Establishment method of shock rat model with liver-spleen combined injury

Abstract

The invention discloses a method for establishing a shock rat model with liver-spleen combined injury, which is characterized in that: on the basis of injury operation on the liver and spleen of a rat, when the liver and spleen of the rat are simultaneously injured, a blood loss intervention experiment is performed on the rat, so that the establishment of the shock rat model with liver-spleen combined injury is realized; the blood loss mode of the rat better conforms to the clinical blood loss condition, and the pathological process of the blood loss shock caused by the rat liver-spleen combined injury can accurately simulate the pathological process of the blood loss shock caused by the human body liver-spleen combined injury and is closer to the clinical practice; the device has the advantages that the treatment effect of a liver-spleen combined injury patient after blood substitutes are infused can be accurately simulated, and measures for coping with serious shock caused by liver-spleen combined injury can be conveniently and comprehensively studied.

Description

Technical Field

[0001] The invention relates to a method for establishing a shock rat model, in particular to a method for establishing a shock rat model with liver and spleen combined injury. Background Art

[0002] Traumatic combined liver and spleen injury is a serious multi-organ injury, often accompanied by massive bleeding and shock. Since the liver and spleen are highly vascularized organs, they are prone to massive internal bleeding after injury, leading to rapid loss of blood volume and then hemorrhagic shock. If not treated in time, it may lead to multiple organ dysfunction syndrome (MODS) and high mortality.

[0003] Due to the complexity of combined liver and spleen injuries, emergency surgery and massive blood transfusions are usually required to maintain vital signs and control bleeding. Blood substitutes have the following advantages in treating traumatic combined liver and spleen injuries in emergency situations: 1. Rapid recovery of circulating blood volume: blood substitutes can quickly expand the circulating blood volume, restore circulating blood volume, help maintain blood pressure, and prevent worsening shock; 2. Reduce the risk of transfusion reactions: compared with traditional blood, blood substitutes reduce transfusion-related risks such as infection and immune response; 3. Improve emergency efficiency: the ready-to-use and portability of blood substitutes enable them to greatly improve emergency and treatment efficiency and shorten treatment time in pre-hospital emergency and resource-constrained environments; 4. Reduce dependence on blood supply: when blood supply is tight or matching is difficult, blood substitutes provide important support and reduce dependence on traditional blood. The above advantages make blood substitutes an important tool for treating combined liver and spleen injuries in emergency situations, which helps to improve the success rate of treatment and improve patient prognosis. In order to further optimize the research and development of blood substitutes, it is of great significance to establish a shock model of combined liver and spleen injury.

[0004] The anatomical structure and physiological functions of the liver and spleen of rats are similar to those of humans, especially in terms of hemodynamics and organ damage, and can better simulate human pathological processes. In addition, because the blood volume of rats is relatively small, the amount of blood loss is easy to accurately control, and the progression of hemorrhagic shock can be clearly observed, which is convenient for quantitative analysis of the degree of injury and shock response. The existing shock rat model is easy to operate and can accurately reproduce different degrees of clinical injury through surgical techniques. It is helpful to study the treatment effects and physiological responses under different degrees of injury. It shows good feasibility in drug testing and intervention studies, and can effectively evaluate the therapeutic effects of drugs on multiple organ damage and shock. These advantages make the shock rat model an ideal experimental model for studying hemorrhagic shock caused by combined liver and spleen injury and its treatment measures.

[0005] However, the existing shock rat model achieves hemorrhagic shock by bleeding through the femoral artery, and does not perform laparotomy on the rats to perform combined liver and spleen injury. Liver injury may affect coagulation function and metabolic capacity, while spleen injury may affect immune response. Compared with single organ injury, this complex interaction of multiple organs involves injury and bleeding of two organs. Shock occurs faster and more severely, and the systemic pathological reactions it triggers are more complex, including more obvious blood loss, shock, and inflammatory reactions. Therefore, there are significant differences in physiological responses between hemorrhagic shock caused by combined liver and spleen injury and hemorrhagic shock caused by femoral artery bleeding. It is difficult for the existing shock rat model to accurately simulate the treatment effect of patients with combined liver and spleen injury after transfusion of blood substitutes, making it difficult to comprehensively study the countermeasures for severe shock caused by combined liver and spleen injury. Summary of the invention

[0006] The technical problem to be solved by the present invention is to provide a method for establishing a rat model of shock caused by combined liver and spleen injury, which can accurately simulate the therapeutic effect of patients with combined liver and spleen injury after transfusion of blood substitutes, and is convenient for comprehensively studying the countermeasures for severe shock caused by combined liver and spleen injury.

