Hemin

E. faecalis OG1RF ATCC 47,077 (abbreviated to Ef) was cultured in Brain Heart Infusion (BHI) broth (Difco, USA) aerobically. F. nucleatum subsp. polymorphum ATCC 10,953 (abbreviated to Fnp) was grown in BHI broth, enriched with 5 g/L yeast extract (Thermo Fisher Scientific, USA), 0.4 g/L L-cysteine HCL (Amresco, USA), 0.005 g/L hemin (Solarbio, China), 0.001 g/L vitamin K (Ronshyn, China) anaerobically (5% CO₂, 5% H₂, 90% N₂).

Laminarin (Eisenia bicylis source) (Phaeophyceae) was purchased from Tokyo Chemical Industry Co., Ltd. (Tokyo, Japan). Sodium alginate and fucoidan (Undaria pinnatifida source) (Phaeophyceae) were brought from Sigma-Aldrich (Saint Louis, United States). P. haitanensis polysaccharides (PHP) was prepared using a method previously reported by our research team (Qiu et al., 2020 (link); Yu et al., 2023 (link)). In brief, the decolorized and deproteinized P. haitanensis powder was boiled in a water bath at 90°C for 2 h after adding 10 times volume distilled water (w/v). Centrifugation was used to extract the mixture’s supernatant, which was then combined with three liters of 95% ethanol. Following centrifugation for isolation, the precipitate was dissolved in water, frozen in a −20°C freezer, and subjected to freeze-drying to obtain the PHP. Standard dextrans with molecular weight from 4.66 kDa to 496 kDa were purchased from Aladdin (Shanghai, China). _D-glucose, _L-cysteine hydrochloride, hemin, vitamin K₁ were procured from Solarbio Science & Technology (Beijing, China). Brain heart infusion (BHI) was brought from Hopebio (Qingdao, China). All other chemicals of analytical grade were acquired from XiLong Scientific Co., Ltd. (Shantou, China).

The culture of Pg was performed as previously described [20 (link)]. Briefly, Pg (strain W83) was cultured in brain heart infusion medium (HuanKai Microbial, China) supplemented with 1% yeast extract (OXOID, UK), 0.001% menadione (Sigma, USA) and 0.5% hemin (Solarbio, China) at 37 °C for two days in an anaerobic atmosphere in use of an AnaeroPack (C-1; MGC, Tokyo, Japan). Then, the bacteria were harvested, centrifuged, and washed with phosphate buffered saline (PBS). The number of Pg was estimated by measurement of the optical density at 600 nm (OD₆₀₀).
Live Pg was suspended in a total of 1.0 × 10⁹ colony-forming units (CFU) in 0.1 mL PBS with 2% carboxymethyl cellulose (CMC) (Solarbio). This suspension was supplied to each mouse by oral gavage three times a week for 1 month, or oral administration every other day for 6 weeks. The control group was administered 0.1 mL PBS with 2% CMC without Pg.

K562 cells were cultured in RPMI-1640 medium (Sigma‒Aldrich, United States) containing 10% fetal bovine serum (FBS, Sigma‒Aldrich, United States) and 1% penicillin and streptomycin (Sigma‒Aldrich, United States). K562 cells were differentiated by erythroid induction at an initial concentration of 1 × 10⁵/mL cells. A final concentration of 40 μM hemin (Solarbio, China) and 0.1 ng/mL cytarabine (Solarbio, China) were added to the medium to induce erythroid differentiation, and the cells were placed at 37°C under hypoxic (5% CO₂, 5% O₂), CoCl (200 μM) and normoxic (5% CO₂, 20% O₂) conditions for 5 days.

PEKK-GF samples were provided by Tianfu Industrial Design (China). Mg and Ag cathode sources were provided by Borui Tiancheng Technology (China). Dulbecco’s modified Eagle’s medium (DMEM), foetal bovine serum (FBS), phosphate-buffered saline (PBS), and penicillin/streptomycin were provided by Gibco (USA). The Cell Counting Kit-8 (CCK-8) was provided by Dojindo (Japan). Live/dead viability/cytotoxicity kit, hemin, vitamin K and 4′,6-diamidino-2-phenylindole (DAPI) were provided by Solarbio (China). The MiniBEST Universal Kit, PrimeScript™ RT Kit with gDNA Eraser, and SYBR^®Premix Ex TaqTM II kit were provided by Takara (Japan). Primary antibodies were purchased from HuaBio (China). Secondary antibodies were provided by Beyotime (China). Ethanol solution was provided by Jinshan Chemical Test (China). Triton X-100, Tween20, paraformaldehyde, glutaraldehyde, phenazine methosulphate, and bovine serum albumin (BSA) were provided by Sigma-Aldrich (USA). Brain heart infusion (BHI) was purchased from OXOID (UK). XTT was purchased from Abcam (UK). The live/dead BacLight bacterial viability kit was provided by Thermo Fisher Scientific (USA). Agar was provided by BIOFROXX (Germany). Sheep blood plates were purchased from Huan Kai Microbial (China).