Journal: PLoS Pathogens

Article Title: Targeting resident memory T cell immunity culminates in pulmonary and systemic protection against Brucella infection

doi: 10.1371/journal.ppat.1008176

Figure Lengend Snippet: (A) Groups of BALB/c mice were orally immunized with 1×10 9 CFUs of znBAZ, RB51 or sPBS on day 0, and nasally boosted with znBAZ (1×10 9 CFUs), RB51 (1×10 8 CFUs), or sPBS on day 28. After 4 weeks, mice were nasally challenged with wild-type (wt) B . abortus 2308 (5×10 4 CFUs). To assess vaccine efficacy, 4 weeks post-challenge (n = 12 mice/group), (B) lungs and (C) spleens were assessed for colonization and (D) splenic weights. Data are the mean ± SEM of tissue CFU levels and splenic weights from individual mice. Analysis of variance with Two-way ANOVA Tukey’s multiple comparisons test was performed; ****P<0.0001, ***P<0.001, **P<0.01, *P<0.05, ns indicates not significant versus sPBS-dosed mice; ++++ P < 0.0001 versus RB51-vaccinated mice. (E-G) Lungs and (H-J) splenic lymphocytes were harvested from separate groups of double-vaccinated mice at 14, 42, 56, and 84 days (4 wks post-challenge [post-chall]) post-primary immunization. Lymphocytes were restimulated with HKRB51 for 3.5 days, and collected supernatants analyzed for (E, H) IFN-γ, (F, I) TNF-α, and (G, J) IL-17a. Data depict n = 8 mice per group, and the mean of two independent experiments. The differences were determined when compared to sPBS-dosed mice: ***P<0.001, **P<0.01, *P<0.05, or compared to RB51-dosed mice §§§ P<0.001, §§ P<0.01, § P<0.05.

Article Snippet: Half of each group received IP 250 μg of either of rat IgG1 isotype control (clone TNP6A7, BioXCell) or anti-mouse IL-17 mAb (Clone 17F3, BioXCell) on days -1, day 2, 7, and 12 post-challenge.

Techniques:

Journal: PLoS Pathogens

Article Title: Targeting resident memory T cell immunity culminates in pulmonary and systemic protection against Brucella infection

doi: 10.1371/journal.ppat.1008176

Figure Lengend Snippet: Groups of BALB/c mice were orally primed, nasally boosted with sPBS, RB51, or znBAZ as described in , and on day 42 post-primary immunization, gated lung CD44 hi CD69 + CD8 + T RM cells were evaluated (A, B) to determine the source of IL-17 production by CD103 + and CD103 - T RM subsets. The total lung CD103 + and CD103 - (B) CD8 + T RM cells and (C) CD4 + T EM cells producing IL-17 were quantified; ***P<0.001, **P<0.01, *P<0.05, significant differences against sPBS-dosed or RB51-vaccinated mice are shown analysis of variance with One-way ANOVA Tukey‘s multiple comparisons test. (D-F) Additional groups of BALB/c mice were vaccinated in the same manner, and then challenged with virulent B . abortus 2308 as described above. Half of all mice were treated with isotype control Ab and the other half with anti-17 mAb on days -1, 2, 7, and 12 post-challenge. Two weeks after challenge, extent of brucellae colonization of the (D) lungs and (E) spleens were measured. (F) Splenic weights were also measured. Analysis of variance with Two-way ANOVA Tukey’s multiple comparisons test was performed: ****P<0.0001, ***P<0.001, **P<0.01, *P<0.05, versus isotype control-treated sPBS-dosed mice; ¶¶ P<0.01, ¶ P<0.05 versus isotype control-treated RB51-vaccinated mice. Data depict results from two experiments.

Article Snippet: Half of each group received IP 250 μg of either of rat IgG1 isotype control (clone TNP6A7, BioXCell) or anti-mouse IL-17 mAb (Clone 17F3, BioXCell) on days -1, day 2, 7, and 12 post-challenge.

