CN118667088A

CN118667088A - Ferrocenyl thio dihydropyrimidinone structure polymer, preparation method and application

Info

Publication number: CN118667088A
Application number: CN202410685722.9A
Authority: CN
Inventors: 潘鹏宇; 毛腾飞; 朱昆源; 梁国标
Original assignee: General Hospital of Shenyang Military Region
Current assignee: General Hospital of Shenyang Military Region
Priority date: 2024-05-30
Filing date: 2024-05-30
Publication date: 2024-09-20

Abstract

The invention discloses a polymer with a ferrocenyl thio-dihydropyrimidinone structure, which is poly (2- (methacryloyloxy) 6-methyl-4-ferrocenyl-dihydropyrimidine-2 (1H) -thioketone-5-carboxylic acid ethyl ester-polyethylene glycol methacrylate), and also discloses a preparation method of the polymer, wherein the preparation method comprises the following steps: adding ferrocene formaldehyde, AEMA, thiourea and anhydrous magnesium chloride into acetic acid, mixing to obtain a reaction solution, pouring the reaction solution into deionized water for precipitation, filtering, ultrasonically washing, filtering again, and drying to obtain an Fc-S-AEMA monomer; adding an Fc-S-AEMA monomer, PEGMA and azodiisoheptanenitrile into DMF, uniformly stirring, introducing nitrogen, reacting at 70 ℃ for 12 hours to obtain a polymerization solution, and performing post-treatment to obtain a ferrocenyl thiodihydropyrimidinone structure polymer. The polymer disclosed by the invention is applied to medicines for relieving early brain injury after subarachnoid hemorrhage.

Description

Ferrocenyl thio dihydropyrimidinone structure polymer, preparation method and application

Technical Field

The invention belongs to the technical field of high molecular biological materials, and particularly relates to a polymer with a ferrocenyl thio-dihydropyrimidinone structure, a preparation method and application thereof.

Background

Subarachnoid hemorrhage (Subarachnoid hemorrhage, abbreviated as SAH) is a neurological disease mainly caused by rupture of intracranial aneurysms, and has high morbidity and mortality. Although management of neurological intensive care has reduced mortality from SAH over the last decades, SAH remains a disease with a very poor prognosis. In recent years, clinical research has focused on the pathophysiological mechanism within 72 hours after hemorrhage, i.e., early brain injury (Early brain injury, abbreviated as EBI). Pathophysiological processes of EBI include oxidative stress, inflammation, cerebral edema and Blood-brain barrier (BBB) disruption. EBI is one of the important factors responsible for poor prognosis in aneurysmal SAH patients. Blood after rupture of an aneurysm produces oxyhemoglobin in the subarachnoid space, resulting in the production of excess reactive oxygen species (Reactive oxygen species, ROS for short) such as superoxide anions, hydrogen peroxide, and hydroxyl radicals. ROS-induced oxidative stress plays a critical role in various pathological mechanisms that arise after SAH. Furthermore, SAH increases mitochondrial and enzyme mediated ROS production while inhibiting endogenous antioxidant protection systems. In recent years, no effective progress has been made in clinical trials of a simple neuroprotectant against subarachnoid hemorrhage, and there is a strong need for a neuroprotectant having a remarkable therapeutic effect on nerve function damage after subarachnoid hemorrhage.

Disclosure of Invention

The invention aims at the problems, overcomes the defects of the prior art, and provides the following technical scheme:

A ferrocenyl thiodihydropyrimidinone structured polymer which is poly (2- (methacryloyloxy) 6-methyl-4-ferrocenyl-dihydropyrimidine-2 (1H) -thione-5-carboxylic acid ethyl ester-polyethylene glycol methacrylate) having a molecular formula (C₂₄H₃₀FeN₂O₄S)_x(C₅H₈O₂(C₂H₄O)_m)_y, wherein x and y are integers of 20 or more, x: y has a value of 0.5-2 and m is an integer between 4-44.

