CN108498496B

CN108498496B - Application of brilliant blue G in preparation of medicine for treating acute CO poisoning

Info

Publication number: CN108498496B
Application number: CN201810665889.3A
Authority: CN
Inventors: 杨坤丽; 李颖虹; 魏林郁; 张利彬; 黄亚迪; 李东亮
Original assignee: Xinxiang Medical University
Current assignee: Xinxiang Medical University
Priority date: 2018-06-26
Filing date: 2018-06-26
Publication date: 2020-04-07
Anticipated expiration: 2038-06-26
Also published as: CN108498496A

Abstract

The invention discloses an application of brilliant blue G in preparing a medicine for treating acute CO poisoning, and experiments prove that: the brilliant blue G is adopted to carry out intraperitoneal injection on a rat suffering from acute CO poisoning, and is found to be capable of obviously reducing the rat mortality rate caused by ACMP, relieving the edema condition of hippocampal tissues of the ACMP rat, reducing inflammatory reaction and improving the learning and memory functions.

Description

Application of brilliant blue G in preparation of medicine for treating acute CO poisoning

Technical Field

The invention relates to the field of acute CO poisoning treatment, in particular to application of brilliant blue G in preparation of a medicine for treating acute CO poisoning.

Background

Carbon monoxide poisoning is caused by inhalation through the respiratory tract of products produced when carbonaceous materials are incompletely combusted. The poisoning mechanism is that the affinity of carbon monoxide and hemoglobin is 200-fold higher than that of oxygen and hemoglobin, so that carbon monoxide is easy to combine with hemoglobin to form carboxyhemoglobin, which loses the oxygen carrying capacity and function of hemoglobin, and thus the tissue is suffocated. Has toxic effect on tissue cells of the whole body, and especially has the most serious influence on cerebral cortex. The clinical manifestations are mainly hypoxia, whose severity is proportional to the saturation of HbCO. The mild patients have headache, weakness, dizziness and difficult breathing in labor, and the saturation degree of HbCO reaches 10-20%. The symptoms are aggravated, the lips of the patients are cherry red, nausea, vomiting, blurred consciousness, collapse or coma can occur, and the saturation degree of HbCO reaches 30% -40%. Severe coma with high fever, increased tension of limb muscles and paroxysmal or tonic spasm, with HbCO saturation greater than 50%. Patients often have cerebral edema, pulmonary edema, myocardial damage, arrhythmia and respiratory depression, which can lead to death.

Brilliant blue G (also called BBG) is P2X₇Specific antagonists of R, in the study of P2X₇R plays an important role in vivo experiments of the role played in disease, and brilliant blue G has become the most widely used antagonist. Research in recent years proves that brilliant blue G has neuroprotective effect on many brain diseases. The intervention of brilliant blue G can relieve the neurological symptoms and pathological results caused by Alzheimer disease, spinal injury and cerebral ischemia. The brain damage mechanism caused by ACMP is unknown, the treatment means is single, the incidence rate of the patient prognosis delayed encephalopathy is high, but the influence of brilliant blue G on ACMP is not researched in a relevant way.

Disclosure of Invention

The invention aims to provide application of brilliant blue G in preparation of a medicine for treating acute CO poisoning so as to solve the problems in the background technology.

In order to achieve the purpose, the invention provides the following technical scheme:

the brilliant blue G is applied to the preparation of medicaments for treating acute CO poisoning.

As a further scheme of the invention: the dosage of the brilliant blue G is 26-38 mg/kg.

Compared with the prior art, the invention has the beneficial effects that: the brilliant blue G is adopted to carry out intraperitoneal injection on a rat suffering from CO poisoning, and is found to be capable of obviously reducing the death rate of the rat caused by ACMP, relieving the edema condition of hippocampal tissues of the rat suffering from ACMP, reducing inflammatory reaction and improving the learning and memory functions, has a protection effect on the hippocampal tissues suffering from acute CO poisoning, is beneficial to improving the prognosis and survival quality of ACMP and provides a new idea for clinical research on new drugs for treating CO poisoning.

Drawings

Fig. 1 is a graph showing the change of HbCO concentration in 6h, n-10, of rats after intraperitoneal injection of CO gas at different doses in the application of brilliant blue G in the preparation of a therapeutic drug for acute CO poisoning.

FIG. 2 is a graph showing the effect of BBG on the proinflammatory factor IL-1 β in hippocampal tissues in the application of brilliant blue G in the preparation of a medicine for treating acute CO poisoning.

FIG. 3 is a graph showing the effect of BBG on the hippocampal tissue pro-inflammatory factor TNF-alpha in the application of Brilliant blue G in the preparation of a drug for treating acute CO poisoning.

FIG. 4 is a graph showing the effect of BBG on the proinflammatory factor IL-6 in hippocampal tissue in the application of brilliant blue G in the preparation of a therapeutic drug for acute CO poisoning.

FIG. 5 is a graph showing the effect of BBG on the reduction of the increase in water content in hippocampal tissue in the use of brilliant blue G in the preparation of a therapeutic agent for acute CO poisoning.

