cd4 depletion microbeads  (Miltenyi Biotec)

Journal: Frontiers in neuroscience

Article Title: The cholinergic anti-inflammatory pathway inhibits inflammation without lymphocyte relay.

doi: 10.3389/fnins.2023.1125492

Figure Lengend Snippet: FIGURE 2 Cholinergic anti-inflammatory pathway is independent of CD4+ T-cells in mice. Wild type (A), CD4:Cre ADRB2fl/fl (B), CD4:Cre CHATfl/fl (C) mice and LysM:Cre ADRB2fl/fl (F) mice carrying (fl/fl) or not (LoxP/LoxP) the Cre transgene and RAG-1−/− (E) mice and control littermates were implanted onto the vagus or main arterial splenic nerve with a micro-cuff electrode 1 week before electrostimulation was applied. (D) Wild type mice were i.p. injected with CD4 depleting antibody or isotype control on day −3 and − 1 relative to electrostimulation. One week after surgery, LPS (5 mg/kg) was injected and electrical stimulation was applied or not (650 μA, 10 Hz, 2 min duration, −20, 0 and + 10 relative to LPS injection) to the vagus (upper panels) or main arterial splenic nerve (lower panels) in freely moving animals. Data show serum TNF levels in individual mice ±SEM of 2–5 experiments in non- electrically stimulated (NS, blue) and electrically stimulated (S, red) mice. (A–F) Mann–Whitney were performed. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.

Article Snippet: Magnetic separation of human splenocyte populations CD4 depletion was performed using human CD4 MicroBeads (Miltenyi Biotec, cat. no. 130-045-101; >99% depletion efficiency) Frontiers in Neuroscience 04 frontiersin.org according to manufacturer’s instructions.

Techniques: Control, Injection, MANN-WHITNEY

Journal: Frontiers in neuroscience

Article Title: The cholinergic anti-inflammatory pathway inhibits inflammation without lymphocyte relay.

doi: 10.3389/fnins.2023.1125492

Figure Lengend Snippet: FIGURE 3 Cholinergic anti-inflammatory pathway in nude (FoxN1−/−) mice. Nude mice were implanted onto the vagus (A) or arterial splenic (A–C) nerve with a micro-cuff electrode 1 week before electrostimulation was applied. (C) One, two or five millions CD4+ T-cell (1 M, 2 M, 5 M) or PBS (0 M) were transferred on day −1 relative to electrostimulation. One week after surgery, LPS was injected and electrical stimulation was applied or not (650 μA, 10 Hz, 2 min duration, −20, 0 and +10 relative to LPS injection) in freely moving animals. Data show serum TNF levels in individual mice ±SEM of 2–5 experiments (n ≥ 5/group) in non-electrically stimulated (NS, blue) and electrically stimulated (S, red) mice. (A,C) Mann–Whitney and (B) Kruskal–Wallis followed by Dunn’s post-hoc test were performed.

Article Snippet: Magnetic separation of human splenocyte populations CD4 depletion was performed using human CD4 MicroBeads (Miltenyi Biotec, cat. no. 130-045-101; >99% depletion efficiency) Frontiers in Neuroscience 04 frontiersin.org according to manufacturer’s instructions.

Techniques: Injection, MANN-WHITNEY

Journal: Frontiers in neuroscience

Article Title: The cholinergic anti-inflammatory pathway inhibits inflammation without lymphocyte relay.

doi: 10.3389/fnins.2023.1125492

Figure Lengend Snippet: FIGURE 5 NA promotes TNF suppression via β2-AR and is independent of T-cells in humans. (A) Flow cytometry data showing the expression of β2-AR in different populations (CD14+ and CD3+) of human splenocytes. Secondary antibody alone is shown for each cell types. (B) Dose dependent change of LPS- induced TNF caused by treatment of human peripheral blood cells with different concentrations of NA (8 × 10−5 – 8 × 102 μM), either in mixed cell culture (all cells) or culture depleted from CD4+ cells. Data are expressed as % over LPS control (n = 4). All data are expressed as mean ± SEM.

Article Snippet: Magnetic separation of human splenocyte populations CD4 depletion was performed using human CD4 MicroBeads (Miltenyi Biotec, cat. no. 130-045-101; >99% depletion efficiency) Frontiers in Neuroscience 04 frontiersin.org according to manufacturer’s instructions.

Techniques: Flow Cytometry, Expressing, Cell Culture, Control

Journal: Frontiers in neuroscience

Article Title: The cholinergic anti-inflammatory pathway inhibits inflammation without lymphocyte relay.

doi: 10.3389/fnins.2023.1125492

Figure Lengend Snippet: FIGURE 6 Cholinergic anti-inflammatory pathway revisited. The efferent arm of the inflammatory reflex, is composed of the vagus and the splanchnic nerve (SN) originating from the dorsal motor nucleus (DMV) and the intermediolateral nucleus (IML) respectively. Both nerve converge to the coeliac ganglion (CG) where they release acetylcholine (ACh) that binds to the α7 subunit of the nicotinic acetylcholine receptor (α7-AChR) relaying the neural information to the adrenergic splenic nerve (SpN). (A) In the previously described CAP (Gallowitsch-Puerta and Pavlov, 2007; Jonge and Ulloa, 2009; Andersson and Tracey, 2012; Olofsson et al., 2012), the noradrenaline (NA) release at the splenic nerve termini, which binds to β2-adrenergic receptor (β2-AR) of CD4+ T-cells, which in turn triggers the release of ACh. Engagement of the α7-AChR at the surface of macrophage by locally produced ACh inhibits their production of TNF following LPS exposition. (B) Our data support a simpler model where, NA binds directly to β2-AR to inhibit LPS-induced TNF secretion.

Article Snippet: Magnetic separation of human splenocyte populations CD4 depletion was performed using human CD4 MicroBeads (Miltenyi Biotec, cat. no. 130-045-101; >99% depletion efficiency) Frontiers in Neuroscience 04 frontiersin.org according to manufacturer’s instructions.

Techniques: Produced