[0007] The technical solution adopted by the present invention to solve the above technical problems is: a method for establishing a shock rat model with liver and spleen combined injury, comprising the following steps:

[0008] Step 1: Weigh the rat, anesthetize it, intubate it, and perform mechanical ventilation with a ventilator;

[0009] Step 2: Perform arteriovenous puncture and catheterization on the rats. A 22F arterial catheter was placed in the right femoral artery of the rats for drawing blood gas and retaining arterial blood during the experiment, and a 22F venous catheter was placed in the left femoral vein of the rats for fluid replacement, drug administration and blood transfusion. The blood pressure and heart rate of the rats were monitored using a multi-channel physiological recorder.

[0010] Step 3, 1.3 ml of arterial blood was drawn through a 22F arterial catheter at the right femoral artery of the rat for blood gas analysis and biochemical function test, and 1.3 ml of normal saline was infused through a 22F venous catheter at the left femoral vein of the rat for supplementation;

[0011] Step 4: Perform injury operation on the rat's liver and spleen, so that the rat suffers from liver and spleen injury at the same time;

[0012] Step 5: Conduct a blood loss intervention experiment on rats. The specific process is as follows:

[0013] S5.1. The time when the rats' liver and spleen were injured at the same time was taken as the 0 minute of the experiment and the timing was started from the 0 minute of the experiment. The rats' liver incisions were wiped continuously with cotton balls to make the bleeding steady and slow while preventing the formation of blood clots. The mean arterial pressure of the rats dropped to 50-40 mmHg within 45 minutes of the experiment. That is, the mean arterial pressure of the rats was 50-40 mmHg at 45 minutes of the experiment. From 45 minutes of the experiment, the mean arterial pressure of the rats was maintained at 50-40 mmHg for 15 minutes until 60 minutes of the experiment.

[0014] S5.2. Stop wiping the rat liver incision at 60 minutes into the experiment and cover the rat liver incision with cotton balls;

[0015] S5.3, on one hand, 1.3 ml of arterial blood is extracted through the 22F arterial catheter at the right femoral artery of the rat for blood gas analysis and biochemical function test, and on the other hand, 1.3 ml of normal saline is first infused through the 22F venous catheter at the left femoral vein of the rat, and normal saline of one times the blood loss during shock blood pressure is infused through the 22F venous catheter at the left femoral vein of the rat at a rate of 2 ml / (kg.min), wherein the total blood volume of the rat is calculated according to 60 ml / kg of the rat's body weight, and 40% of the total blood volume of the rat is set as one times the blood loss during shock blood pressure;

[0016] S5.4, obtaining the mean artery of the rat at this time. If the mean artery of the rat is ≤10 mmHg at this time, the rat is determined to be dead, otherwise the rat is determined to be alive. When the rat is determined to be dead, relevant tissue specimens of the rat are retained, and the shock rat model of liver and spleen combined injury is established. When the rat is determined to be alive, proceed to step S5.5;

[0017] S5.5. The mean artery of the rat is obtained again at 120 minutes after the experiment. If the mean artery of the rat is ≤10 mmHg at this time, the rat is determined to be dead. Otherwise, the rat is determined to be alive. When the rat is determined to be dead, relevant tissue specimens of the rat are collected, and the shock rat model of liver and spleen combined injury is established. When the rat is determined to be alive, 1.3 ml of arterial blood is drawn through the 22F arterial catheter at the right femoral artery of the rat for blood gas analysis and biochemical function test. At the same time, 1.3 ml of normal saline is infused through the 22F venous catheter at the left femoral vein of the rat for supplementation, and then step S5.6 is entered;

[0018] S5.6. At 180 minutes into the experiment, the mean artery of the rat was obtained again. If the mean artery of the rat was ≤10 mmHg at this time, the rat was judged to be dead. Otherwise, the rat was judged to be alive. When the rat was judged to be dead, relevant tissue specimens of the rat were collected, and the shock rat model with combined liver and spleen injury was established. When the rat was judged to be alive, 1.3 ml of arterial blood was drawn through a 22F arterial catheter at the right femoral artery of the rat, and the rat was killed to collect relevant tissue specimens of the rat, and the shock rat model with combined liver and spleen injury was established.

