Journal: Brain structure & function

Article Title: The α7 nicotinic acetylcholine receptors regulate hippocampal adult-neurogenesis in a sexually dimorphic fashion

doi: 10.1007/s00429-018-1799-6

Figure Lengend Snippet: Effect of loss of α7 nAChRs on maturation of neurons. a The distance that immature neurons had moved in from the hilus was measured to determine if maturation was increasing. To do this, maximum projection images were analyzed in ImageJ by marking all visible cell bodies also stained with DCX. When the edge of the image was aligned with the edge of the hilus, the inward movement of cells could be measured. b No difference was seen in knock-out mice of either sex in average inward mobility of immature neurons. c There was no difference in the percent of cells that had moved inward from the edge of the hilus by more than 10 μm in male or female mice of either genotype. d–f In order to examine the effect of loss of α7 nAChRs on dendritic structure, 63X images were captured and analyzed using Imaris Filament function. These images delineate the process. d A region of interest was randomly selected from the upper arm of the dentate gyrus in the dorsal region of the hippocampus and imaged at 63X. e Five DCX cells were randomly selected from all cells present in the image. Cells were not selected if the dendritic structure traveled out of the plane created by the section. Cells were manually traced using the filament function on Imaris. Cells were checked for accuracy in 3D. f Tracings were used to count tertiary filaments and measure overall filament length. g There was no significant difference in the number of tertiary dendrites (p = 0.068). h Compared with wild-type mice, mice that lacked the α7 nAChR had an overall increase in total filament length [F(1,22) = 5.051, p = 0350, n = 6 mwt, 6 fwt, 7 mko, 7 fko] when both sexes are considered together. Statistical analysis uses two-way ANOVA with Sidak’s multiple comparison test and Graphpad Prism 7.0 Software. Experimental groups contained 5–7 mice per group including male wild type, male knock-out, female wild type, female knock-out. In each mouse, four hippocampi sections were analyzed to provide a mean density per mouse. Data expressed as mean ± SEM. Asterisk indicates *p < 0.05

Article Snippet: Furthermore, prenatal stress decreases adult neurogenesis ( Belnoue et al. 2013 ), and also affects the levels of α7 nAChRs in adult rodents ( Baier et al. 2015 ).

Techniques: Staining, Knock-Out, Comparison, Software

Journal: Brain structure & function

Article Title: The α7 nicotinic acetylcholine receptors regulate hippocampal adult-neurogenesis in a sexually dimorphic fashion

doi: 10.1007/s00429-018-1799-6

Figure Lengend Snippet: Loss of α7 nAChRs impairs performance in DNTP task in male mice and results in lost sexual dimorphism. Schematic (a) and timeline (b) of DNTP task. Mice were kept in 12 h light:12 h dark reverse light dark and placed on restricted diet beginning 10 days prior to experiment, and handled and weighed daily during this time. Prior to the experiment, mice were habituated to the room and maze. Habituation included 8 min free exploration of the maze with reward pellets at the end of each arm, and 2 min restricted exploration of the maze with reward pellets at the end of 2 open arms. During the two-phase task, mice were placed in one arm of an eight-arm radial maze and retrieved a reward pellet placed in another arm (sample arm). After a short delay, a novel arm was loaded with a reward pellet. Mice were placed in the maze in the original arm and allowed to retrieve the treat. We measured whether the mouse chose the novel arm or the familiar arm to enter first. c Our results show that there is a sex by genotype interaction in this task (p = 0.0025, n = 53), with wild-type males preferring the novel arm while females did not (p = 0.0009, n = 10 m, 8 f). This sexual dimorphism disappears when the α7 nAChR is knocked out (p = 0.9957; n = 19 m, 16 f). Transgenic male mice performed significantly closer to chance than wild-type males (p = 0.0408, n = 10 wt, 19 tg) while females do not show a significant change (p = 0.2780, n = 8 wt 16 tg) in their choice of novel or familiar arm. Asterisk indicates *p < 0.05, **p < 0.01, ***p < 0.001. Statistical analysis using a two-way ANOVA and Sidak’s post hoc using SPSS and Graphpad Prism 7.0 software. Data expressed as mean ± SEM

Article Snippet: Furthermore, prenatal stress decreases adult neurogenesis ( Belnoue et al. 2013 ), and also affects the levels of α7 nAChRs in adult rodents ( Baier et al. 2015 ).