It is still another object of the present invention to provide a method for preparing the above polymer of ferrocenyl thiodihydropyrimidinone structure, comprising the steps of: adding the mixture into acetic acid in a molar ratio of 1.5:0.2, mixing, oscillating at 100 ℃, reacting for 4 hours to obtain a reaction solution, wherein the concentration of ferrocene formaldehyde in the acetic acid is 2mol/L, pouring the reaction solution into deionized water for precipitation, filtering to obtain first filter residues, ultrasonically washing, filtering to obtain second filter residues, and drying the second filter residues for 24 hours to obtain an Fc-S-AEMA monomer;

Adding Fc-S-AEMA monomer, PEGMA and azodiisoheptanenitrile into DMF at a molar ratio of 1:1:0.04, wherein the concentration of the Fc-S-AEMA monomer in DMF is 0.5mol/L; stirring uniformly, introducing nitrogen to remove air in the system, then placing the mixture in a constant-temperature oil bath at 70 ℃ for reaction for 12 hours to obtain a polymerization solution, and performing aftertreatment to obtain the ferrocenyl thiodihydropyrimidinone structure polymer.

Preferably, the PEGMA has an average molecular weight of 950g/mol.

Preferably, the ultrasonic washing is to wash 3 times respectively with deionized water and a mixed solvent, wherein the time of each washing is 15min, and the mixed solvent is composed of diethyl ether and petroleum ether in a volume ratio of 1:4.

Preferably, the post-treatment process is as follows: the polymerization solution was cooled, diethyl ether was added dropwise with stirring until no more precipitate was formed, the mixture was dissolved in 5ml of THF, diethyl ether was added dropwise until no more precipitate was formed, and the mixture was filtered and dried for 24 hours to obtain a ferrocenyl thiodihydropyrimidinone-structured polymer.

Another object of the present invention is to provide a drug for alleviating early brain injury after subarachnoid hemorrhage, wherein the component of the drug is the polymer of ferrocenyl thiodihydropyrimidinone structure.

The invention has the beneficial effects that:

The ferrocenyl thio-dihydropyrimidinone structure polymer reduces oxidative stress injury of mice after SAH through Nrf2/HO-1 signal path; the protective effect is shown in the nervous system after SAH, and the effect of reducing early brain injury after subarachnoid hemorrhage is achieved by improving the nerve function, protecting the integrity of blood brain barrier, inhibiting activation of glial cells, reducing neuronal apoptosis and reducing inflammatory response.

Drawings

FIG. 1 shows the modified Garcia score results at 24h and 72h after SAH.

Fig. 2 shows the results of the balance beam experiments at 24h and 72h after SAH.

FIG. 3 shows the results of measuring the brain water content 24 hours after SAH.

Fig. 4 shows the evans blue permeability 24h after SAH.

FIG. 5 shows Western blot expression of Claudin-5, occludin, ZO-1 and beta-actin.

FIG. 6 is a graph of the relative densities of Claudin-5, occludin, ZO-1.

FIG. 7 shows Western blot expression of Nrf2, HO-1 and beta-actin.

Fig. 8 is a graph of comparative densities of Nrf2 and HO-1.

FIG. 9 shows Western blot expression of Bax, bcl-2, caspase3, CHOP1 and beta-actin.

FIG. 10 is a graph of relative densities of Bax, bcl-2, caspase3, CHOP 1.

FIG. 11 shows Western blot expression of IL-1. Beta. IL-18 and. Beta. -actin.

FIG. 12 is a graph of IL-1. Beta. And IL-18 relative density comparisons.

FIG. 13 shows the results of astrocyte GFAP immunofluorescence staining.

FIG. 14 shows the ratio of GFAP positive cells to normal cells.

FIG. 15 shows the result of immunofluorescent staining of microglial cells Iba-1.

FIG. 16 is a graph showing the ratio of Iba-1 positive cells to normal cells.

FIG. 17 shows the result of neuronal Tunel immunofluorescence staining.

Fig. 18 is a ratio of Tunel positive neurons to normal neurons.

In the figure, represents a set of data, such as: six mice generated six sets of data, six.

In the figure, the sign of the significance difference is given. Wherein represents P <0.05; * Represents P <0.01; * Represents P <0.001; * Represents P <0.0001; ns represents P.gtoreq.0.05.