[0019] Compared with the prior art, the advantage of the present invention is that when the rat suffers from simultaneous liver and spleen damage based on an injury operation on the rat liver and spleen, a blood loss intervention experiment is performed on the rat to achieve the establishment of a shock rat model with combined liver and spleen damage. During the establishment of the shock rat model, the rat's bleeding pattern is more consistent with the clinical blood loss situation, and the pathological process of hemorrhagic shock caused by combined liver and spleen damage in rats can accurately simulate the pathological process of hemorrhagic shock caused by combined liver and spleen damage in humans, which is closer to clinical reality. Therefore, the shock rat model can accurately simulate the therapeutic effect of patients with combined liver and spleen damage after transfusion of blood substitutes, making the research results more clinically relevant, and the evaluation of drug and therapeutic intervention effects is more comprehensive, which is convenient for comprehensive research on countermeasures for severe shock caused by combined liver and spleen damage.

[0020] Furthermore, in the step 1, the rat is first anesthetized, and then the rat is intubated and the rat is ventilator-assisted mechanical ventilation. The specific process is: injecting sodium pentobarbital solution into the abdominal cavity along the ventral side of the rat for general anesthesia, wherein the dose of sodium pentobarbital solution is 30 mg / kg, the anesthetized rat is fixed in a supine position on the rat operating table, tracheal intubation is established, and the catheter is fixed to the mouth or head of the rat with tape to prevent the catheter from shifting, and then the rat is ventilator-assisted mechanical ventilation is implemented.

[0021] Furthermore, the parameters of the rat ventilator were set as follows: tidal volume of 8 ml / kg, respiratory rate of 80 times / min, and inspired oxygen concentration of 21%.

[0022] Furthermore, in step 2, arteriovenous puncture and catheterization were performed on the rats, a 22F arterial catheter for extracting blood gas and retaining arterial blood during the experiment was placed in the right femoral artery of the rats, and a 22F venous catheter for fluid replacement, drug administration and blood transfusion was placed in the left femoral vein of the rats, and a multi-channel physiological recorder was used to monitor the blood pressure and heart rate of the rats; the specific process is:

[0023] S2.1. Make a longitudinal incision of 2-3 cm below the midline of the rat's neck along the horizontal line from the larynx to the top of the sternum, then separate the skin and muscle tissue layer by layer to expose the rat's internal carotid artery. Perform arterial puncture at the distal end with a 20G arterial puncture needle under direct vision, and fix the catheter and the tissue around the rat's carotid artery with surgical thread to prevent the catheter from falling off. Finally, connect the catheter to a multi-channel physiological recorder to monitor the rat's blood pressure and heart rate.

[0024] S2.2. Touch the femoral artery pulsation with your fingers in the right groin surgical area of ​​the rat, then use ophthalmic straight scissors to make a 2 cm long incision at the strongest point of the pulsation to fully expose the right femoral artery of the rat, and insert a 22F arterial catheter into the right femoral artery of the rat for drawing blood gas and retaining arterial blood during the experiment;

[0025] S2.3. Touch the femoral vein pulsation with your fingers in the surgical area of ​​the left groin of the rat, and then use ophthalmic straight scissors to cut a 2 cm long incision at the strongest point of the pulsation to fully expose the rat's left femoral vein. Insert a 22F venous catheter into the rat's left femoral vein for fluid replacement, medication, and blood transfusion.

[0026] Furthermore, in step 4, the rat liver and spleen are damaged, and the specific process of causing simultaneous liver and spleen damage in the rat is as follows: a 5 cm incision is made in the middle of the rat's abdomen, and a laparotomy is performed. After the laparotomy is completed, the rat spleen is completely severed into three sections without damaging its blood supply vessels, and a 2 cm incision is made along the longitudinal axis starting from the lower pole of the left half of the rat's liver with ophthalmic scissors, i.e., the liver incision. At this time, it is considered that the rat has simultaneous liver and spleen damage. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 This is a graph showing changes in blood pressure in a sham operation group in an embodiment of a method for establishing a shock rat model of liver and spleen combined injury of the present invention;

[0028] Figure 2 This is a graph showing changes in blood pressure in a saline resuscitation group in an embodiment of the method for establishing a shock rat model of liver and spleen combined injury of the present invention;

[0029] Figure 3 This is a graph showing changes in blood pressure in the non-resuscitation group in an embodiment of the method for establishing a shock rat model with combined liver and spleen injury of the present invention. DETAILED DESCRIPTION

[0030] The present invention is further described in detail below with reference to the accompanying drawings.