Techniques: Transgenic Assay, Software

Journal: Brain structure & function

Article Title: The α7 nicotinic acetylcholine receptors regulate hippocampal adult-neurogenesis in a sexually dimorphic fashion

doi: 10.1007/s00429-018-1799-6

Figure Lengend Snippet: Loss of α7 nAChRs in nestin+ cells results in decreased NSCs in males. a Administering tamoxifen to nestin-CreER/T2 × ROSAtdTomato × α7 nAChRflox male mice results in the loss of α7 nAChR expression in nestin+ NSCs and their progeny of male mice homozygous for floxed α7 nAChR, as well as the expression of tdTomato expression in nestin+ cells and their progeny. In this experiment females were not tested. b Male mice were dosed with tamoxifen through intraperitoneal injection in 5 consecutive doses (0.15 mg/g) at 8 weeks of age. Mice then aged 9 weeks and were euthanized at 17 weeks, allowing sufficient time for α7 nAChR−/nestin+ cells to divide, differentiate and mature into the dentate gyrus granule cell layer. c 3-D images were collected of dentate gyrus of α7 nAChRflox × nestin-Cre/ERT2 × ROSATdTomato wild-type and knock-out mice euthanized 9 weeks after final tamoxifen injection. This delay allows activated NSCs to divide and mature without α7 nAChR expression. Images show NSCs, immature neurons (gp-antiDCX 1:500, Gt-antiGp Alexa 488 1:500), and granule cells (Rb-antiProx1 1:500, Gt-antiRb Alexa 647 1:500), as well as DAPI (2 μg/ml). Scale bar: 10 μm. d The NSC population was significantly decreased (p = 0.0416, n = 5, 5). This is comparable to the result seen in whole body knock-out of the α7 nAChR, in which there is an approximate 50% decline in NSCs in the knock-out compared to the wild type. e The percent of NSCs compared to the overall population of cells in the dentate gyrus was significantly decreased (p = 0.0468, n = 5, 5). f There were no significant differences in numbers of immature neurons in these mice (p = 0.495, n = 5, 5). This is in contrast to the whole body knock-out of α7 nAChR, where male mice showed a significant increase in immature neurons. Female mice were not analyzed. Asterisk indicates *p < 0.05. Statistical analysis with unpaired Student’s t test calculated using Graphpad Prism 7.0. Data in bar graph expressed as mean ± SEM

Article Snippet: Furthermore, prenatal stress decreases adult neurogenesis ( Belnoue et al. 2013 ), and also affects the levels of α7 nAChRs in adult rodents ( Baier et al. 2015 ).

Techniques: Expressing, Injection, Knock-Out

Journal: Brain structure & function

Article Title: The α7 nicotinic acetylcholine receptors regulate hippocampal adult-neurogenesis in a sexually dimorphic fashion

doi: 10.1007/s00429-018-1799-6

Figure Lengend Snippet: Loss of α7 nAChRs in nestin+ cells results in greater inward mobility in males. a The average distance inward from the hilus edge that cells had traveled was measured by taking a maximum intensity projection and importing it into ImageJ. b Results indicated that loss of the α7 nAChR in NSCs was associated with greater inward mobility of these cells from the hilus prior to their maturation into adult neurons (p = 0.0288, n = 6,6). c Loss of α7 nAChRs from NSCs results in a larger percentage of cells that have moved more than 10 μm from the edge of the hilus as immature neurons (p = 0.0168, n = 6,6). d Tertiary dendrites were counted by imaging regions of interest in dorsal dentate gyrus and analyzing using filament feature in Imaris. Loss of α7 nAChRs does not impact on tertiary dendrite numbers. d–f In order to examine the effect of loss of α7 nAChRs on dendritic structure, images were captured and analyzed using Imaris Filament function. These images delineate the process. d A region of interest was randomly selected from the upper arm of the dentate gyrus in the dorsal region of the hippocampus and imaged. e Four DCX cells were randomly selected from all cells present in the image. Cells were manually traced using the filament function on Imaris and checked for accuracy in 3D. f Tracings were used to count tertiary filaments and measure overall filament length. Female mice were not analyzed. Asterisk indicates *p < 0.05. Statistical analysis with unpaired Student’s t test calculated using Graphpad Prism 7.0. Data in bar graph expressed as mean ± SEM

Article Snippet: Furthermore, prenatal stress decreases adult neurogenesis ( Belnoue et al. 2013 ), and also affects the levels of α7 nAChRs in adult rodents ( Baier et al. 2015 ).