Techniques: Mouse Assay

Journal: International Journal of Molecular Sciences

Article Title: Interleukin-17 and Th17 Lymphocytes Directly Impair Motoneuron Survival of Wildtype and FUS-ALS Mutant Human iPSCs

doi: 10.3390/ijms22158042

Figure Lengend Snippet: Impact of IL-17A on MNs: ( A ) the MAP-2 immunostaining after different concentrations (5 ng/mL, 50 ng/mL, and 500 ng/mL) of IL-17A-stimulated MNs for three days; ( B ) the viability of MNs under the same IL-17A treatment for three days was evaluated using CCK-8 assay; ( C ) the MAP-2 positive MNs, counting manually in different IL-17A treatments; ( D ) the neurite length of MAP-2 positive MNs was calculated using Neuron J in the FIJI software; ( E ) the immunostaining of MAP-2 with the same IL-17A treatment at different times (three days and six days); ( F ) the MAP-2 positive MNs’ counting in IL-17A treatment at different times; ( G ) the length of the neurite was measured under the same treatment. * p < 0.05, ** p < 0.01, *** p < 0.001 versus controls, ns: no significance.

Article Snippet: Further, 0.5 µg/mL of IL-17A neutralizing antibody (1:1,000, BPS Bioscience, San Diego, CA, USA) and 1µg/mL of human IL17RA/IL-17R antibody pretreatment (1:500, R&D Systems, Minneapolis, MN, USA) for 0.5 h prior to adding IL-17A was performed for three days.

Techniques: Immunostaining, CCK-8 Assay, Software

Journal: International Journal of Molecular Sciences

Article Title: Interleukin-17 and Th17 Lymphocytes Directly Impair Motoneuron Survival of Wildtype and FUS-ALS Mutant Human iPSCs

doi: 10.3390/ijms22158042

Figure Lengend Snippet: Effect of IL-17F on MNs: ( A ) immunostaining of MAP-2 in IL-17F (5 ng/mL, 50 ng/mL, and 500 ng/mL) stimulated MNs; ( B ) the viability of MNs under different IL-17F treatments was measured using CCK-8 assay; ( C ) MAP-2 positive MNs counting under different IL-17F treatments; (D) the neurite length was calculated, using Neuron J in the FIJI software, under different treatment conditions in both cells, ns: no significance.

Article Snippet: Further, 0.5 µg/mL of IL-17A neutralizing antibody (1:1,000, BPS Bioscience, San Diego, CA, USA) and 1µg/mL of human IL17RA/IL-17R antibody pretreatment (1:500, R&D Systems, Minneapolis, MN, USA) for 0.5 h prior to adding IL-17A was performed for three days.

Techniques: Immunostaining, CCK-8 Assay, Software

Journal: International Journal of Molecular Sciences

Article Title: Interleukin-17 and Th17 Lymphocytes Directly Impair Motoneuron Survival of Wildtype and FUS-ALS Mutant Human iPSCs

doi: 10.3390/ijms22158042

Figure Lengend Snippet: Immunostaining of the IL-17 receptor (IL-17RA and IL-17RC) and MHCI in MNs: ( A ) immunostaining of IL-17RA, IL-17RC, and MHCI in FUS-WT-EGFP, FUS-P525L-EGFP, PBMCs, and a fibroblast: ( B ) co-staining of MAP-2 with IL-17RA, IL-17RC, and MHCI in FUS-WT-EGFP and FUS-P525L-EGFP cells; ( C ) the counting of IL-17RA positive cells in MNs; ( D ) IL-17RA positive staining in MNs; ( E ) MHCI positive staining in MNs. * p < 0.05 versus controls, ns: no significance.

Article Snippet: Further, 0.5 µg/mL of IL-17A neutralizing antibody (1:1,000, BPS Bioscience, San Diego, CA, USA) and 1µg/mL of human IL17RA/IL-17R antibody pretreatment (1:500, R&D Systems, Minneapolis, MN, USA) for 0.5 h prior to adding IL-17A was performed for three days.