[0031] Embodiment 1: A method for establishing a shock rat model with liver and spleen combined injury, comprising the following steps:

[0032] Step 1: Weigh the rat, anesthetize it, intubate it, and perform mechanical ventilation with a ventilator;

[0033] Step 2: Perform arteriovenous puncture and catheterization on the rats. A 22F arterial catheter was placed in the right femoral artery of the rats for drawing blood gas and retaining arterial blood during the experiment, and a 22F venous catheter was placed in the left femoral vein of the rats for fluid replacement, drug administration and blood transfusion. The blood pressure and heart rate of the rats were monitored using a multi-channel physiological recorder.

[0034] Step 3, 1.3 ml of arterial blood was drawn through a 22F arterial catheter at the right femoral artery of the rat for blood gas analysis and biochemical function test, and 1.3 ml of normal saline was infused through a 22F venous catheter at the left femoral vein of the rat for supplementation;

[0035] Step 4: Perform injury operation on the rat's liver and spleen, so that the rat suffers from liver and spleen injury at the same time;

[0036] Step 5: Conduct a blood loss intervention experiment on rats. The specific process is as follows:

[0037] S5.1. The time when the rats' liver and spleen were injured at the same time was taken as the 0 minute of the experiment and the timing was started from the 0 minute of the experiment. The rats' liver incisions were wiped continuously with cotton balls to make the bleeding steady and slow while preventing the formation of blood clots. The mean arterial pressure of the rats dropped to 50-40 mmHg within 45 minutes of the experiment. That is, the mean arterial pressure of the rats was 50-40 mmHg at 45 minutes of the experiment. From 45 minutes of the experiment, the mean arterial pressure of the rats was maintained at 50-40 mmHg for 15 minutes until 60 minutes of the experiment.

[0038] S5.2. Stop wiping the rat liver incision at 60 minutes into the experiment and cover the rat liver incision with cotton balls;

[0039] S5.3, on one hand, 1.3 ml of arterial blood is extracted through the 22F arterial catheter at the right femoral artery of the rat for blood gas analysis and biochemical function test, and on the other hand, 1.3 ml of normal saline is first infused through the 22F venous catheter at the left femoral vein of the rat, and normal saline of one times the blood loss during shock blood pressure is infused through the 22F venous catheter at the left femoral vein of the rat at a rate of 2 ml / (kg.min), wherein the total blood volume of the rat is calculated according to 60 ml / kg of the rat's body weight, and 40% of the total blood volume of the rat is set as one times the blood loss during shock blood pressure;

[0040] S5.4, obtaining the mean artery of the rat at this time. If the mean artery of the rat is ≤10 mmHg at this time, the rat is determined to be dead, otherwise the rat is determined to be alive. When the rat is determined to be dead, relevant tissue specimens of the rat are retained, and the shock rat model of liver and spleen combined injury is established. When the rat is determined to be alive, proceed to step S5.5;

[0041] S5.5. The mean artery of the rat is obtained again at 120 minutes after the experiment. If the mean artery of the rat is ≤10 mmHg at this time, the rat is determined to be dead. Otherwise, the rat is determined to be alive. When the rat is determined to be dead, relevant tissue specimens of the rat are collected, and the shock rat model of liver and spleen combined injury is established. When the rat is determined to be alive, 1.3 ml of arterial blood is drawn through the 22F arterial catheter at the right femoral artery of the rat for blood gas analysis and biochemical function test. At the same time, 1.3 ml of normal saline is infused through the 22F venous catheter at the left femoral vein of the rat for supplementation, and then step S5.6 is entered;

[0042] S5.6. At 180 minutes into the experiment, the mean artery of the rat was obtained again. If the mean artery of the rat was ≤10 mmHg at this time, the rat was judged to be dead. Otherwise, the rat was judged to be alive. When the rat was judged to be dead, relevant tissue specimens of the rat were collected, and the shock rat model with combined liver and spleen injury was established. When the rat was judged to be alive, 1.3 ml of arterial blood was drawn through a 22F arterial catheter at the right femoral artery of the rat, and the rat was killed to collect relevant tissue specimens of the rat, and the shock rat model with combined liver and spleen injury was established.

