Agomir nc

Specific pathogen-free Sprague-Dawley rats (aged 8 weeks, weighing 200–250 g) were purchased from Hunan SJA Laboratory Animal (Changsha, Hunan, China). The rats were reared individually in a sterile environment with constant temperature (22°C), 50% relative humidity, and 12-h:12-h light/dark cycle. LPS (5 mg/kg) was instilled into the tracheas of the anesthetized rats to induce ALI.⁴⁴ Subsequently, the rats were treated with normal saline or 5 mg/kg LPS. Rats administered with 5 mg/kg LPS were further manipulated with NC agomir or miR-495 agomir, which were both synthesized by Guangzhou RiboBio (Guangzhou, Guangdong, China). After 12 h of LPS treatment, all rats were euthanized, and the specimens were collected for further experimentation.

ALL cell lines including Jurkat (BNCC338495), BALL-1 (BNCC102176), HPBALL (BNCC342583), Nb4 (BNCC341963), and CEM/C1 (BNCC100321) and the normal human peripheral blood mononuclear cell line PBMC (BNCC341622) were supplied by the BeNa Culture Collection (BNCC, Beijing, China). All of them were maintained in a humid thermostatic incubator containing 5% CO₂ at 37 °C. The medium used was RPMI-1640 (HyClone, Logan, UT, USA) with 10% fetal bovine serum (FBS; Gibco, Waltham, MA, USA), 100 U/mL streptomycin (Gibco), and 100 U/mL penicillin (Gibco). The medium was replaced regularly.
GenePharma (Shanghai, China) synthesized the miRNA-181b-5p mimic, miRNA-181b-5p inhibitor, small interfering RNA targeting SSX2IP, and corresponding negative controls (NCs). The miRNA-181b-5p agomir and NC agomir were procured from RiboBio (Guangzhou, China) for mouse experiments. Transfection was performed with Lipofectamine 3000 reagent (Life Technologies Corporation, Carlsbad, CA, USA).

To verify the effect of miRNA-181b-5p on ALL tumors in vivo, female BALB/c mice (SLAC Animal Center, Shanghai, China) aged 4-6 weeks were selected and randomly divided into two groups. After the BALL-1 cells were incubated with NC agomir or miRNA-181b-5p agomir (RiboBio, Guangzhou, China) for 3 days, a subcutaneous tumorigenesis test was performed on the mice. Tumor volume was evaluated once a week and tumor growth curves were plotted. The formula for this was: volume = length x width² x 0.5. Five weeks later, the mice were euthanized and the tumors were isolated and weighed.

BALB/C nude mice (n=15, female, aged 6 weeks, weighed 16–18 g, purchased from Laboratory Animal Resources, Chinese Academy of Sciences) were raised in a 12-h light/dark cycle at a temperature of 25°C±1°C and relative humidity of 50±10%. All the mice were given standard diet and sterile water ad libitum. Mice were randomly divided into A, B and C groups with 5 mice in each group. Mice in group A were inoculated with A549 cells transfected with NC Agomir (Guangzhou RiboBio Co., Ltd.) and mice in groups B and C were inoculated with A549 cells transfected with miR-205-5p Agomir (Guangzhou RiboBio Co., Ltd.). Mice in group C were additionally injected with ICA (10 mg/kg) every 3 days (4 (link)).
Tumor diameter was measured every 7 days. After the experiment, the mice were sacrificed by inhalation of CO₂ (50% of the chamber volume/min) and the tumors were resected and weighed. The flow of CO₂ was maintained for at least 2 min after respiratory arrest. All animal experiments were approved by the Institutional Animal Care and Use Committee of the First Affiliated Hospital of Zhejiang Chinese Medical University (approval No. 2021026) in accordance with the National Research Council Guide for Care and Use of Laboratory Animals.

Female BALB/c athymic nude mice (4‐6 weeks old and weighing 16‐20 g) were purchased from the Experimental Animal Centre of Soochow University and bred under pathogen‐free conditions. All animal experiments were carried out in accordance with the Soochow University Guide for the Care and Use of Experimental Animals. An miR‐338‐3p agomir and a NC agomir (RiboBio) were directly injected into the tumour, which was formed from implanted A549 cells, at a dose of 2 nmol (in 20 μL of PBS) per mouse every 4 days. After seven treatments, chemically stabilized miRNAs may have markedly improved the pharmacological properties, as described previously.13 The tumor volume (V) was determined by measuring the tumour length (L) and width (W) using Vernier callipers and applying the formula V = (L × W²) × 0.5.