Journal: PLoS ONE

Article Title: Fine-Tuning Roles of Endogenous Brain-Derived Neurotrophic Factor, TrkB and Sortilin in Colorectal Cancer Cell Survival

doi: 10.1371/journal.pone.0025097

Figure Lengend Snippet: (A): RT-PCR of BDNF, its high (TrkB) and low (p75 NTR ) affinity receptors, TrkA, and TrkC from cells cultured in basal (10% FCS medium), WiDr (lane: 1), SW480 (lane: 2), SW620 (lane: 3), COLO 205 (lane: 4). GAPDH mRNA levels was used as an internal control. Positive control (lane 5) was the neuroblastoma cell line (IMR32) for BDNF, TrkB and p75 NTR ; and the erythromyeloblastoid leukemia cells (K562) for TrkA and C. A representative result of at least three to five independent experiments. (B) Comparison of TrkB95 and p75 NTR on total RNA extracted from WiDr cells cultured in 10% FCS or after 24 to 72 h serum deprivation (0% FCS). TrkB95 and p75 NTR mRNA/GAPDH mRNA quantification of band intensities evaluated by densitometry are shown above lanes and expressed in arbitrary units (mean of three independent experiments). (C) Same experiment: RT-PCR of TrkA and TrkC on total RNA extracted from WiDr cells cultured under basal culture conditions (10% FCS) and after 24–72 h of serum starvation in comparison to positive control (K562) (D) Expression of pro-BDNF and BDNF, full length TrkB 145 and truncated TrkB 95 and p75 NTR proteins in CRC cell lines cultured in 10% FCS. TrkA and TrkC were not detected. Actin was used as loading protein control. WiDr (lane: 1), SW480 (lane: 2), SW620 (lane: 3), COLO 205 (lane: 4). Positive control (lane: 5) were IMR32 cells for BDNF, pro-BDNF, TrkB and p75 NTR and K562 cells or TrkA and TrkC. A representative result of at least three independent experiments.

Article Snippet: The following Abs were used: rabbit anti-BDNF Ab (1 µg/ml; Santa Cruz Biotechnology), rabbit anti-pro-BDNF Ab (8 µg/ml; Alomone Labs), mouse anti-TrkB Ab (2.5 µg/ml; R&D Systems), rabbit anti-p75 NTR (2 µg/ml; Santa Cruz Biotechnology) and goat anti-sortilin Ab (1 µg/ml; Santa Cruz Biotechnology).

Techniques: Reverse Transcription Polymerase Chain Reaction, Cell Culture, Positive Control, Expressing

Journal: PLoS ONE

Article Title: Fine-Tuning Roles of Endogenous Brain-Derived Neurotrophic Factor, TrkB and Sortilin in Colorectal Cancer Cell Survival

doi: 10.1371/journal.pone.0025097

Figure Lengend Snippet: BDNF release and effect of exogenous BDNF, pro-BDNF, TrkB inhibitor (K252a) and neutralizing anti-BDNF on apoptosis and proliferation in primary CRC cell lines.

Article Snippet: The following Abs were used: rabbit anti-BDNF Ab (1 µg/ml; Santa Cruz Biotechnology), rabbit anti-pro-BDNF Ab (8 µg/ml; Alomone Labs), mouse anti-TrkB Ab (2.5 µg/ml; R&D Systems), rabbit anti-p75 NTR (2 µg/ml; Santa Cruz Biotechnology) and goat anti-sortilin Ab (1 µg/ml; Santa Cruz Biotechnology).

Techniques:

Journal: PLoS ONE

Article Title: Fine-Tuning Roles of Endogenous Brain-Derived Neurotrophic Factor, TrkB and Sortilin in Colorectal Cancer Cell Survival

doi: 10.1371/journal.pone.0025097

Figure Lengend Snippet: BDNF release and effect of exogenous BDNF, pro-BDNF, TrkB inhibitor (K252a) and neutralizing anti-BDNF on apoptosis and proliferation in metastatic CRC cell lines.