[0043] In this embodiment, based on the injury operation on the rat liver and spleen, when the rat has liver and spleen injury at the same time, the rat is subjected to a blood loss intervention experiment to establish a shock rat model with liver and spleen combined injury. Compared with the previous shock rat model, it has the following characteristics:

[0044] 1. Closer to clinical reality: Liver-spleen combined injury is common in actual trauma patients, especially in cases of traffic accidents or blunt abdominal trauma. Compared with the shock rat model involving only single organ injury of the liver or spleen, the shock rat model of liver-spleen combined injury better simulates the pathological process of complex multi-organ injury and hemorrhagic shock in the clinic, making the research results more clinically relevant. And the innovative blood loss method is more in line with clinical blood loss.

[0045] 2. Synergistic effect of multiple organ injury: The shock rat model of combined liver and spleen injury allows the study of the synergistic effect of multiple organ injuries in hemorrhagic shock. For example, liver injury may affect coagulation function and metabolic capacity, while spleen injury may affect immune response. This complex interaction of multiple organs cannot be effectively simulated and studied in the shock rat model of single organ injury.

[0046] 3. More complex pathological reactions: Compared with the shock rat model of single organ injury, the systemic pathological reactions caused by combined liver and spleen injury are more complex, including more obvious blood loss, shock and inflammatory reactions. This makes the shock rat model of combined liver and spleen injury more able to reflect the complexity of post-traumatic systemic reactions, and thus better used to study the stress response of multiple systems in the body.

[0047] 4. Higher severity of injury, significant observation effect: Since combined liver and spleen injury involves injury and bleeding of two organs, shock occurs faster and is more severe, so the pathophysiological changes of severe hemorrhagic shock are more easily observed in the shock rat model of combined liver and spleen injury. In contrast, the blood loss and shock degree of the shock rat model of single organ injury are relatively mild, making it difficult to comprehensively study the countermeasures for severe shock.

[0048] 5. More comprehensive evaluation of drug and therapeutic intervention effects: The shock rat model of combined liver and spleen injury can more comprehensively test the effects of treatment strategies, especially for complex pathological processes caused by multi-organ injury. For example, when evaluating drugs or intervention measures for the treatment of hemorrhagic shock, this model can simultaneously test their protective or repair effects on multiple organs such as the liver and spleen, rather than being limited to a single organ.

[0049] 6. Multi-system impact assessment: Since liver and spleen injury affects multiple systems such as blood circulation, immunity, and metabolism, this shock rat model with combined liver and spleen injury provides a better platform for studying the interaction between multiple systems, especially in the occurrence and development of systemic inflammatory response syndrome (SIRS) and multiple organ dysfunction syndrome (MODS) after hemorrhagic shock. Therefore, the shock rat model with combined liver and spleen injury is more suitable for simulating complex clinical scenarios and can more comprehensively evaluate the pathophysiological mechanisms of post-traumatic multi-organ hemorrhagic shock and the effectiveness of intervention measures. Compared with previous shock rat models with single organ injury, it provides important advantages for multi-level and multi-angle research.

[0050] Embodiment 2: This embodiment is basically the same as Embodiment 1, except that: in this embodiment, in step 1, the rat is first anesthetized, and then the rat is intubated and the rat is ventilator-assisted mechanical ventilation. The specific process is as follows: a sodium pentobarbital solution (at a dose of 30 mg / kg) is injected into the abdominal cavity along the ventral side of the rat for general anesthesia, the anesthetized rat is fixed supinely on the rat operating table, an endotracheal intubation is established, and the catheter is fixed to the mouth or head of the rat with tape to prevent the catheter from shifting, and then the rat is ventilated and assisted mechanical ventilation is implemented,

[0051] Embodiment 3: This embodiment is basically the same as Embodiment 2, except that: in this embodiment, the parameters of the rat ventilator are set as follows: tidal volume is 8 ml / kg, respiratory rate is 80 times / minute, and inspired oxygen concentration is 21%.

[0052] Example 4: This example is basically the same as Example 3, except that: in this example, in step 2, arteriovenous puncture and catheterization are performed on rats, a 22F arterial catheter for extracting blood gas and retaining arterial blood during the experiment is inserted into the right femoral artery of the rat, and a 22F venous catheter for fluid replacement, drug administration and blood transfusion is inserted into the left femoral vein of the rat, and a multi-channel physiological recorder is used to monitor the blood pressure and heart rate of the rat; the specific process is:

[0053] S2.1. Make a longitudinal incision of 2-3 cm below the midline of the rat's neck along the horizontal line from the larynx to the top of the sternum, then separate the skin and muscle tissue layer by layer to expose the rat's internal carotid artery. Perform arterial puncture at the distal end with a 20G arterial puncture needle under direct vision, and fix the catheter and the tissue around the rat's carotid artery with surgical thread to prevent the catheter from falling off. Finally, connect the catheter to a multi-channel physiological recorder to monitor the rat's blood pressure and heart rate.