Techniques: Immunostaining, Staining

Journal: International Journal of Molecular Sciences

Article Title: Interleukin-17 and Th17 Lymphocytes Directly Impair Motoneuron Survival of Wildtype and FUS-ALS Mutant Human iPSCs

doi: 10.3390/ijms22158042

Figure Lengend Snippet: Antagonizing IL17 protects from IL17-induced neurodegeneration: ( A ) IL-17A-neutralizing pretreatment for 0.5 h, 50 ng/mL IL-17A stimulated the MNs for three days, MAP-2 immunostaining was performed after the treatment; ( B ) the viability of MNs under the same treatment mentioned before was calculated using CCK-8 assay; ( C ) MAP-2 positive MNs were counted and (D) the neurite length of MNs were measuredunder different treatments; ( E ) immunostaining of MAP-2 under anti-IL-17RA/IL-17R receptor pretreatment for 0.5 h, then 50 ng/mL IL-17A-stimulated for three days; ( F ) viability of MNs; ( G ) MAP-2 positive MNs counting; (H) neurite length of MAP-2 positive MNs using the aforementioned treatment.* p < 0.05, ** p < 0.01 versus controls, ns: no significance.

Article Snippet: Further, 0.5 µg/mL of IL-17A neutralizing antibody (1:1,000, BPS Bioscience, San Diego, CA, USA) and 1µg/mL of human IL17RA/IL-17R antibody pretreatment (1:500, R&D Systems, Minneapolis, MN, USA) for 0.5 h prior to adding IL-17A was performed for three days.

Techniques: Immunostaining, CCK-8 Assay

Journal: Journal of Neuroinflammation

Article Title: RhANP attenuates endotoxin-derived cognitive dysfunction through subdiaphragmatic vagus nerve-mediated gut microbiota–brain axis

doi: 10.1186/s12974-021-02356-z

Figure Lengend Snippet: Roles of SDV in rhANP-mediated reduction of LPS-induced systemic inflammation and neuroinflammation. a Treatment schedule. Mice were intraperitoneally injected with lipopolysaccharides (LPS, 5 mg/kg). Recombinant human ANP (rhANP; 1.0 mg/kg) or 0.9% saline were intraperitoneally injected to mice at 24 h before and 10 min after LPS injection. Subdiaphragmatic vagotomy (SDV) was performed 14 days prior to LPS injection. Plasma and hippocampus were collected 24 h after LPS injection. b Body weight loss in each group (two-way ANOVA: rhANP: F 1,36 = 7.058, P = 0.0117; SDV: F 1,36 = 12.72, P = 0.0010; interaction: F 1,36 = 4.923, P = 0.0329). The plasma levels of interleukin (IL)-6 ( c ; two-way ANOVA: rhANP: F 1,36 = 13.07, P = 0.0009; SDV: F 1,36 = 15.60, P = 0.0003; interaction: F 1,36 = 20.51, P < 0.0001), IL-17A ( d ; two-way ANOVA: rhANP: F 1,36 = 6.111, P = 0.0183; SDV: F 1,36 = 7.794, P = 0.0083; interaction: F 1,36 = 6.027, P = 0.0191), interferon (IFN)-γ ( e ; two-way ANOVA: rhANP: F 1,36 = 6.460, P = 0.0155; SDV: F 1,36 = 8.447, P = 0.0062; interaction: F 1,36 = 7.836, P = 0.0082), and tumor necrosis factor (TNF)-α ( f ; two-way ANOVA: rhANP: F 1,36 = 4.828, P = 0.0345; SDV: F 1,36 = 6.217, P = 0.0174; interaction: F 1,36 = 7.883, P = 0.0080). Western blot analysis of ionized calcium-binding adapter molecule 1 (iba-1) ( g ; two-way ANOVA: rhANP: F 1,36 = 6.208, P = 0.0175; SDV: F 1,36 = 4.772, P = 0.0355; interaction: F 1,36 = 4.996, P = 0.0317), IL-6 ( h ; two-way ANOVA: rhANP: F 1,36 = 6.286, P = 0.0168; SDV: F 1,36 = 6.894, P = 0.0126; interaction: F 1,36 = 8.569, P = 0.0059), IL-17A ( i ; two-way ANOVA: rhANP: F 1,36 = 4.268, P = 0.0461; SDV: F 1,36 = 3.363, P = 0.0750; interaction: F 1,36 = 5.503, P = 0.0246), interferon (IFN)-γ ( j ; two-way ANOVA: rhANP: F 1,36 = 4.706, P = 0.0367; SDV: F 1,36 = 5.685, P = 0.0225; interaction: F 1,36 = 3.530, P = 0.0684), TNF-α ( k ; two-way ANOVA: rhANP: F 1,36 = 1.598, P = 0.2144; SDV: F 1,36 = 12.72, P = 0.0010; interaction: F 1,36 = 4.345, P = 0.0443), inducible nitric oxide synthase (iNOS) ( l ; two-way ANOVA: rhANP: F 1,36 = 4.764, P = 0.0357; SDV: F 1,36 = 2.933, P = 0.0954; interaction: F 1,36 = 5.462, P = 0.0251) and their respective β-actin in the hippocampus. Data are shown as mean ± SEM, n = 10/group. * P < 0.05, ** P < 0.01, *** P < 0.0001; N.S. not significant