Article Snippet: The following Abs were used: rabbit anti-BDNF Ab (1 µg/ml; Santa Cruz Biotechnology), rabbit anti-pro-BDNF Ab (8 µg/ml; Alomone Labs), mouse anti-TrkB Ab (2.5 µg/ml; R&D Systems), rabbit anti-p75 NTR (2 µg/ml; Santa Cruz Biotechnology) and goat anti-sortilin Ab (1 µg/ml; Santa Cruz Biotechnology).

Techniques:

Journal: PLoS ONE

Article Title: Fine-Tuning Roles of Endogenous Brain-Derived Neurotrophic Factor, TrkB and Sortilin in Colorectal Cancer Cell Survival

doi: 10.1371/journal.pone.0025097

Figure Lengend Snippet: Confocal microscopy of WiDr (A, B) and COLO 205 (C, D) cells, stained with an anti-BDNF Ab (red), anti-TrkB mAb (green) or both (overlay) cultured with 10% FCS (A, C) or after 24-h serum deprivation (B, D). Under basal culture conditions (10% FCS), TrkB and BDNF were sequestered in the cytoplasm (arrows) in WiDr (A) and COLO 205 (C) cells. The same staining patterns were obtained with the two other cell lines (data not shown). After serum starvation relocation to the cell membrane and colocalization of TrkB and BDNF (yellow in merged, arrows) were detected in WiDr (B) and COLO 205 (D). Images were representative for at least three to five independent experiments.

Article Snippet: The following Abs were used: rabbit anti-BDNF Ab (1 µg/ml; Santa Cruz Biotechnology), rabbit anti-pro-BDNF Ab (8 µg/ml; Alomone Labs), mouse anti-TrkB Ab (2.5 µg/ml; R&D Systems), rabbit anti-p75 NTR (2 µg/ml; Santa Cruz Biotechnology) and goat anti-sortilin Ab (1 µg/ml; Santa Cruz Biotechnology).

Techniques: Confocal Microscopy, Staining, Cell Culture

Journal: PLoS ONE

Article Title: Fine-Tuning Roles of Endogenous Brain-Derived Neurotrophic Factor, TrkB and Sortilin in Colorectal Cancer Cell Survival

doi: 10.1371/journal.pone.0025097

Figure Lengend Snippet: (A) Role of endogenous BDNF and its receptor TrkB on CRC cell proliferation: effects of exogenous BDNF and suppressing endogenous TrkB receptor on cell proliferation. The four cell lines were cultured for 24 h in FCS-free medium (FCS 10%, −) in the presence of exogenous BDNF (+), K252a (+) alone or in combination. Cell proliferation was determined by flow cytometry analysis using EdU Alexa Fluor 488. The data are presented as histograms of proliferating cells in relative units ± SEM of five independent experiments. *, p <0.05; **, p <0.01; ***, p <0.001, when compared with serum-free medium. (B, C) Effects of exogenous BDNF and suppressing endogenous TrkB receptor on cell survival. Apoptotic ratios of soluble nucleosomes were detected by ELISA Cell for WiDr, SW480, SW620, and COLO 205 induced by serum deprivation alone (FCS 10%, −) or in association either with exogenous BDNF (+), or with K252a (+), during 24–72 h of serum deprivation. Histograms, mean ratio of apoptotic cells ± SEM of at least three independent experiments. *, p <0.05; **, p <0.01; ***, p <0.001, when compared with serum-free condition alone. (D, E) apoptotic ratio after 24 h serum deprivation alone (0% FCS) or with combination with a neutralizing anti-BDNF mAb (0% anti-BDNF). Histograms, mean ratio of apoptotic cells ± SEM of three independent experiments. *, p <0.05; **, p <0.01; ***, p <0.001, when compared with serum-free condition alone.

Article Snippet: The following Abs were used: rabbit anti-BDNF Ab (1 µg/ml; Santa Cruz Biotechnology), rabbit anti-pro-BDNF Ab (8 µg/ml; Alomone Labs), mouse anti-TrkB Ab (2.5 µg/ml; R&D Systems), rabbit anti-p75 NTR (2 µg/ml; Santa Cruz Biotechnology) and goat anti-sortilin Ab (1 µg/ml; Santa Cruz Biotechnology).