Fig. 6 is a graph of the localized voyage training of brilliant blue G in the application of the preparation of a therapeutic drug for acute CO poisoning.

Fig. 7 is a bar graph of the localization navigation training of brilliant blue G in the preparation of a therapeutic drug for acute CO poisoning.

Fig. 8 is a graph showing the results of a space exploration experiment in the application of brilliant blue G in the preparation of a therapeutic drug for acute CO poisoning.

Fig. 9 is a trace diagram of a space exploration experiment in the application of brilliant blue G in the preparation of a therapeutic drug for acute CO poisoning.

FIG. 10 is a graph showing the neuronal cell damage in the CA1 region of hippocampus 3 days after 200 times carbon monoxide poisoning in the application of brilliant blue G in the preparation of a therapeutic drug for acute CO poisoning.

FIG. 11 shows the tissue P2X of hippocampus of ACMP group 6h after rat contamination in the application of brilliant blue G in the preparation of the drug for treating acute CO poisoning₇And (3) a change graph of R.

Detailed Description

The technical solution of the present patent will be described in further detail with reference to the following embodiments.

The results after injection of different doses of CO gas show: the rat mortality rate was 0 when 70ml/kg of CO gas was intraperitoneally injected, 50% when 100ml/kg of CO gas was injected, and 100% when 150ml/kg of CO gas was injected. The death rate of rats after ACMP has concentration dependence on the CO gas dose injected in the abdominal cavity, and the dose with the death rate of 45 percent is selected as the optimal molding dose, namely the dose of CO gas injected in the abdominal cavity is 100 ml/kg. See table 1.

TABLE 1 mortality of animals following intraperitoneal injection of different doses of CO gas

Dosage/ml/kg	Number/only	Death number/body	Mortality rate/%)
				70	10	0	0
100	10	5	50
				150	10	10	100

Injecting CO gas with different dosages into the abdominal cavity, and detecting for 0.5 h; 2 h; the 6h HbCO hemoglobin concentration results show: except for the control group, the HbCO protein concentration of the other three groups rises after CO gas is injected for 30min, reaches a peak within 1-2 hours, then gradually decreases, the HbCO concentration of a rat reaches 67.5% at most after 70ml/kg of CO gas is injected for 30min, and decreases to 23.43% after the rat is infected with the virus within 6 h. The highest HbCO concentration of CO gas injected by 100ml/kg reaches 80.7 percent, and the HbCO concentration is reduced to 26.78 percent after 6 hours of contamination. The HbCO concentration of the CO gas injected at 150ml/kg reaches 89.97 percent at most, and remains high after 6 hours of infection, which is shown in figure 1.

After different doses of gas CO are injected, results of the death rate of rats and the concentration of HbCO are integrated, and 100ml/kg is selected as the optimal molding concentration, so that the survival rate of experimental animals can be guaranteed, and the change trend of the HbCO after CO poisoning is closest to the reality.

After the abdominal cavity is injected with CO gas for 10min, some rats have symptoms of dysphoria, limb weakness, tachypnea, unstable walking, lethargy and the like. The symptoms are the most serious at 2h, the death rate is the highest, the rats have limb weakness and difficult walking, the skin of the exposed parts such as limbs, corners of mouths and the like is cherry red, and the dissected dead rats have visceral congestion and edema and pink lungs.

Example 1

An application of brilliant blue G in preparing a medicine for treating acute CO poisoning comprises the following specific steps:

firstly, selecting 80 rats with the age of 8-10 weeks, randomly and averagely dividing the rats into a Control group, an ACMP + brilliant blue G group and a brilliant blue G group, and then injecting CO with different doses into the abdominal cavity of the rat to prepare an ACMP model of the rat, wherein the dose of the injected CO is 100 ml/kg;

injecting brilliant blue G into rats of an ACMP + brilliant blue G group and a brilliant blue G group, wherein the injection dose of the brilliant blue G is 30mg/kg, detecting HbCO concentration in venous blood to judge CO poisoning degree, detecting the expression of P2X7RmRNA of rat hippocampal tissues by adopting an RT-PCR method, detecting the edema degree of the hippocampal tissues by adopting a dry-wet weighing method, detecting the contents of IL-1 β, TNF-alpha and IL-6 proinflammatory factors of the hippocampal tissues by adopting an ELISA method, observing the change of cytomorphology in a CA1 region of the hippocampal by HE dyeing, and evaluating learning and memory capacity by adopting a Morris water maze experiment.

As a result, it was found that: first, the survival rates were 100% for both Control and brilliant blue G groups, while the survival rate for ACMP group was 55% for the significantly lower Control group (P < 0.05); the survival rate of the ACMP + brilliant blue G group is 75 percent, is obviously improved (P is less than 0.05) compared with the ACMP group, and the brilliant blue G can reduce the death rate of rats caused by ACMP.