Article Snippet: The membranes were blocked in 5% non-fat dried milk for 1 h at room temperature, and then incubated with the following primary antibodies: rabbit polyclonal anti-ionized calcium-binding adapter molecule 1 (iba-1; 1:1000, #016-20001: Wako Pure Chemical Industries, Ltd., Tokyo, Japan), rabbit monoclonal anti-IL-6 (1:1000, #12912S, Cell Signaling Technology, Inc., Danvers, MA, USA), rabbit monoclonal anti-IL-17A (1:1000, #A0688, ABclonal, Wuhan, China), rabbit monoclonal anti-IFN-γ (1:1000, #A12450, ABclonal, Wuhan, China), rabbit polyclonal anti-TNF-α (1:1000, #A0277, ABclonal, Wuhan, China), rabbit monoclonal anti-inducible nitric oxide synthase (iNOS; 1:1000, #ab3523, Abcam, Cambridge, MA, USA), brain-derived neurotrophic factor (BDNF; 1:1000, #A4873, ABclonal, Wuhan, China), rabbit polyclonal phosphorylated tyrosine kinase receptor B (p-TrkB; 1:1000, #AP0423, ABclonal, Wuhan, China), rabbit polyclonal total TrkB (t-TrkB; 1:1000, #A2099, ABclonal, Wuhan, China), and mouse monoclonal anti-β-actin (1:1000, #A2319, ABclonal, Wuhan, China) overnight at 4 ℃.

Techniques: Injection, Recombinant, Western Blot, Binding Assay

Journal: Journal of Neuroinflammation

Article Title: RhANP attenuates endotoxin-derived cognitive dysfunction through subdiaphragmatic vagus nerve-mediated gut microbiota–brain axis

doi: 10.1186/s12974-021-02356-z

Figure Lengend Snippet: Effects of rhANP on plasma inflammatory cytokines after LPS-triggered endotoxemia. a Treatment schedule. Mice were intraperitoneally injected with lipopolysaccharides (LPS, 5 mg/kg) or 0.9% saline. Recombinant human ANP (rhANP; 1.0 mg/kg) or 0.9% saline were intraperitoneally injected to mice 24 h before and 10 min after LPS injection. Spleen and plasma were collected 24 h after injection of LPS or 0.9% saline. b Body weight loss in mice treated with rhANP or 0.9% saline 24 h after injection of LPS or 0.9% saline (one-way ANOVA: F 2,27 = 58.21, P < 0.0001). c Representative picture of spleen and spleen weight (one-way ANOVA: F 2,28 = 27.53, P < 0.0001). d Ratio of spleen weight/body weight (one-way ANOVA: F 2,28 = 20.17, P < 0.0001). e Plasma levels of interleukin (IL)-6 (one-way ANOVA: F 2,28 = 65.35, P < 0.0001). f Plasma levels of IL-17A (one-way ANOVA: F 2,28 = 16.82, P < 0.0001). g Plasma levels of interferon (IFN)-γ (one-way ANOVA: F 2,28 = 12.62, P < 0.0001). h Plasma levels of tumor necrosis factor (TNF)-α (one-way ANOVA: F 2,28 = 51.43, P < 0.0001). i There was a positive correlation (r = 0.797, P < 0.001) between spleen weight and plasma IL-6. j There was a positive correlation ( r = 0.629, P < 0.001) between spleen weight and plasma IL-17A. k There was a positive correlation ( r = 0.479, P = 0.006) between spleen weight and plasma IFN-γ. l Positive correlation ( r = 0.637, P < 0.001) between spleen weight and plasma TNF-α was observed. Data are shown as mean ± SEM, n = 10 or 11/group. ** P < 0.01, *** P < 0.0001