Techniques: Cell Culture, Flow Cytometry, Enzyme-linked Immunosorbent Assay

Journal: Theranostics

Article Title: A Novel Peptide Interfering with proBDNF-Sortilin Interaction Alleviates Chronic Inflammatory Pain

doi: 10.7150/thno.29703

Figure Lengend Snippet: Mapping of proBDNF-sortilin interaction interface. (A) Schematic diagram illustrating the design of human BDNF prodomain variants. Six variants with a series of amino acids deletions were used in this study. V66M, Val66Met; S, Signal peptide (amino acid 1-18); BDNF, mature BDNF (amino acid 129-247); HA, C-terminal HA epitope. (B) Immunoblots of co-immunoprecipitation (Left) or inputs (Right) of sortilin and proBDNF variants obtained with antibody against HA or Myc epitope. IP, Immunoprecipitation; IB, Immunoblot. (C) Left , live fluorescence micrograph of BiFC mapping assay (Yellow) using HEK293T cells with VC155-sortilin (Sort1) and VN155(I152L)-BDNF prodomain variants. Scale bar represents 50 µm. Right , quantification of fluorescence intensity of BiFC assay. P <0.0001, one-way ANOVA. ** P <0.01 versus Fl group; *** P <0.0001 versus Fl group; ## P <0.001 versus variant I; ### P <0.0001 versus variant I group; n.s., non-significant. Values are means ± SEM. n = 100 individual cells from 4 images. (D) Left , representative immunoblots of supernatant and cell lysates of HEK293 cells overexpressing sortilin and BDNF prodomain variants obtained with antibodies against HA. Right , densitometry analyses of secreted BDNF in supernatants, normalized to input in lysates. P <0.001, one-way ANOVA. * P <0.05, versus Fl group; *** P <0.001, versus Fl group; ## P <0.05, versus I group. Values are means ± SEM. n = 4. IB, Immunoblot. (E) BDNF ELISA quantification of supernatant collected from HEK293 cells overexpressing sortilin and BDNF prodomain variants. P <0.0001, one-way ANOVA. * P <0.05; ** P <0.01; *** P <0.001, versus Sort1/Fl group; ## P <0.05, versus Sort1/variant I group. Values are means ± SEM. Each data point represents the average of 3 independent experiments.

Article Snippet: The primary antibodies used included anti-proBDNF antibody (#ANT-006-AG, Alomone Labs Ltd, Jerusalem, Israel), anti-phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) antibody (Cell signalling Technology, Danvers, MA, USA), anti-NeuN antibody (MAB377, EMD Millipore Corporation, Billerica, MA, USA) and anti-sortilin antibody (Abcam, Cambridge, UK).

Techniques: Western Blot, Immunoprecipitation, Fluorescence, Mapping Assay, Bimolecular Fluorescence Complementation Assay, Variant Assay, Enzyme-linked Immunosorbent Assay

Journal: Theranostics

Article Title: A Novel Peptide Interfering with proBDNF-Sortilin Interaction Alleviates Chronic Inflammatory Pain

