2.2 Methods for Establishing Models and Evaluation

2.2 Methods for Establishing Models and Evaluation

2.2.1 Methods

2.2.1.1 Physical Methods

Duan selected healthy mice, half male and half female, and subjected them to a single whole-body irradiation with ⁶⁰Co-γ rays at a dose of 7.0 Gy, at a distance of 2 meters, with a radiation dose rate of 2.275 Gy/min, creating an AA animal model.

2.2.1.2 Chemical Methods

Zhou prepared a 0.05% suspension of miliary using distilled water and continuously administered it to mice at a dose of 4 mg/(kg·d) via gavage for 45 days, creating an AA animal model.

2.2.1.3 Mixed Methods (Physical-Chemical Methods)

Sun selected healthy Kunming strain mice, males aged 6–7 weeks, weighing 20 ± 0.3 g, and subjected them to ³⁰Co-γ ray irradiation at a dose of 3.0 Gy. On the fourth day, they began receiving cyclophosphamide (CTX) at 50.0 mg/kg and chloramphenicol (CH) at 62.5 mg/kg for three consecutive days, creating an AA animal model.

2.2.1.4 Immune-Mediated

Yao selected Balb/c mice, aged 8–12 weeks, weighing 16–20 g, regardless of sex, as recipients; DBA/2 mice, aged 8–10 weeks, regardless of sex, as donors. A lymphocyte suspension was prepared from DBA/2 mice, and Balb/c mice were immediately injected with this suspension via tail vein within 4 hours after being exposed to 6.0 Gy of γ rays, creating an AA animal model.

2.2.2 Evaluation

The animals used in all the above models are mice, each with its own advantages and disadvantages. The physical method model involves irradiating mice with sublethal or lethal doses of X-rays or gamma rays, resulting in severe hematopoietic dysfunction and a reduction in all peripheral blood cells, which is similar to acute AA in humans. However, gamma-ray irradiation affects DNA replication, inhibits cell mitosis, reduces hematopoietic stem cells, impairs immune function, and damages various organs throughout the body, leading to significant toxic side effects. If the transient damage is too severe, the mortality rate of mice is very high; if the transient damage is too mild, the hematopoietic function of mice will recover on its own. The drawback of the chemical method is that the modeling period is long and it easily causes permanent bone marrow damage. In the mixed method, CTX is an alkylating agent with cytotoxic effects that inhibits DNA synthesis and is sensitive to the hematopoietic system, allowing for relatively slow but sustained suppression of the bone marrow. CH suppresses bone marrow hematopoiesis, causing AA, which has also been reported clinically. Gamma-ray irradiation affects DNA replication, inhibits cell mitosis, reduces hematopoietic stem cells, and impairs immune function. This model is simple to operate, has a short replication cycle, a high success rate, and a low mortality rate. The disadvantage is that around day 25, the hematopoietic function of the model animals will return to normal. If this model is used for AA research, the animals should be euthanized around day 20 to measure various indicators. Currently, the most commonly used model is the immune-mediated AA model. Due to its good stability, this model has been replicated by many scholars as an animal model for AA research. However, the experimental procedure is complex, the requirements are high, and the model animals die quickly, with a mortality rate of 100%. This may be due to sublethal dose irradiation, where the dose is too large, damaging various organs throughout the body and causing excessive toxic side effects. Through preliminary experiments, we observed that whole-body irradiation of mice with ⁶⁰Co gamma rays at a height of 100 cm for 1 minute at a dose of 3.0 Gy results in a high success rate of modeling and prolongs the time until animal death.

Experimental Study