Article Snippet: The membranes were blocked in 5% non-fat dried milk for 1 h at room temperature, and then incubated with the following primary antibodies: rabbit polyclonal anti-ionized calcium-binding adapter molecule 1 (iba-1; 1:1000, #016-20001: Wako Pure Chemical Industries, Ltd., Tokyo, Japan), rabbit monoclonal anti-IL-6 (1:1000, #12912S, Cell Signaling Technology, Inc., Danvers, MA, USA), rabbit monoclonal anti-IL-17A (1:1000, #A0688, ABclonal, Wuhan, China), rabbit monoclonal anti-IFN-γ (1:1000, #A12450, ABclonal, Wuhan, China), rabbit polyclonal anti-TNF-α (1:1000, #A0277, ABclonal, Wuhan, China), rabbit monoclonal anti-inducible nitric oxide synthase (iNOS; 1:1000, #ab3523, Abcam, Cambridge, MA, USA), brain-derived neurotrophic factor (BDNF; 1:1000, #A4873, ABclonal, Wuhan, China), rabbit polyclonal phosphorylated tyrosine kinase receptor B (p-TrkB; 1:1000, #AP0423, ABclonal, Wuhan, China), rabbit polyclonal total TrkB (t-TrkB; 1:1000, #A2099, ABclonal, Wuhan, China), and mouse monoclonal anti-β-actin (1:1000, #A2319, ABclonal, Wuhan, China) overnight at 4 ℃.

Techniques: Injection, Recombinant

Journal: Journal of Neuroinflammation

Article Title: RhANP attenuates endotoxin-derived cognitive dysfunction through subdiaphragmatic vagus nerve-mediated gut microbiota–brain axis

doi: 10.1186/s12974-021-02356-z

Figure Lengend Snippet: Effects of rhANP on the neuroinflammation and cognitive function after LPS-triggered endotoxemia. a Treatment schedule. Mice were intraperitoneally injected with lipopolysaccharides (LPS, 5 mg/kg) or 0.9% saline. Recombinant human ANP (rhANP; 1.0 mg/kg) or 0.9% saline were intraperitoneally injected to mice 24 h before and 10 min after LPS injection. The prefrontal cortex (PFC) and hippocampus were collected 24 h after injection of LPS or 0.9% saline. b , c Western blot analysis of ionized calcium-binding adapter molecule 1 (iba-1) in the prefrontal cortex (PFC) (one-way ANOVA: F 2,27 = 6.170, P = 0.0062) and hippocampus (one-way ANOVA: F 2,27 = 5.250, P = 0.0119). d , e Western blot analysis of interleukin (IL)-6 in the PFC (one-way ANOVA: F 2,27 = 5.958, P = 0.0072) and hippocampus (one-way ANOVA: F 2,27 = 17.60, P < 0.0001). f , g Western blot analysis of IL-17A in the PFC (one-way ANOVA: F 2,27 = 4.498, P = 0.0206) and hippocampus (one-way ANOVA: F 2,27 = 6.268, P = 0.0058). h , i Western blot analysis of interferon (IFN)-γ in the PFC (one-way ANOVA: F 2,27 = 11.18, P = 0.0003) and hippocampus (one-way ANOVA: F 2,27 = 7.611, P = 0.0024). j , k Western blot analysis of tumor necrosis factor (TNF)-α in the PFC (one-way ANOVA: F 2,27 = 5.530, P = 0.0097) and hippocampus (one-way ANOVA: F 2,27 = 8.550, P = 0.0013). l , m Western blot analysis of inducible nitric oxide synthase (iNOS) in the PFC (one-way ANOVA: F 2,27 = 7.328, P = 0.0029) and hippocampus (one-way ANOVA: F 2,27 = 7.597, P = 0.0024). n , o Entries in the novel arm (one-way ANOVA: F 2,27 = 31.33, P < 0.0001) and duration in the novel arm (one-way ANOVA: F 2,27 = 13.34, P < 0.0001) in the Y maze test. p Latency to eat food in the buried food test (one-way ANOVA: F 2,27 = 7.129, P = 0.0033). Data are shown as mean ± SEM, n = 10/group. * P < 0.05, ** P < 0.01, *** P < 0.0001; N.S. not significant