doi: 10.7150/thno.29703

Figure Lengend Snippet: Construction of a novel peptide targeting proBDNF-sortilin Interaction. (A) Schematic diagram illustrating the design of a series of blocking peptides targeting proBDNF-sortilin interaction interface. Scr-bdnf84-83 (scr), scrambled peptide with permutation of peptide sequence at amino acid 84-93. (B) Immunoblots of co-immunoprecipitation (upper) or reverse co-immunoprecipitation (lower) of sortilin and proBDNF variants in cell lysates overexpressing full-length proBDNF and sortilin followed by peptide incubation obtained with antibodies against HA or Myc. IP, Immunoprecipitation; IB, Immunoblot. (C) Schematic diagram of the design of the Tat-tagged interfering peptide. Scr-bdnf89-98-Tat, Tat-tagged scrambled peptide with permutation of peptide sequence at amino acid 89-98. (D) Live fluorescence micrographs of 5FAM (green) or bright field in rat DRG neuronal cultures before (0 min) and incubated with 5 µM 5FAM-tagged bdnf89-98-Tat (bdnf-Tat) for 2 h. Scale bar represents 150 µm. (E) Fluorescence micrograph of 5FAM (green), sortilin (red) or DAPI (blue) in rat DRG neuronal cultures with peptides pre-treatment for 30 min. Scale bars represent 25 µm. (F) Left , fluorescence micrograph of BiFC assay (yellow) of HEK293T cells overexpressing full-length BDNF and sortilin with vehicle, 10 µM scr-Tat or bdnf-Tat pre-treatment for 30 min. Scale bars represent 25 µm. Right , Determination of the corrected total cell fluorescence (CTCF) of the BiFC signal. P <0.0001 , one-way ANOVA. *** P <0.0001, scr-Tat treatment versus bdnf-Tat treatment. Mean values ± SEM. n = 100 individual cells from 5 images. (G) Effect of 30 min peptide pre-treatment on activity-dependent secretion of BDNF in rat DRG neuronal cultures with high-potassium stimulus using ELISA. P <0.05, one-way ANOVA. * P <0.05, vehicle versus scr-Tat treatment or scr-Tat treatment versus bdnf-Tat treatment. Values are means ± SEM. Each data point represents the average of 3 independent experiments.

Article Snippet: The primary antibodies used included anti-proBDNF antibody (#ANT-006-AG, Alomone Labs Ltd, Jerusalem, Israel), anti-phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) antibody (Cell signalling Technology, Danvers, MA, USA), anti-NeuN antibody (MAB377, EMD Millipore Corporation, Billerica, MA, USA) and anti-sortilin antibody (Abcam, Cambridge, UK).

Techniques: Blocking Assay, Sequencing, Western Blot, Immunoprecipitation, Incubation, Fluorescence, Bimolecular Fluorescence Complementation Assay, Activity Assay, Enzyme-linked Immunosorbent Assay

Journal: Mediators of Inflammation

Article Title: Dexmedetomidine Attenuates Neurotoxicity in Developing Rats Induced by Sevoflurane through Upregulating BDNF-TrkB-CREB and Downregulating ProBDNF-P75NRT-RhoA Signaling Pathway

doi: 10.1155/2020/5458061

Figure Lengend Snippet: Dexmedetomidine could decrease the level of proBDNF and restore the ratio of proBDNF/mBDNF and alleviates activation of the proBDNF-P75NRT-RHOA pathway after sevoflurane. (a) Western blot band. (b) ProBDNF/mBDNF. (c–e) Bar graph of Western blot. (f) Immunofluorescence of proBDNF (scale bar = 50 μ m). ∗ Compare with the control group, P < 0.05; # Compared with the Sev group, P < 0.05.

Article Snippet: The primary antibodies were added: anti-BDNF (1 : 2000, Abcam, Cambridge, UK), anti-proBDNF (1 : 500, Alomone Labs, Israel), P75NRT (1 : 2000, Cell Signaling Technology, USA), TrkB (1 : 1000, ABclonal, Woburn, MA, USA), p-TrkB (1 : 1000, ABclonal, Woburn, MA, USA), CREB (1 : 1000, Cell Signaling Technology, USA), anti-RhoA (1 : 500, Proteintech, China), PSD95 (1 : 500, Proteintech Biotechnology, Chicago, IL, USA), SYP (1 : 1000, Abcam, USA), anti- β -actin (1 : 500, Proteintech, China), and GAPDH (1 : 1000, Cell Signaling Technology, Boston, MA, USA).

