptor (rabbit polyclonal, 1:200, Alomone Labs, Jerusalem, Israel [APC-002]) primary antibodies were dropped and kept at +4 °C for one night and incubated the next day for 20 min. As a negative control, PBS was used instead of the primary antibody.

Techniques: Control

Journal: Biomedicines

Article Title: Increased ACh-Associated Immunoreactivity in Autonomic Centers in PTZ Kindling Model of Epilepsy

doi: 10.3390/biomedicines8050113

Figure Lengend Snippet: Examples of immunoreactivity for M2 receptors and Kir3.1 channel within the heart tissue (atria). Immunoreactivity was significantly increased for both ACh-associated centers and Kir3.1 channel, as demonstrated in the represented images. Results for female rats are demonstrated in section A , whereas for male rats they are demonstrated in section B . Pictures were taken at a magnification of ×200. Scale bar: 50 μm. Fold change in immunoreactivity for M2 receptors and Kir3.1 channel within the heart tissue ( C ). For female PTZ-kindled rats, there was a 2.1-fold change in M2 and a 6.1-fold change in the Kir3.1 channel when compared to the controls. On the other hand, for male PTZ-kindled rats, there was a 3.8-fold change in M2 receptors and a 3.7-fold change in the Kir3.1 channel when compared to the controls, * p < 0.05.

Article Snippet: Immediately, Kir3.1 (mouse monoclonal, 1:200, Alomone Labs, Jerusalem, Israel [APC-005]) and M2 receptor (rabbit polyclonal, 1:200, Alomone Labs, Jerusalem, Israel [APC-002]) primary antibodies were dropped and kept at +4 °C for one night and incubated the next day for 20 min. As a negative control, PBS was used instead of the primary antibody.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: Six2 Is a Coordinator of LiCl-Induced Cell Proliferation and Apoptosis

doi: 10.3390/ijms17091504

Figure Lengend Snippet: LiCl promotes cell proliferation in mK3 and mK4 cells. ( A ) mK3 cells were treated with LiCl of increasing dosages (0, 10, 20, 30, 40, and 50 mM) for 12 h and performed with 5-ethynyl-20-deoxyuridine (EdU) assays. Proliferating mK3 cells were labeled with EdU (red) and cell nucleuses were stained with DAPI (blue). The EdU results were accessed by fluorescent microscope (200×) with the scale bar representing 20 µm and the respective pictures were merged to the purple one; ( B ) Statistical analysis of mK3 cell proliferation. Values were presented as mean ± SEM ( n = 3). p -values were calculated by Student t -test, ** p < 0.01 relative to control; ( C ) mK4 cells were treated in the same way as in ( A ) and the EdU assay were conducted; ( D ) Statistical analysis of mK4 cell proliferation. Values were presented as mean ± SEM ( n = 3). p -values were calculated by Student t -test, ** p < 0.01 relative to control.

Article Snippet: mK3 and mK4 cells were cultured in 6-well plates for 24 h and treated with LiCl of increasing dosages for 12 h. Furthermore, the Western blot assays was processed with antibodies Six2 (1:600, proteintech, Chicago, IL, USA) and internal control β-tubulin (1:5000, proteintech) followed by the research “Identification of a thyroid microsomal antigen by Western blot and immune-precipitation” [ ].

Techniques: Labeling, Staining, Microscopy, Control, EdU Assay

Journal: International Journal of Molecular Sciences

Article Title: Six2 Is a Coordinator of LiCl-Induced Cell Proliferation and Apoptosis

doi: 10.3390/ijms17091504

Figure Lengend Snippet: LiCl activates Wnt and BMP signaling pathway. ( A ) mK3 cells were treated with LiCl of increasing dosages (0, 10, 20 M, 30, 40, and 50 mM) for 12 h. The expression Wnt and BMP Signal markers were detected for confirming the function of LiCl by RT-PCR; ( B ) The relative mRNA expressions in mK3 cells was quantified by gray scan, normalized to the internal control 18S. Values were presented as mean ± SEM ( n = 3). p -values were calculated by Student t -test, ** p < 0.01, *** p < 0.001 relative to control; ( C ) mK4 cells were treated same with mK3 cells and the mRNA expression of the same genes was tested by RT-PCR; ( D ) The relative mRNA expressions in mK4 cells were quantified by gray scan, normalized to the internal control 18S. Values were presented as mean ± SEM ( n = 3). p -values were calculated by Student t -test, * p < 0.05, ** p < 0.01, *** p < 0.001 relative to control.

Article Snippet: mK3 and mK4 cells were cultured in 6-well plates for 24 h and treated with LiCl of increasing dosages for 12 h. Furthermore, the Western blot assays was processed with antibodies Six2 (1:600, proteintech, Chicago, IL, USA) and internal control β-tubulin (1:5000, proteintech) followed by the research “Identification of a thyroid microsomal antigen by Western blot and immune-precipitation” [ ].