[0054] S2.2. Touch the femoral artery pulsation with your fingers in the right groin surgical area of ​​the rat, then use ophthalmic straight scissors to make a 2 cm long incision at the strongest point of the pulsation to fully expose the right femoral artery of the rat, and insert a 22F arterial catheter into the right femoral artery of the rat for drawing blood gas and retaining arterial blood during the experiment;

[0055] S2.3. Touch the femoral vein pulsation with your fingers in the surgical area of ​​the left groin of the rat, and then use ophthalmic straight scissors to cut a 2 cm long incision at the strongest point of the pulsation to fully expose the rat's left femoral vein. Insert a 22F venous catheter into the rat's left femoral vein for fluid replacement, medication, and blood transfusion.

[0056] Example 5: This example is basically the same as Example 10, except that: in this example, the rat liver and spleen are injured in step 4, and the specific process of causing simultaneous liver and spleen injury in the rat is as follows: a 5 cm incision is made in the middle of the rat's abdomen, and a laparotomy is performed. After the laparotomy is completed, the rat spleen is completely severed into three sections without damaging its blood supply vessels, and a 2 cm incision is made along the longitudinal axis starting from the lower pole of the left half of the rat's liver with ophthalmic scissors, i.e., the liver incision. At this time, it is considered that the rat has simultaneous liver and spleen injury.

[0057] In order to verify the performance of the shock rat model of combined liver and spleen injury established in the present invention, 18 SD rats were prepared, each weighing about 500 g, and the 18 SD rats were divided into three groups according to different intervention measures, with 6 rats in each group. The three groups were: sham operation group, saline resuscitation group and no resuscitation group.

[0058] The experiments were started for the 6 SD rats in the sham operation group, saline resuscitation group and no resuscitation group at the same time. The 6 SD rats in the sham operation group were operated only according to steps 1 to 3 of the method for establishing a rat model of shock with combined liver and spleen injury of the present invention. The 6 SD rats in the saline resuscitation group were operated according to the method for establishing a rat model of shock with combined liver and spleen injury of the present invention. The 6 SD rats in the no resuscitation group were operated according to steps 1 to 4 of the method for establishing a rat model of shock with combined liver and spleen injury of the present invention and S5.1 and S5.2 in step 5. During the experimental operation of the 6 SD rats in the sham operation group, saline resuscitation group and no resuscitation group, the dynamic blood pressure of each SD rat was recorded every 10 minutes from the 0 minute of the experiment until the rat died or until the 180 minute of the experiment. The sham operation group took the completion time of step 3 as the 0 minute of the experiment.

[0059] Among them, the blood pressure changes of a SD rat in the sham operation group were as follows: Figure 1 As shown in Figure 2, the blood pressure changes of a SD rat in the saline resuscitation group were as follows: Figure 2 As shown in Figure 2, the blood pressure changes of a SD rat in the no-resuscitation group were as follows: Figure 3 shown.

[0060] Figure 1 The figure shows the changes in blood pressure of SD rats in the sham operation group from 0 to 180 minutes of the experiment. Figure 1 It can be seen that the blood pressure of SD rats in the sham operation group showed a slow downward trend after the experiment began.

[0061] Figure 2 Figure 2 shows the blood pressure changes of SD rats in the saline resuscitation group from 0 to 180 minutes of the experiment. At 60 minutes, the SD rats were infused with the corresponding saline solution. Figure 2 It can be seen that the blood pressure of SD rats infused with normal saline showed an upward trend at 60 minutes of the experiment, but the blood pressure showed a downward trend around 90 minutes, and the blood pressure of SD rats was at the point of death at 180 minutes.

[0062] Figure 3 The blood pressure changes of rats in the non-resuscitation group from 0 to 180 minutes of the experiment were analyzed. Figure 3 It can be seen that the blood pressure of rats in the non-resuscitation group showed a downward trend after the experiment began, and the rats died at 160 minutes into the experiment.