Techniques: Imaging

Journal: Brain structure & function

Article Title: The α7 nicotinic acetylcholine receptors regulate hippocampal adult-neurogenesis in a sexually dimorphic fashion

doi: 10.1007/s00429-018-1799-6

Figure Lengend Snippet: Effects of α7 nAChRs by sex and genotype. Schematic shows a summary of the effects of loss of α7 nAChRs by sex and genotype. Overall, cell divisions increased in α7 nAChR knock-out mice (shown by a larger arrow), with an increase in intermediate progenitor cells. In adult male mice only, loss of α7 nAChRs also led to a decrease in NSCs and an increase in immature neurons, supporting the idea that adult neurogenesis was increased in these mice after the loss of the α7 nAChR. Despite their apparent higher levels, we would anticipate poor survival for these immature neurons, and a loss of neurons at maturity. Future studies will test survival. In addition, the loss of performance in the DNTP radial-arm maze task is consistent with reduced functionality of the dentate gyrus, supporting the hypothesis that the observed neurogenesis is dysregulated and aberrant, rather than healthy. Wild-type adult female mice demonstrate a lack of preference for novelty in the DNTP radial-arm maze task, with no significant change in mice lacking the α7 nAChR. In addition, in adult female mice levels of NSCs and immature neurons remain unchanged, indicating that increased cell division is not having an easily identifiable lasting impact on adult neurogenesis

Article Snippet: Furthermore, prenatal stress decreases adult neurogenesis ( Belnoue et al. 2013 ), and also affects the levels of α7 nAChRs in adult rodents ( Baier et al. 2015 ).

Techniques: Knock-Out

Journal: JHEP Reports

Article Title: Natural antibodies are required for clearance of necrotic cells and recovery from acute liver injury

doi: 10.1016/j.jhepr.2024.101013

Figure Lengend Snippet: Rag2 -/- mice have impaired cell debris clearance, prolonged inflammation and delayed recovery from liver injury. (A) Representative intravital microscopy images of WT and Rag2 -/- mice showing the deposition of DNA (sytox green + ) in the liver 24 and 48 h after APAP overdose (600 mg/kg). Scale bar = 100 μm. (B), Quantification of the sytox green + area in the liver. (C) Quantification of the fibrin(ogen) + area fraction in liver cryosections. See Fig. S3A for IgM and IgG stainings. (D) Serum ALT levels in WT and Rag2 -/- mice 24 and 48 h after APAP overdose. Data are from WT and Rag2 -/- mice 24 and 48 h after an oral gavage of 600 mg/kg APAP (A-D) and are represented as mean ± SEM. Each dot in the graph represents a single mouse (B-D). Image quantifications were pooled from 10 random pictures per liver (B–C). ∗ p ≤0.05 (Student’s t test). ALT, alanine aminotransferase; APAP, acetaminophen; Rag2 , recombination activating gene 2; WT, wild-type.

Article Snippet: C57BL/6J and C57BL/6NRj mice were purchased from Janvier Labs. C57BL/6N-Rag2Tm1/CipheRj ( Rag2 -/- ) were bred in specific pathogen-free conditions at the Animal Facility of the Rega Institute (KU Leuven).