Techniques: Activation Assay, Western Blot, Immunofluorescence

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Pro-BDNF Contributes to Hypoxia/Reoxygenation Injury in Myocardial Microvascular Endothelial Cells: Roles of Receptors p75 NTR and Sortilin and Activation of JNK and Caspase 3

doi: 10.1155/2018/3091424

Figure Lengend Snippet: Effects of the anti-pro-BDNF antibody on apoptosis, migration, and capillary-like-structure formation among MMECs after exposure to HG and H/R. (a) Effects of pro-BDNF on apoptosis were analyzed by flow cytometry of MMECs after different treatments: control, H/R, H/R + anti-pro-BDNF, and H/R + vehicle. (b, c) The functional effects of pro-BDNF on MMECs were assessed by capillary-like-structure formation and cell scratch assays. (d) Relative apoptosis levels and fold changes are expressed in relation to the control group. The H/R group showed markedly increased relative apoptosis levels, decreased capillary-like-structure formation, and reduced cell migration when compared with the control group. These effects were reversed by treatment with the anti-pro-BDNF antibody to the levels similar to those in the control group. The data were subjected to one-way ANOVA. The error bars represent SEM. ∗ P < 0.05 as compared with the control group; # P < 0.05 as compared with the H/R group or the H/R + vehicle group.

Article Snippet: After blocking in 5% fat-free dry milk, antibodies against pro-BDNF (Alomone, 1 : 400), BDNF (Abcam, 1 : 500), P75 (Santa Cruz Biotechnology, Santa Cruz, CA, USA; 1 : 100), sortilin (Abcam, 1 : 500), JNK (Santa Cruz Biotechnology, 1 : 100), cleaved caspase 3 (Asp175, Cell Signaling Technology, 1 : 1000), caspase 3, and β -actin (Santa Cruz Biotechnology) were employed.

Techniques: Migration, Flow Cytometry, Functional Assay

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Pro-BDNF Contributes to Hypoxia/Reoxygenation Injury in Myocardial Microvascular Endothelial Cells: Roles of Receptors p75 NTR and Sortilin and Activation of JNK and Caspase 3

doi: 10.1155/2018/3091424

Figure Lengend Snippet: Effects of the anti-pro-BDNF antibody on the expression of p75 NTR and sortilin and apoptosis-related proteins. (a) Representative immunofluorescent images of pro-BDNF (red, first column), p-JNK (green, second column), BDNF (red, fourth column), JNK (green, fifth column), sortilin (red, seventh column), and p75 NTR (green, eighth column) in groups control, H/R, H/R + anti-pro-BDNF, and H/R + vehicle. (b–e) Representative Western blots and quantitative analysis of pro-BDNF, BDNF (b), p75 NTR , sortilin (c), JNK, p-JNK (d), caspase 3 and cleaved-caspase 3 expression (e) in response to different treatments. All the data were normalized to β -actin, and fold changes are expressed in relation to the control group. Exposure of MMECs to H/R resulted in significantly higher expression levels of pro-BDNF, p75 NTR , and sortilin and in activation of JNK and caspase 3 as compared with MMECs maintained under normal conditions (control). Nonetheless, there were no significant differences in BDNF, JNK, and caspase 3 expression levels after H/R. Treatment with the anti-pro-BDNF antibody significantly reversed the increase in the protein expression of pro-BDNF, p75 NTR , sortilin, p-JNK, and cleaved caspase 3 in MMECs after exposure to HG and H/R (H/R + anti-pro-BDNF). The data were subjected to one-way ANOVA. The error bars represent SEM. ∗ P < 0.05 as compared with the control group; # P < 0.05 as compared with the H/R or H/R + vehicle group.

Article Snippet: After blocking in 5% fat-free dry milk, antibodies against pro-BDNF (Alomone, 1 : 400), BDNF (Abcam, 1 : 500), P75 (Santa Cruz Biotechnology, Santa Cruz, CA, USA; 1 : 100), sortilin (Abcam, 1 : 500), JNK (Santa Cruz Biotechnology, 1 : 100), cleaved caspase 3 (Asp175, Cell Signaling Technology, 1 : 1000), caspase 3, and β -actin (Santa Cruz Biotechnology) were employed.