Article Snippet: The membranes were blocked in 5% non-fat dried milk for 1 h at room temperature, and then incubated with the following primary antibodies: rabbit polyclonal anti-ionized calcium-binding adapter molecule 1 (iba-1; 1:1000, #016-20001: Wako Pure Chemical Industries, Ltd., Tokyo, Japan), rabbit monoclonal anti-IL-6 (1:1000, #12912S, Cell Signaling Technology, Inc., Danvers, MA, USA), rabbit monoclonal anti-IL-17A (1:1000, #A0688, ABclonal, Wuhan, China), rabbit monoclonal anti-IFN-γ (1:1000, #A12450, ABclonal, Wuhan, China), rabbit polyclonal anti-TNF-α (1:1000, #A0277, ABclonal, Wuhan, China), rabbit monoclonal anti-inducible nitric oxide synthase (iNOS; 1:1000, #ab3523, Abcam, Cambridge, MA, USA), brain-derived neurotrophic factor (BDNF; 1:1000, #A4873, ABclonal, Wuhan, China), rabbit polyclonal phosphorylated tyrosine kinase receptor B (p-TrkB; 1:1000, #AP0423, ABclonal, Wuhan, China), rabbit polyclonal total TrkB (t-TrkB; 1:1000, #A2099, ABclonal, Wuhan, China), and mouse monoclonal anti-β-actin (1:1000, #A2319, ABclonal, Wuhan, China) overnight at 4 ℃.

Techniques: Injection, Recombinant, Western Blot, Binding Assay

Journal: PLoS ONE

Article Title: Bone marrow derived-mesenchymal stem cells downregulate IL17A dependent IL6/STAT3 signaling pathway in CCl 4 -induced rat liver fibrosis

doi: 10.1371/journal.pone.0206130

Figure Lengend Snippet: Sequence of primers used in RT-qPCR analysis.

Article Snippet: Slides were washed with TBS-Triton three times and one time with TBS for 5 min. Nonspecific binding was blocked using 5% goat serum for 30 min and the blocking buffer was then removed and sections were incubated with a polyclonal anti–IL17A antibody (1:2000) (USBiological, Salem, MA, USA), polyclonal anti–IL17RA antibody (2.5μg/ml) (GeneTex, Irvine, CA, USA) and polyclonal antibody to PCNA (2.5mg/ml) (ThermoFisher Scientific, Rockford, IL, USA).

Techniques: Sequencing

Journal: PLoS ONE

Article Title: Bone marrow derived-mesenchymal stem cells downregulate IL17A dependent IL6/STAT3 signaling pathway in CCl 4 -induced rat liver fibrosis

doi: 10.1371/journal.pone.0206130

Figure Lengend Snippet: The data are presented as fold induction of mRNA expression of a . Il17a , b . Il17f , c . Il17ra , and d . Il17rc in CCl 4 /untreated (n = 5) rats compared to MSCs transplanted (CCl 4 +BM-MSCs) groups (n = 5) after 2, 4 and 6 weeks. All groups are compared with normal/diet control and oil/vehicle control rats (n = 5). Each experiment was performed in duplicates. Representative data are shown as mean±SD. * P <0.05. # P <0.05 compared to control groups.

Article Snippet: Slides were washed with TBS-Triton three times and one time with TBS for 5 min. Nonspecific binding was blocked using 5% goat serum for 30 min and the blocking buffer was then removed and sections were incubated with a polyclonal anti–IL17A antibody (1:2000) (USBiological, Salem, MA, USA), polyclonal anti–IL17RA antibody (2.5μg/ml) (GeneTex, Irvine, CA, USA) and polyclonal antibody to PCNA (2.5mg/ml) (ThermoFisher Scientific, Rockford, IL, USA).

