Citations

… For macrophage ablation, CD11b-DTR mice received a tail vein injection of 5µg/kg diphtheria toxin (DT, cat. #150, List Bio, Campbell, CA) …

• Product #150 – Diphtheria Toxin, Unnicked, from Corynebacterium diphtheriae

Reagents

… Shiga toxin 2 from Escherichia coli was obtained from List Labs (Campbell, CA), …

• Product #162L – Shiga Toxin 2 from Escherichia coli

… Mice were injected intraperitoneally with 200 ng of pertussis toxin (List Biological laboratories, INC) on days 0 and 2 unless otherwise specified …

Author did not specify which List Labs Pertussis Toxin was utilized. List Labs provides the following Pertussis Toxin products:
• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer
• Product #181 – Pertussis Toxin from B. pertussis, Lyophilized (Salt-Free)
• Product #179A – Pertussis Toxin from B. pertussis (in Glycerol)

Induction of experimental autoimmune myositis

… BALB/c mice were each immunized intracutaneously with 50-100 μl of an emulsion into three to four locations (a total of 200 μl) on the back on days 0, 7, and 14. One hour after the first immunization, pertussis toxin (500 ng in 100 μl saline; List Biological Laboratories) was intraperitoneally injected into each animal. In the present study, all treated animals underwent successful EAM, defined by a significant increase in spleen weight.

Author did not specify which List Labs Pertussis Toxin was utilized. List Labs provides the following Pertussis Toxin products:
• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer
• Product #181 – Pertussis Toxin from B. pertussis, Lyophilized (Salt-Free)
• Product #179A – Pertussis Toxin from B. pertussis (in Glycerol)

… For PTX experiments, cells were incubated with 10 µM PTX (List Labs) overnight (o/n) at 37°C. Light stimulation employed the Xenon white light flash lamp (1 mW nominal output power, 10 ms flash separation) of the microplate reader. …

Author did not specify which List Labs Pertussis Toxin was utilized. List Labs provides the following Pertussis Toxin products:
• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer
• Product #181 – Pertussis Toxin from B. pertussis, Lyophilized (Salt-Free)
• Product #179A – Pertussis Toxin from B. pertussis (in Glycerol)

Experimental autoimmune encephalomyelitis (EAE) induction

Female C57BL/6 wildtype mice (Jackson Laboratories) or NG2creER:MAPTmGFP mice (8–12 weeks old) were injected subcutaneously with 200 μg MOG35-55 peptide (synthesized by Protein and Nucleic acid facility, Stanford University) emulsified in complete Freund’s adjuvant (CFA) (Thermo Fisher Scientific) containing 10 mg/ml of heat inactivated Mycobacterium tuberculosis H37RA (Sigma-Aldrich); injections were at one site into each hind flank. On the day of immunization and 48 h later, animals received intraperitoneal injections of 300 ng of pertussis toxin (List Biological Laboratories). …

Author did not specify which List Labs Pertussis Toxin was utilized. List Labs provides the following Pertussis Toxin products:
• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer
• Product #181 – Pertussis Toxin from B. pertussis, Lyophilized (Salt-Free)
• Product #179A – Pertussis Toxin from B. pertussis (in Glycerol)

Cell isolation and propagation

The uterine horns were collected from one 8-week-old C57BL female mice (Wuhan University Center for Animal Experiment). The Animal Ethics Committee of Wuhan University Center for Animal Experiment approved this study. Cell isolation and culture procedures were carried out according to the previous studies with minor modifications [24, 25, 29, 30]. In brief, fresh murine uterus from one female mouse was dissected and the uterine horns were collected as illustrated in Fig. 1A. Uterine horns were minced into pieces and dispersed into single cells by digestion with collagenase (StemCell, Vancouver, BC, Canada) plus trypsin. The primary mouse endometrial epithelial cells were co-cultured with irradiated mouse fibroblast 3T3 cells (J2 strain) (YongTech, Shenzhen, China) in primary epithelial culture basic medium (PECBM). PECBM contains DMEM and nutrient F-12 Ham (3:1) (v/v) (Sigma-Aldrich), supplemented with 5% FBS (GIBCO), 2 nM triiodothyronine (Sigma), 0.5% insulin–transferrin–selenium reagent (Life Technologies), 5 μg/ml transferrin (Life Technologies), 10 ng/mL epidermal growth factor (Sigma), 0.4 μg/mL hydrocortisone (Sigma), 1 nM cholera toxin (List Biological Labs), 0.5 μg/mL amphotericin B (Fungizone; Bristol-Myers Squibb), 40 μg/mL gentamicin (Gentacin; Life Technologies), and 5 to 10 mol/L Y-27632 (Enzo Life Sciences). …

• Product #100B – Cholera Toxin (AZIDE-FREE) from Vibrio cholerae

… cholera toxin (List Biological Labs, Campbell, CA) in a humid environment with 5% CO2 at 37°C. …

• Product #100B – Cholera Toxin (AZIDE-FREE) from Vibrio cholerae

… On days 0 and 2, the immunized mice received additional ip injections of 200 ng pertussis toxin (List Biological Labs). Starting 1 wk after the first immunization, EAE …

Author did not specify which List Labs Pertussis Toxin was utilized. List Labs provides the following Pertussis Toxin products:
• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer
• Product #181 – Pertussis Toxin from B. pertussis, Lyophilized (Salt-Free)
• Product #179A – Pertussis Toxin from B. pertussis (in Glycerol)

2.6 Immunocytochemistry

The following process was based on previously published procedures in the lab (Aicher et al., 2013; Hegarty et al., 2018). Prior to immunocytochemical processing, fixed stellate ganglia (SG) were rinsed with 0.1 M PB followed by 0.1 M Tris-buffered saline pH 7.6 (TS) and placed in 0.5% bovine serum albumin (BSA, Sigma-Aldrich) in TS for 30 min. SG were then incubated for 48 h at 4°C in primary antibody solution of goat anti-cholera toxin b (CTb) (1:25000, cat. 703, List Biological, RRID: AB_10013220) (Experiment 1), or goat anti-vesicular acetylcholine transporter (VAChT) (1:1000, cat. ABN100, EMD Millipore, RRID: AB_2630394) (Experiment 2), in 0.25% Triton X-100 (Sigma)/0.1% BSA in 0.1 M TS. SG were then rinsed and incubated for 2 h at room temperature in donkey anti-goat Alexa Fluor (AF) 647 secondary antibody (1:800, cat. 705–605–147, Jackson ImmunoResearch, RRID: AB_2340437) in 0.1% BSA in TS. SG were rinsed, whole mounted, and cover-slipped using Prolong Gold Antifade mountant (ThermoFisher). The goat anti-CTb antibody has been used extensively to detect the beta subunit of cholera toxin (Hegarty et al., 2010, 2018) and preadsorption of this antibody with CTb abolished immunolabeling in rat and rabbit spinal cord sections (Llewellyn-Smith et al., 1995). The VAchT antibody has been used extensively in recent rodent studies to identify cholinergic neurons using immunocytochemistry (Avila et al., 2020; Webber et al.,) and VAchT protein levels using Western blotting (Li et al., 2021; Yokoi et al., 2021).

• Product #703 – Anti-Cholera Toxin B Subunit (Goat)