Techniques: Expressing, Western Blot, Activation Assay

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Pro-BDNF Contributes to Hypoxia/Reoxygenation Injury in Myocardial Microvascular Endothelial Cells: Roles of Receptors p75 NTR and Sortilin and Activation of JNK and Caspase 3

doi: 10.1155/2018/3091424

Figure Lengend Snippet: Effects of the anti-pro-BDNF antibody on apoptosis, migration, and capillary-like-structure formation among MMECs after exposure to HG and H/R. (a) Effects of pro-BDNF on apoptosis were analyzed by flow cytometry of MMECs after different treatments: control, H/R, H/R + anti-pro-BDNF, and H/R + vehicle. (b, c) The functional effects of pro-BDNF on MMECs were assessed by capillary-like-structure formation and cell scratch assays. (d) Relative apoptosis levels and fold changes are expressed in relation to the control group. The H/R group showed markedly increased relative apoptosis levels, decreased capillary-like-structure formation, and reduced cell migration when compared with the control group. These effects were reversed by treatment with the anti-pro-BDNF antibody to the levels similar to those in the control group. The data were subjected to one-way ANOVA. The error bars represent SEM. ∗ P < 0.05 as compared with the control group; # P < 0.05 as compared with the H/R group or the H/R + vehicle group.

Article Snippet: Cells were fixed with 4% paraformaldehyde at room temperature (RT) for 30 min and permeabilized or not permeabilized with 0.5% Triton X-100 (Sigma-Aldrich), and nonspecific binding was blocked by incubation with 5% donkey serum (Jackson ImmunoResearch Laboratories) at RT for 30 min. Coverslips were incubated overnight at 4°C with the following primary antibodies: rabbit anti-BDNF (Abcam, 1 : 500), rabbit anti-pro-BDNF (Alomone Labs, 1 : 400), anti-JNK (Santa Cruz Biotechnology, 1 : 100), anti-phosphorylated-JNK (p-JNK) (Santa Cruz Biotechnology, 1 : 100), rabbit anti-p75 NTR (Santa Cruz Biotechnology, 1 : 100), and goat anti-sortilin (Abcam, 1 : 500) antibodies.

Techniques: Migration, Flow Cytometry, Functional Assay

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Pro-BDNF Contributes to Hypoxia/Reoxygenation Injury in Myocardial Microvascular Endothelial Cells: Roles of Receptors p75 NTR and Sortilin and Activation of JNK and Caspase 3

doi: 10.1155/2018/3091424

Figure Lengend Snippet: Effects of the anti-pro-BDNF antibody on the expression of p75 NTR and sortilin and apoptosis-related proteins. (a) Representative immunofluorescent images of pro-BDNF (red, first column), p-JNK (green, second column), BDNF (red, fourth column), JNK (green, fifth column), sortilin (red, seventh column), and p75 NTR (green, eighth column) in groups control, H/R, H/R + anti-pro-BDNF, and H/R + vehicle. (b–e) Representative Western blots and quantitative analysis of pro-BDNF, BDNF (b), p75 NTR , sortilin (c), JNK, p-JNK (d), caspase 3 and cleaved-caspase 3 expression (e) in response to different treatments. All the data were normalized to β -actin, and fold changes are expressed in relation to the control group. Exposure of MMECs to H/R resulted in significantly higher expression levels of pro-BDNF, p75 NTR , and sortilin and in activation of JNK and caspase 3 as compared with MMECs maintained under normal conditions (control). Nonetheless, there were no significant differences in BDNF, JNK, and caspase 3 expression levels after H/R. Treatment with the anti-pro-BDNF antibody significantly reversed the increase in the protein expression of pro-BDNF, p75 NTR , sortilin, p-JNK, and cleaved caspase 3 in MMECs after exposure to HG and H/R (H/R + anti-pro-BDNF). The data were subjected to one-way ANOVA. The error bars represent SEM. ∗ P < 0.05 as compared with the control group; # P < 0.05 as compared with the H/R or H/R + vehicle group.