Techniques: Expressing

Journal: PLoS ONE

Article Title: Bone marrow derived-mesenchymal stem cells downregulate IL17A dependent IL6/STAT3 signaling pathway in CCl 4 -induced rat liver fibrosis

doi: 10.1371/journal.pone.0206130

Figure Lengend Snippet: Upper panels are representatives of IL17 expression and it is noted that it was very weak in normal and paraffin/oil control group, severe in CCl 4 /fibrosis and recovery groups, high in CCl 4 +BM-MSCs/2 weeks group, moderate in CCl 4 +BM-MSCs/4weeks group and mild CCl 4 +BM-MSCs/6 weeks group. Lower panels are representatives of IL17RA expression and it is noted that it was absent in normal and paraffin/oil control group, high in CCl 4 /fibrosis and recovery groups, moderate in CCl 4 +BM-MSCs/2weeks group, mild in CCl 4 +BM-MSCs/4weeks group and nil in CCl 4 +BM-MSCs/6 weeks group (400X).

Article Snippet: Slides were washed with TBS-Triton three times and one time with TBS for 5 min. Nonspecific binding was blocked using 5% goat serum for 30 min and the blocking buffer was then removed and sections were incubated with a polyclonal anti–IL17A antibody (1:2000) (USBiological, Salem, MA, USA), polyclonal anti–IL17RA antibody (2.5μg/ml) (GeneTex, Irvine, CA, USA) and polyclonal antibody to PCNA (2.5mg/ml) (ThermoFisher Scientific, Rockford, IL, USA).

Techniques: Expressing

Journal: iScience

Article Title: Mesenchymal stem cell-derived small extracellular vesicles alleviate the immunometabolic dysfunction in murine septic encephalopathy

doi: 10.1016/j.isci.2024.110573

Figure Lengend Snippet: MSC-derived sEVs affect cytokine concentration in septic mouse cerebellum TNF-α and IL-17α assessed by immunofluorescence. (A) Representative photomicrograph of TNF-α, PV and IL-17α staining in low (x1.4, left) and high (×60, right) magnification of the area included in the hatched box. TNF-α was expressed and co-localized with the PC and their dendrites. (B and C) TNF-α expression significantly increased in SE compared to controls (5.2 ± 1.1 vs. 1.4 ± 0.2, p = 0.0085), however, treatment with MSC-derived sEVs restored its expression by more than 50% (1.9 ± 0.2, p = 0.03). Notably, TNF-α was not expressed in parvalbumin (PV)+ interneurons (a and c) that surround the PCs. (D) The expression of IL-17α was similar in control and septic mice (5.62E+09 ± 2.7E+08 vs. 7.8E+09 ± 3.7E+08, p = 0.1096), however, treatment with MSC-derived sEVs doubled its expression (1.52E+10 ± 1.3E+09, p =<0.0001). Data are represented as mean ± SEM, one-way ANOVA. Scale bar = 10 μm, Control ( n = 6), Sepsis+media [for TNF-α ( n = 7) and IL-17α ( n = 8)], Sepsis+MSC-derived sEVs [(for TNF-α ( n = 5) and IL-17α ( n = 6)]. MSC-derived sEV: mesenchymal stem cell-derived small extracellular vesicles, NS: non-significant, TNFα: tumor necrosis factor alpha, IL-17α: interleukin 17 alpha, ANOVA: Analysis of Variance, PV: parvalbumin.

Article Snippet: Antibodies against TNF-a (ServiceBio, GB11188, dilution 1:200), parvalbumin (PV) (Abcam, ab11427, dilution 1:100), IL-17a (ServiceBio, GB11110-1, dilution 1:200), GFAP (Abcam, ab7260, dilution 1:500) and appropriate secondary antibodies were used for immunofluorescent analysis.

Techniques: Derivative Assay, Concentration Assay, Immunofluorescence, Staining, Expressing, Control