Secondly, the ELISA method is used for detecting the contents of IL-1 β -6 and TNF-alpha of four groups of rat hippocampal tissue proinflammatory factors, namely IL-1 β -6 and TNF-alpha, compared with a Control group and a brilliant blue G group, the contents of IL-1 β, IL-6 and TNF-alpha of an ACMP group are obviously increased (P <0.05), compared with the ACMP group, the contents of IL-1 β -alpha and IL-6 of an ACMP + brilliant blue G group are obviously reduced (P <0.05), compared with the Control group, the contents of the inflammatory factors IL-1 β -6 and TNF-alpha of the brilliant blue G group are slightly lower, but have no statistical significance, and brilliant blue G pretreatment can effectively inhibit CO from inducing release of the hippocampal tissue proinflammatory factors, as shown in figures 2-4.

Thirdly, the water content of the hippocampus tissues is detected by a dry-wet weighing method. The water content of the hippocampal tissues of the Control group and the brilliant blue G group was 79.22% + -0.82% and 78.97% + -0.71%, respectively, with no significant difference therebetween. The water content of hippocampal tissue in ACMP group is 80.72% + -0.93%, which is significantly higher than that in Control group (P < 0.05). Whereas the water content of hippocampal tissue was 79.56% ± 0.63% in ACMP + brilliant blue G group, which was significantly lower than that in ACMP group (P < 0.05). It was demonstrated that brilliant blue G pretreatment significantly improved ACMP-induced hippocampal edema, as shown in fig. 5.

Fourthly, performing positioning navigation training for 7 days, and displaying the result: escape latency for all rats was 46.25s on day 1, decreased to 14.02s on day 7, and there was no significant difference between the four groups (P >0.05), see fig. 6-a. After the rats in each group are treated by corresponding medicaments on day 8, the escape latency between the brilliant blue G group and the Control group is not statistically different, the escape latency of the ACMP group is remarkably prolonged (P <0.05) compared with that of the Control group, and the escape latency of the ACMP + brilliant blue G group is obviously shorter than that of the ACMP group (P < 0.05); a space exploration experiment is carried out, and the result shows that: the search time in the target quadrant was statistically different in the bright blue G group compared to the Control group, while the search residence time in the target quadrant was significantly shortened (P <0.05) and the trajectory was decreased in the ACMP group compared to the Control group, while the search time in the SE quadrant was significantly prolonged (P <0.05) and the trajectory was increased in the ACMP + bright blue G group compared to the ACMP group. It was demonstrated that brilliant blue G pretreatment improved the decline in learning and memory ability of rats caused by ACMP, as shown in fig. 6-9.

Fifth, HE staining method detects the morphological structure change of four groups of rat hippocampal CA1 zone nerve cells, and the result shows that: the cells of the Control group are densely and regularly arranged, the cell morphology is regular, the cytoplasm is rich, and the boundary between the nucleus and the cytoplasm is clear and visible; the ACMP group hippocampal CA1 area has reduced cell number, disordered cell arrangement, incomplete cell morphology, sparse cytoplasm, fuzzy boundary between nucleus and cytoplasm, deep staining and shrinking of nucleus, and triangular or irregular shape. Compared with the ACMP group, the nucleus fixation phenomenon of the rat hippocampus CA1 area nerve cells in the brilliant blue G + ACMP group is obviously improved, the shape is regular, and the arrangement is regular. The bright blue G group and the Control group have no obvious difference. The results show that brilliant blue G pretreatment can improve neuronal injury in the CA1 region of the hippocampus of rats caused by acute CO poisoning. The appropriate concentration of brilliant blue G alone had no effect on the morphological structure of the CA1 region of rat hippocampal neurons, as shown in fig. 10.

Sixth, RT-PCR results showed P2X in hippocampal tissue of ACMP group₇The expression level of R mRNA was significantly higher than that of Control group (P)<0.01), ACMP + BBG group P2X₇The expression level of R mRNA was significantly lower than that of ACMP group (P)<0.05), there was no statistical difference between the BBG group and the Control group, see FIG. 11.

It will be evident to those skilled in the art that the invention is not limited to the details of the foregoing illustrative embodiments, and that the present invention may be embodied in other specific forms without departing from the spirit or essential attributes thereof. The present embodiments are therefore to be considered in all respects as illustrative and not restrictive, the scope of the invention being indicated by the appended claims rather than by the foregoing description, and all changes which come within the meaning and range of equivalency of the claims are therefore intended to be embraced therein. Any reference sign in a claim should not be construed as limiting the claim concerned.

Furthermore, it should be understood that although the present description refers to embodiments, not every embodiment may contain only a single embodiment, and such description is for clarity only, and those skilled in the art should integrate the description, and the embodiments may be combined as appropriate to form other embodiments understood by those skilled in the art.

Claims

1. The brilliant blue G is applied to the preparation of medicaments for treating acute CO poisoning.

2. The use of brilliant blue G in claim 1 for the preparation of a medicament for the treatment of acute CO poisoning, wherein brilliant blue G is administered in an amount of 26-38 mg/kg.

3. Use of brilliant blue G according to claim 2 in the preparation of a medicament for the treatment of acute CO poisoning, in a dose of 30 mg/kg.