Techniques: Intravital Microscopy

Journal: JHEP Reports

Article Title: Natural antibodies are required for clearance of necrotic cells and recovery from acute liver injury

doi: 10.1016/j.jhepr.2024.101013

Figure Lengend Snippet: Restitution of NAbs to Rag2 -/- mice improves the recovery from liver injury. (A) Representative images showing immunostaining of liver cryosections from APAP-challenged Rag2 -/- mice treated with 150 μl of WT or Rag2 -/- serum. Gray: fibrin(ogen), red: IgM, green: IgG. Scale bar = 100 μm. (B) Quantification of the fibrin(ogen) + area fraction in liver cryosections. (C) Serum ALT levels at 24 and 48 h after APAP administration. (D) Quantification of Ki67 + hepatocytes in liver cryosections of Rag2 -/- mice after serum transfer. (E) Quantification of Ki67 + hepatocytes in liver cryosections of Rag2 -/- mice treated with 100 μg (i.v.) of purified IgM and IgG. See also for Ki67 staining. Data are from Rag2 -/- mice that received an intravenous injection of WT serum or Rag2 -/- serum (A-D) or 100 μg purified IgG and IgM (E) 4 h after an oral gavage with 600 mg/kg APAP and were sacrificed 24 and 48 h after the APAP challenge. Each dot in the graph represents a single mouse (B-E). Image quantifications were pooled from 10 random pictures per liver (B, D and E). ∗ p ≤0.05 (Student’s t test). ALT, alanine aminotransferase; APAP, acetaminophen; NAbs, natural antibodies; Rag2 , recombination activating gene 2; WT, wild-type.

Article Snippet: C57BL/6J and C57BL/6NRj mice were purchased from Janvier Labs. C57BL/6N-Rag2Tm1/CipheRj ( Rag2 -/- ) were bred in specific pathogen-free conditions at the Animal Facility of the Rega Institute (KU Leuven).

Techniques: Immunostaining, Purification, Staining, Injection

Journal: JHEP Reports

Article Title: Natural antibodies are required for clearance of necrotic cells and recovery from acute liver injury

doi: 10.1016/j.jhepr.2024.101013

Figure Lengend Snippet: Natural antibodies drive the phagocytosis of necrotic cell debris through FcγRs and CD11b. (A) Phagocytosis by primary mouse neutrophils 3 h after adding debris opsonized with normal or HI serum. FcγRs were blocked by adding 100 μg/ml IgG to the cells. CD11b was blocked with 10 μg/ml anti-mouse CD11b. Phagocytosis was blocked with 10 μM Latrunculin B. (B,C) Phagocytosis by primary mouse neutrophils 3 h after adding IgG- or IgM-opsonized debris. Receptors were blocked using 10 μg/ml anti-CD16.2, anti-CD16/CD32, anti-CD11b or 100 μg/ml purified mouse IgG. (D) Phagocytosis by primary human neutrophils 3 h after adding debris opsonized with normal or HI serum. FcγRs were blocked with 100 μg/ml IgG and Latrunculin B was used at 10 μM. (E) Representative confocal images from human neutrophils containing pHrodo + phagosomes. FcγRs were blocked with 100 μg/ml purified human IgG. Scale bar = 10 μm. See also supplementary video 6. (F) Representative image from immunostaining of liver cryosections from WT and Rag2 -/- mice 12 h after FTI showing deposition of IgM (red) and IgG (cyan). Scale bar = 200 μm. (G) Representative intravital microscopy images showing neutrophils (Ly6G + ) phagocytosing necrotic debris as in H. Scale bar = 20 μm. Yellow square represents the area zoomed. Also see supplementary videos 3-5. (H) Quantification of the percentage of neutrophils (Ly6G + , green) phagocytosing necrotic debris 6 h after a FTI in the liver of WT, Rag2 -/- mice. A separate group of Rag2 -/- mice were restituted with total purified Abs (100 μg/mouse) (n ≥4). The FTI was labeled with a droplet of 4 μM pHrodo Red succinimidyl ester. Each dot in the graph represents a mouse. (I) Quantification of Ki67 + cells in liver cryosections of WT, Rag2 -/- and Rag2 -/- mice treated with purified IgM and IgG (100 μg/mouse). Mice were treated with purified antibodies 30 min prior to the FTI (n ≥4). Data are represented as mean ± SEM. ∗ p ≤0.05 (one-way ANOVA for panels A, B, D, H and I); (Student’s t test for panel C). Abs, antibodies; FTI, focal thermal injury; HI, heat-inactivated; Rag2 , recombination activating gene 2; WT, wild-type.