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Control

Journal: International Journal of Molecular Sciences

Article Title: Six2 Is a Coordinator of LiCl-Induced Cell Proliferation and Apoptosis

doi: 10.3390/ijms17091504

Figure Lengend Snippet: LiCl regulates the expression of Six2 at mRNA and protein level. ( A ) pGL3- Six2 promoter-Luciferase construction was simulate by diagram. The Six2 promoter ranging from −2322 to −323 (Six genome sequence) was obtained from NCBI; ( B ) HEK293T cells were co-transfected with pRL-SV40 (renilla control) and pGL3- Six2 promoter-LuC for 36 h. Luciferase activity was normalized to Renilla control. p -values were calculated by Student t -test. Values represents mean values ± SEM of triplicate experiments, *** p < 0.001 relative to control; ( C ) HEK293T cells were co-transfected with pRL-SV40 (renilla control) and pGL3- Six2 -LuC for 36 h then were treated with LiCl of increasing dosages for 12 h. Luciferase activity was measured using dual luciferase reporter assay, normalized to Renilla control. Values were presented as mean ± SEM ( n = 3), *** p < 0.001 relative to control; ( D ) mK3 cells were treated with LiCl of increasing dosages for 12 h. The Six2 expression at protein level was tested by Western-blot. Values were presented as mean ± SEM ( n = 3), ** p < 0.01 relative to control.

Article Snippet: mK3 and mK4 cells were cultured in 6-well plates for 24 h and treated with LiCl of increasing dosages for 12 h. Furthermore, the Western blot assays was processed with antibodies Six2 (1:600, proteintech, Chicago, IL, USA) and internal control β-tubulin (1:5000, proteintech) followed by the research “Identification of a thyroid microsomal antigen by Western blot and immune-precipitation” [ ].

Techniques: Expressing, Luciferase, Sequencing, Transfection, Control, Activity Assay, Reporter Assay, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Six2 Is a Coordinator of LiCl-Induced Cell Proliferation and Apoptosis

doi: 10.3390/ijms17091504

Figure Lengend Snippet: Knockdown of Six2 gene inhibits cell proliferation while LiCl treatment of low-concentration promotes cell proliferation in mK3 and mK4 cells. ( A ) mK3 cells were transfected with negative shRNA control and Six2 -shRNA for 36 h and treated with LiCl of increasing dosages for 12 h. Proliferating mK3 cells were labeled with EdU (red) and cell nucleus were stained with hoechst (blue). The EdU results were accessed by fluorescent microscope (200×) with the scale bar representing 20 μm and the respective pictures were merged to the purple one; ( B , C ) Statistical analysis of cell proliferation. Values were presented as mean ± SEM ( n = 3), ** p < 0.01, *** p < 0.001 relative to control; ( D ) mK4 cells were transfected and were detected by EdU assay as same as mK3 cells in ( A ); ( E , F ) Statistical analysis of cell proliferation. Values were presented as mean ± SEM ( n = 3), ** p < 0.01 relative to control.

Article Snippet: mK3 and mK4 cells were cultured in 6-well plates for 24 h and treated with LiCl of increasing dosages for 12 h. Furthermore, the Western blot assays was processed with antibodies Six2 (1:600, proteintech, Chicago, IL, USA) and internal control β-tubulin (1:5000, proteintech) followed by the research “Identification of a thyroid microsomal antigen by Western blot and immune-precipitation” [ ].

Techniques: Knockdown, Concentration Assay, Transfection, shRNA, Control, Labeling, Staining, Microscopy, EdU Assay

Journal: International Journal of Molecular Sciences

Article Title: Six2 Is a Coordinator of LiCl-Induced Cell Proliferation and Apoptosis

doi: 10.3390/ijms17091504

Figure Lengend Snippet: Knockdown of Six2 gene accelerates cell apoptosis while LiCl treatment of low-concentration inhibits cell apoptosis in mK3 cells. ( A ) mK3 cells were transfected with negative shRNA control and Six2 -shRNA for 36 h and treated with LiCl of increasing dosages. The apoptosis was detected by FCM; ( B ) Statistical analysis of cell apoptosis and histogram was drawn in GraphPad Prism 5; ( C ) The efficiency of knockdown Six2 at mRNA and protein level, compared with internal control 18S and β-tubulin, respectively. Values were presented as mean ± SEM ( n = 3), *** p < 0.001 relative to control.

Article Snippet: mK3 and mK4 cells were cultured in 6-well plates for 24 h and treated with LiCl of increasing dosages for 12 h. Furthermore, the Western blot assays was processed with antibodies Six2 (1:600, proteintech, Chicago, IL, USA) and internal control β-tubulin (1:5000, proteintech) followed by the research “Identification of a thyroid microsomal antigen by Western blot and immune-precipitation” [ ].

Techniques: Knockdown, Concentration Assay, Transfection, shRNA, Control

Journal: Cell Reports Medicine

Article Title: A target discovery pipeline identified ILT3 as a target for immunotherapy of multiple myeloma

doi: 10.1016/j.xcrm.2023.101110

Figure Lengend Snippet:

Article Snippet: KCNA3 , Prosci , 57–918.

Techniques: Enzyme-linked Immunosorbent Assay, Isolation, Recombinant, Magnetic Beads, Software, CRISPR