Article Snippet: Cells were fixed with 4% paraformaldehyde at room temperature (RT) for 30 min and permeabilized or not permeabilized with 0.5% Triton X-100 (Sigma-Aldrich), and nonspecific binding was blocked by incubation with 5% donkey serum (Jackson ImmunoResearch Laboratories) at RT for 30 min. Coverslips were incubated overnight at 4°C with the following primary antibodies: rabbit anti-BDNF (Abcam, 1 : 500), rabbit anti-pro-BDNF (Alomone Labs, 1 : 400), anti-JNK (Santa Cruz Biotechnology, 1 : 100), anti-phosphorylated-JNK (p-JNK) (Santa Cruz Biotechnology, 1 : 100), rabbit anti-p75 NTR (Santa Cruz Biotechnology, 1 : 100), and goat anti-sortilin (Abcam, 1 : 500) antibodies.

Techniques: Expressing, Western Blot, Activation Assay

Journal: Frontiers in Cell and Developmental Biology

Article Title: Maternal immune activation-induced proBDNF-mediated neural information processing dysfunction at hippocampal CA3-CA1 synapses associated with memory deficits in offspring

doi: 10.3389/fcell.2022.1018586

Figure Lengend Snippet: The performance in the conditioning and fear memory test. PolyI:C-treated offspring were intra -CA1 or -CA3 infused with anti-proBDNF antibody or p75 NTR inhibitor TAT-Pep5 30 min before the testing. The freezing levels of polyI:C-treated offspring during the training (A) and the memory test (B) are lower while blocking the activation of proBDNF/p75 NTR signaling can effectively reversed the impaired memory behavior. Data are presented as mean ± SEM. * p < 0.05, Control + ACSFCA1 vs. other groups in A and the group vs. Control + ACSFCA1, PolyI:C + AntiCA1 and PolyI:C + Pep5CA1 in B. Training phase: Control + ACSFCA1: n = 6; PolyI:C + ACSFCA1: n = 6; PolyI:C + AntiCA1: n = 6; PolyI:C + AntiCA3: n = 6; PolyI:C + Pep5CA1: n = 6. Memory test: Control + ACSFCA1: n = 6; PolyI:C + ACSFCA1: n = 4; PolyI:C + AntiCA1: n = 7; PolyI:C + AntiCA3: n = 7; PolyI:C + Pep5CA1: n = 7.

Article Snippet: The cleavage-resistant proBDNF (2 ng/ml; Cat#B257 Alomone Labs), TAT-Pep5 (4 ng/μl; Cat#506181, EMD Millipore), or artificial CSF (ACSF) was infused bilaterally at a rate of 0.5 μl/min/side for 2 min.

Techniques: Blocking Assay, Activation Assay

Journal: Frontiers in Cell and Developmental Biology

Article Title: Maternal immune activation-induced proBDNF-mediated neural information processing dysfunction at hippocampal CA3-CA1 synapses associated with memory deficits in offspring

doi: 10.3389/fcell.2022.1018586

Figure Lengend Snippet: (A) The changes in EPSC of prymidal CA1 neurons. Typical consecutive sample traces of sEPSCs from each group (Top). The frequency of sEPSC (B) is not altered but the amplitude of sEPSC (C) is decreased in polyI:C-treated offspring. Incubation with anti-proBDNF antibody or TAT-Pep5 inhibitor can significantly enhance the declined amplitude. Data are presented as mean ± SEM. * p < 0.05, vs. Control + ACSFCA1, PolyI:C + AntiCA1 and PolyI:C + Pep5CA1. Control + ACSFCA1: n = 5; PolyI:C + ACSFCA1: n = 5; PolyI:C + AntiCA1: n = 7; PolyI:C + AntiCA3: n = 7; PolyI:C + Pep5CA1: n = 6.

Article Snippet: The cleavage-resistant proBDNF (2 ng/ml; Cat#B257 Alomone Labs), TAT-Pep5 (4 ng/μl; Cat#506181, EMD Millipore), or artificial CSF (ACSF) was infused bilaterally at a rate of 0.5 μl/min/side for 2 min.

Techniques: Incubation