Article Snippet: C57BL/6J and C57BL/6NRj mice were purchased from Janvier Labs. C57BL/6N-Rag2Tm1/CipheRj ( Rag2 -/- ) were bred in specific pathogen-free conditions at the Animal Facility of the Rega Institute (KU Leuven).

Techniques: Purification, Immunostaining, Intravital Microscopy, Labeling

Journal: IBRO Neuroscience Reports

Article Title: Electroacupuncture alleviates paradoxical sleep deprivation-induced postoperative hyperalgesia via a7nAChR mediated BDNF/TrkB-KCC2 signaling pathway in the spinal cord

doi: 10.1016/j.ibneur.2024.10.002

Figure Lengend Snippet: The protein expression of α7nAChR (n=10 in each group). *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001.

Article Snippet: The filter membranes were blocked with 5 % nonfat milk for 1 h at room temperature and incubated with the primary antibody α7nAChR (1:1000, Genetex, USA), anti-BDNF (1:1000, Genetex, USA), anti-trkB (1:1000, Genetex, USA) and anti-KCC (1:1000, Genetex, USA) for overnight at 4°C.

Techniques: Expressing

Journal: IBRO Neuroscience Reports

Article Title: Electroacupuncture alleviates paradoxical sleep deprivation-induced postoperative hyperalgesia via a7nAChR mediated BDNF/TrkB-KCC2 signaling pathway in the spinal cord

doi: 10.1016/j.ibneur.2024.10.002

Figure Lengend Snippet: Schematic diagram of the fluorescence of α7nAChR in the spinal cord (n=10 in each group). Scale bars are 50 μm.

Article Snippet: The filter membranes were blocked with 5 % nonfat milk for 1 h at room temperature and incubated with the primary antibody α7nAChR (1:1000, Genetex, USA), anti-BDNF (1:1000, Genetex, USA), anti-trkB (1:1000, Genetex, USA) and anti-KCC (1:1000, Genetex, USA) for overnight at 4°C.

Techniques: Fluorescence

Journal: PLoS ONE

Article Title: Vimentin, a Novel NF-κB Regulator, Is Required for Meningitic Escherichia coli K1-Induced Pathogen Invasion and PMN Transmigration across the Blood-Brain Barrier

doi: 10.1371/journal.pone.0162641

Figure Lengend Snippet: A-B: Immunehistochemical analysis of vimentin (A) and α7 nAChR (B) expression in mouse brain cortex. Images (A and B) photographed at 200X magnification C: Western blotting analysis of vimentin, α7 nAChR, phospho-IKKα/β and P65 expression levels in mouse vascular endothelial cells.

Article Snippet: All mouse monoclonal (MMAb)/ rabbit polyclonal (RPAb) antibodies and other reagents were commercially obtained from the following sources: (A) MMAbs against vimentin (V9) (sc-6260), NF-kB (p65) (sc-109) and a RPAb against β-actin (sc-130657) from Santa Cruz Biotechnology (Santa Cruz, CA); (B) RPAbs against phospho-IKK α/β (Ser176/180) (#2697), phospho-ERK1/2 (Thr202/Tyr204) (#4370), and a MMAb against IkBa (#4814) from Cell Signaling Technology (Danvers, MA); (C) a RPAb recognizing phoshothreonine286 of CaM kinase II from Promega and a MMAb recognizing CaM kinase II from BD Biosciences; (D) a RPAb against α7 nAChR from Genescript (Piscataway, NJ) and a rat anti-mouse Ly-6G (Gr-1) antibody from eBiosciences (San Diego, CA).

Techniques: Expressing, Western Blot

Journal: PLoS ONE

Article Title: Vimentin, a Novel NF-κB Regulator, Is Required for Meningitic Escherichia coli K1-Induced Pathogen Invasion and PMN Transmigration across the Blood-Brain Barrier