[0063] The experimental data statistics of 6 SD rats in the sham operation group, saline resuscitation group and no resuscitation group are shown in Table 1:

[0064] Table 1 Experimental data statistics

[0065] 180min survival rate 120min PH value 120min lactate value 120min blood sugar level 120min BE value Sham operation group 100％ 7.42±0.02 0.7±0.5 13.3±2.0 0.3±4.8 Saline resuscitation group 66％ 7.23±0.06 9.3±2.4 29.7±2.0 -15.8±5.1 Do not resuscitate group 33％ 7.21±0.05 10.9±1.9 31.3±4.6 -18.6±3.3

[0066] Analysis of the data in Table 1 shows that: 1. Survival rate: The sham operation group was significantly higher than the other two groups; the non-resuscitation group was the lowest, only 33%. 2. pH value: The sham operation group remained within the normal range, and the resuscitation group and the non-resuscitation group showed mild acidosis. 3. Lactate value: The sham operation group had the lowest lactate level, and the resuscitation group and the non-resuscitation group had significantly increased lactate levels, indicating insufficient tissue perfusion. 4. Blood glucose level: The resuscitation group and the non-resuscitation group were significantly higher than the sham operation group, which may be related to stress response. 5. BE value: The BE values ​​of the resuscitation group and the non-resuscitation group were negative, indicating obvious metabolic acidosis, and the non-resuscitation group was the most serious.

[0067] In summary, the experiment clarified the pathological characteristics and physiological responses of the liver-spleen combined injury shock rat model established by the method for establishing the liver-spleen combined injury shock rat model of the present invention by comparing the sham operation group, the saline resuscitation group and the no-resuscitation group, verified the role of saline resuscitation in improving hemorrhagic shock, and provided a reliable experimental basis for further research on multiple organ injury and therapeutic intervention.

Claims

1. A method for establishing a shock rat model with liver and spleen combined injury, characterized in that The following steps are involved: Step 1: Weigh the rat, anesthetize it, intubate it, and perform mechanical ventilation with a ventilator; Step 2: Perform arteriovenous puncture and catheterization on the rats. A 22F arterial catheter was placed in the right femoral artery of the rats for drawing blood gas and retaining arterial blood during the experiment, and a 22F venous catheter was placed in the left femoral vein of the rats for fluid replacement, drug administration and blood transfusion. The blood pressure and heart rate of the rats were monitored using a multi-channel physiological recorder. Step 3, 1.3 ml of arterial blood was drawn through a 22F arterial catheter at the right femoral artery of the rat for blood gas analysis and biochemical function test, and 1.3 ml of normal saline was infused through a 22F venous catheter at the left femoral vein of the rat for supplementation; Step 4: Perform injury operation on the rat's liver and spleen, so that the rat suffers from liver and spleen injury at the same time; Step 5: Conduct a blood loss intervention experiment on rats. The specific process is as follows: S5.

1. The time when the rats' liver and spleen were injured at the same time was taken as the 0 minute of the experiment and the timing was started from the 0 minute of the experiment. The rats' liver incisions were wiped continuously with cotton balls to make the bleeding steady and slow while preventing the formation of blood clots. The mean arterial pressure of the rats dropped to 50-40 mmHg within 45 minutes of the experiment. That is, the mean arterial pressure of the rats was 50-40 mmHg at 45 minutes of the experiment. From 45 minutes of the experiment, the mean arterial pressure of the rats was maintained at 50-40 mmHg for 15 minutes until 60 minutes of the experiment. S5.

2. Stop wiping the rat liver incision at 60 minutes into the experiment and cover the rat liver incision with cotton balls; S5.3, on one hand, 1.3 ml of arterial blood is extracted through the 22F arterial catheter at the right femoral artery of the rat for blood gas analysis and biochemical function test, and on the other hand, 1.3 ml of normal saline is first infused through the 22F venous catheter at the left femoral vein of the rat, and normal saline of one times the blood loss during shock blood pressure is infused through the 22F venous catheter at the left femoral vein of the rat at a rate of 2 ml / (kg.min), wherein the total blood volume of the rat is calculated according to 60 ml / kg of the rat's body weight, and 40% of the total blood volume of the rat is set as one times the blood loss during shock blood pressure; S5.4, obtaining the mean artery of the rat at this time. If the mean artery of the rat is ≤10 mmHg at this time, the rat is determined to be dead, otherwise the rat is determined to be alive. When the rat is determined to be dead, relevant tissue specimens of the rat are retained, and the shock rat model of liver and spleen combined injury is established. When the rat is determined to be alive, proceed to step S5.5; S5.