doi: 10.1371/journal.pone.0162641

Figure Lengend Snippet: Elevation of intracellular calcium flux in HBMEC stimulated with E44 or ZD1 strains. HBMEC transfected with siRNAs of a scrambled sequence (CON), vimentin (Vim KD) and α7 nAChR (A7 KD) were loaded with Fura-2 AM as described in Methods and Materials. The monolayer was monitored for intracellular calcium flux for 10 minutes with 4 s intervals under an automated fluorescent microscope. Monolayer cells were stimulated with E44 or ZD (10 8 CFU) at the 120 s time point. The intensity of fluorescence at 340 nm and 380 nm was measured. The ratios of intensity of fluorescence at 340 nm and 380 nm were calculated for each time interval and depicted as continuous lines in (A–C). (G) The y axis represents the ratio, and x axis represents time (s). The 340 nm/380 nm ratio changes in each treatment were calculated and subjected to statistical analysis. WT HBMEC without any pre-treatment served as a control and are defined as one-fold (1.0). (*P<0.05; **P<0.01).

Article Snippet: All mouse monoclonal (MMAb)/ rabbit polyclonal (RPAb) antibodies and other reagents were commercially obtained from the following sources: (A) MMAbs against vimentin (V9) (sc-6260), NF-kB (p65) (sc-109) and a RPAb against β-actin (sc-130657) from Santa Cruz Biotechnology (Santa Cruz, CA); (B) RPAbs against phospho-IKK α/β (Ser176/180) (#2697), phospho-ERK1/2 (Thr202/Tyr204) (#4370), and a MMAb against IkBa (#4814) from Cell Signaling Technology (Danvers, MA); (C) a RPAb recognizing phoshothreonine286 of CaM kinase II from Promega and a MMAb recognizing CaM kinase II from BD Biosciences; (D) a RPAb against α7 nAChR from Genescript (Piscataway, NJ) and a rat anti-mouse Ly-6G (Gr-1) antibody from eBiosciences (San Diego, CA).

Techniques: Transfection, Sequencing, Microscopy, Fluorescence, Control

Journal: PLoS ONE

Article Title: Vimentin, a Novel NF-κB Regulator, Is Required for Meningitic Escherichia coli K1-Induced Pathogen Invasion and PMN Transmigration across the Blood-Brain Barrier

doi: 10.1371/journal.pone.0162641

Figure Lengend Snippet: E . coli K1 infection triggers IbeA-dependent activation of the vimentin signaling pathway at the host cell membrane. IbeA binds to its receptor Vim and co-receptor PSF that interact with α7n nAChR through lipid rafts. These communications trigger phosphorylation of signaling proteins (e.g., Vim, TAK1), which in turn activates the nuclear factor-kappaB (NF-κB) pathways via activation of the IκB kinase (IKK) complex. NF-κB activation resulted to the nuclear translocation of NF-κB, which induces the production of cytokines, chemokines, and others proinflammatory molecules in response to bacterial stimuli.

Article Snippet: All mouse monoclonal (MMAb)/ rabbit polyclonal (RPAb) antibodies and other reagents were commercially obtained from the following sources: (A) MMAbs against vimentin (V9) (sc-6260), NF-kB (p65) (sc-109) and a RPAb against β-actin (sc-130657) from Santa Cruz Biotechnology (Santa Cruz, CA); (B) RPAbs against phospho-IKK α/β (Ser176/180) (#2697), phospho-ERK1/2 (Thr202/Tyr204) (#4370), and a MMAb against IkBa (#4814) from Cell Signaling Technology (Danvers, MA); (C) a RPAb recognizing phoshothreonine286 of CaM kinase II from Promega and a MMAb recognizing CaM kinase II from BD Biosciences; (D) a RPAb against α7 nAChR from Genescript (Piscataway, NJ) and a rat anti-mouse Ly-6G (Gr-1) antibody from eBiosciences (San Diego, CA).

Techniques: Infection, Activation Assay, Membrane, Phospho-proteomics, Translocation Assay

Journal: Advanced Pharmaceutical Bulletin

Article Title: Alpha7 Nicotinic Acetylcholine Receptor Mediates Nicotine-induced Apoptosis and Cell Cycle Arrest of Hepatocellular Carcinoma HepG2 Cells

doi: 10.15171/apb.2020.008

Figure Lengend Snippet: The sequence of primers for alpha7 nicotinic acetylcholine receptor (α7nAChR), caspase-3, cyclin B1, and GAPDH genes

Article Snippet: The primers sequences for α7nAChR and other genes were obtained from Sinaclon (Tehran, Iran) and listed in .