5. The mean artery of the rat is obtained again at 120 minutes after the experiment. If the mean artery of the rat is ≤10 mmHg at this time, the rat is determined to be dead. Otherwise, the rat is determined to be alive. When the rat is determined to be dead, relevant tissue specimens of the rat are collected, and the shock rat model of liver and spleen combined injury is established. When the rat is determined to be alive, 1.3 ml of arterial blood is drawn through the 22F arterial catheter at the right femoral artery of the rat for blood gas analysis and biochemical function test. At the same time, 1.3 ml of normal saline is infused through the 22F venous catheter at the left femoral vein of the rat for supplementation, and then step S5.6 is entered; S5.

6. At 180 minutes into the experiment, the mean artery of the rat was obtained again. If the mean artery of the rat was ≤10 mmHg at this time, the rat was judged to be dead. Otherwise, the rat was judged to be alive. When the rat was judged to be dead, relevant tissue specimens of the rat were collected, and the shock rat model with combined liver and spleen injury was established. When the rat was judged to be alive, 1.3 ml of arterial blood was drawn through a 22F arterial catheter at the right femoral artery of the rat, and the rat was killed to collect relevant tissue specimens of the rat, and the shock rat model with combined liver and spleen injury was established.

2. The method for establishing a shock rat model with liver and spleen combined injury according to claim 1, characterized in that In the step 1, the rat is first anesthetized, and then the rat is intubated and the rat is ventilator-assisted mechanical ventilation. The specific process is: injecting sodium pentobarbital solution into the abdominal cavity along the ventral side of the rat for general anesthesia, wherein the dose of sodium pentobarbital solution is 30 mg / kg, the anesthetized rat is fixed on the rat operating table in a supine position, and the tracheal intubation is established. At the same time, the catheter is fixed to the mouth or head of the rat with tape to prevent the catheter from shifting, and then the rat is ventilator-assisted mechanical ventilation is implemented.

3. The method for establishing a shock rat model with liver and spleen combined injury according to claim 1, characterized in that The parameters of the rat ventilator were set as follows: tidal volume of 8 ml / kg, respiratory rate of 80 times / min, and inspired oxygen concentration of 21%.

4. The method for establishing a shock rat model with liver and spleen combined injury according to claim 1, characterized in that In step 2, arteriovenous puncture and catheterization were performed on the rats. A 22F arterial catheter for extracting blood gas and retaining arterial blood during the experiment was placed in the right femoral artery of the rats, and a 22F venous catheter for fluid replacement, drug administration and blood transfusion was placed in the left femoral vein of the rats, and a multi-channel physiological recorder was used to monitor the blood pressure and heart rate of the rats. The specific process is as follows: S2.

1. Make a longitudinal incision of 2-3 cm below the midline of the rat's neck along the horizontal line from the larynx to the top of the sternum, then separate the skin and muscle tissue layer by layer to expose the rat's internal carotid artery. Perform arterial puncture at the distal end with a 20G arterial puncture needle under direct vision, and fix the catheter and the tissue around the rat's carotid artery with surgical thread to prevent the catheter from falling off. Finally, connect the catheter to a multi-channel physiological recorder to monitor the rat's blood pressure and heart rate. S2.

2. Touch the femoral artery pulsation with your fingers in the right groin surgical area of ​​the rat, then use ophthalmic straight scissors to make a 2 cm long incision at the strongest point of the pulsation to fully expose the right femoral artery of the rat, and insert a 22F arterial catheter into the right femoral artery of the rat for drawing blood gas and retaining arterial blood during the experiment; S2.

3. Touch the femoral vein pulsation with your fingers in the surgical area of ​​the left groin of the rat, and then use ophthalmic straight scissors to cut a 2 cm long incision at the strongest point of the pulsation to fully expose the rat's left femoral vein. Insert a 22F venous catheter into the rat's left femoral vein for fluid replacement, medication, and blood transfusion.

5. The method for establishing a shock rat model with liver and spleen combined injury according to claim 1, characterized in that The specific process of damaging the rat liver and spleen in step 4 to cause simultaneous liver and spleen damage in the rat is as follows: a 5 cm incision is made in the middle of the rat abdomen, and a laparotomy is performed. After the laparotomy is completed, the rat spleen is completely severed into three sections without damaging its blood supply vessels, and a 2 cm incision is made along the longitudinal axis starting from the lower pole of the left half of the rat liver with ophthalmic scissors, i.e., the liver incision. At this time, it is considered that the rat has simultaneous liver and spleen damage.

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