Techniques: Sequencing

Journal: Scientific Reports

Article Title: Alpha7 nicotinic acetylcholine receptor is required for amyloid pathology in brain endothelial cells induced by Glycoprotein 120, methamphetamine and nicotine

doi: 10.1038/srep40467

Figure Lengend Snippet: HBMECs were incubated with gp120 (50 ng/ml) ( a ) or METH (50 nM) ( b ) for 2, 6, and 24 h, respectively. Expression of α7 nAChR (AchR7), S100B, Tau, and RAGE, the biomarkers of Alzheimer’s-like brain pathology, was determined by Western blot using the antibodies as described in Materials and methods. 0 h: the control HBMECs without METH or gp120 stimulation. β-actin in both fractions was detected as internal loading controls.

Article Snippet: All mouse monoclonal (MMAb)/rabbit polyclonal (RPAb)/goat polyclonal (PGAb) antibodies used for immunostaining, ELISA and Western blot analyses were as follows: (A) PRAb against α7 nAChR (1:60 for immunostaining, 1:500 for Western blot, Genescript, Piscataway, NJ); (B) PRAb against S100B (1:60 for immunostaining, 1:500 for Western blot, LifeSpan BioScience); (C) Rhodamine green-labeled MMAb recognizing β-Amyloid(1–40), MMAb against β-Amyloid(1–42)(1:1000 for ELISA) and PRAb recognizing Tau (Pser199/202) (1: 1000 for ELISA, 1:500 for Western blot) from Ana Spec Inc (Fremont, CA); (D) MMAb against Aβ (17–24) (1:1000 for Western blot, Covance, Emeryville, CA); (E) PRAb against UCHL1(1: 1000 for ELISA, Protein Tech); and (F) PGAbs against RAGE (sc-8230)) (1:500 for Western blot) and PRAb against β-actin (sc-7210) (1:2000 for Western blot) from Santa Cruz Biotechnology (CA).

Techniques: Incubation, Expressing, Western Blot, Control

Journal: Scientific Reports

Article Title: Alpha7 nicotinic acetylcholine receptor is required for amyloid pathology in brain endothelial cells induced by Glycoprotein 120, methamphetamine and nicotine

doi: 10.1038/srep40467

Figure Lengend Snippet: Immunofluorescence microscopy showed the expression of α7 nAChR (AchR7) in DG of the hippocampus of C57BL/6J mouse brains upon treatment with HIV-1 gp120 (50 ng/mouse, 2d), METH (2, 4, 6, 8, 10, 10, 10, 10, 10, 10 mg/kg/d), and NT (1.5 mg/kg/d, 3d). DAPI was used to stain the nuclei in the brain in the merged pictures. ( CON ) mice without any treatment. (gp120) mice injected with gp120. (METH) mice injected with METH. (NT) mice treated with NT. Images are 200×. Scale bar = 100 μm.

Article Snippet: All mouse monoclonal (MMAb)/rabbit polyclonal (RPAb)/goat polyclonal (PGAb) antibodies used for immunostaining, ELISA and Western blot analyses were as follows: (A) PRAb against α7 nAChR (1:60 for immunostaining, 1:500 for Western blot, Genescript, Piscataway, NJ); (B) PRAb against S100B (1:60 for immunostaining, 1:500 for Western blot, LifeSpan BioScience); (C) Rhodamine green-labeled MMAb recognizing β-Amyloid(1–40), MMAb against β-Amyloid(1–42)(1:1000 for ELISA) and PRAb recognizing Tau (Pser199/202) (1: 1000 for ELISA, 1:500 for Western blot) from Ana Spec Inc (Fremont, CA); (D) MMAb against Aβ (17–24) (1:1000 for Western blot, Covance, Emeryville, CA); (E) PRAb against UCHL1(1: 1000 for ELISA, Protein Tech); and (F) PGAbs against RAGE (sc-8230)) (1:500 for Western blot) and PRAb against β-actin (sc-7210) (1:2000 for Western blot) from Santa Cruz Biotechnology (CA).

Techniques: Immunofluorescence, Microscopy, Expressing, Staining, Injection