Detailed Description

In order to make the technical problems, technical schemes and beneficial effects solved by the invention more clear, the invention is further described in detail below with reference to specific embodiments. It should be understood that the detailed description is presented by way of example only and is not intended to limit the invention.

Example 1

10Mmol of ferrocene formaldehyde, 10mmol of AEMA, 15mmol of thiourea and 2mmol of anhydrous magnesium chloride are added to 5mL of acetic acid, and then poured into a centrifuge tube, wherein the anhydrous magnesium chloride is used as a catalyst and the acetic acid is used as a solvent. And (3) placing the centrifuge tube in a mixing instrument at the temperature of 100 ℃ for oscillation reaction. After 4H of reaction, reaction liquid is obtained, the reaction liquid is poured into deionized water for precipitation, the first filter residue is obtained by filtering, the deionized water and the mixed solvent are respectively washed for 3 times by ultrasonic, the time of each washing is 15min, the second filter residue is obtained by filtering, and the second filter residue is dried for 24H to obtain black powdery ethyl 2- (methacryloyloxy) 6-methyl-4-ferrocenyl-dihydropyrimidine-2 (1H) -thione-5-carboxylate which is marked as Fc-S-AEMA monomer.

Into a 10mL Schlenk flask were charged 2mmol of Fc-S-AEMA monomer, 2mmol of PEGMA, 0.08mmol of azobisisoheptonitrile and 4mL of DMF as solvent. Adding a magneton, uniformly stirring, sealing by a rubber plug, introducing nitrogen into a 10mL Schlenk reaction bottle, bubbling for 15min, removing air in the system, then placing into a constant-temperature oil bath at 70 ℃ for reaction for 12h, obtaining a polymerization solution after polymerization, and obtaining a ferrocenyl thio-dihydropyrimidinone structure polymer, which is abbreviated as DHPMs.

The post-treatment process comprises the following steps: the polymerization solution was cooled, diethyl ether was added dropwise with stirring until no more precipitate was formed, the mixture was dissolved in 5ml of THF, diethyl ether was added dropwise until no more precipitate was formed, and the mixture was filtered and dried for 24 hours to obtain a ferrocenyl thiodihydropyrimidinone-structured polymer.

1. Description of the experiment

1. Experimental animals and design

All experimental animals in the experiment of the invention are approved by the ethical committee of the general hospital in the North war zone of the liberation army of Chinese people, and conform to the laboratory animal use and care guidelines and ARRIVE guidelines issued by the national institutes of health. These animals were from the national center for laboratory animals in the North war zone of the liberated army.

A total of 126 mice were randomly divided into 7 groups: sham group (n=30), SAH group (n=6), sah+vehicle group (n=30), sah+ DHPMs50mg/kg group (n=6), sah+ DHPMs mg/kg group (n=30), sah+ DHPMs mg/kg group (n=6) and sah+eda10mg/kg group (n=18). The experimental animal is in an SPF-grade experimental animal house, the environment is standardized, and sufficient food and water are ensured. The illumination environment is a standard photoperiod (12 h darkness/12 h illumination).

2. Dosage of drug

15Min after SAH induction, mice were intraperitoneally injected with ferrocenyl thiodihydropyrimidinone structured polymers at doses of 50mg/kg, 300mg/kg and 500mg/kg, respectively, and 10mg/kg of EDA. sham group received no injection as control group, while sah+vehicle group received injection of sterile physiological saline containing 1% alcohol.

3. Mouse SAH model

The experiment adopts an intravascular threading method to establish a mouse SAH model in the left brain, and the specific operation steps are as follows: first, 1% sodium pentobarbital was injected intraperitoneally into mice at a dose of 40mg/kg, and the mice were anesthetized. When the mice were free of palpebral reflex, they were supine and the limbs and head were fixed on the mouse fixation plate. The body temperature of the mice was maintained at 37 ℃ by monitoring the rectal temperature throughout. Neck hair was removed under a microscope and the skin was disinfected with iodophor. An incision of 25-30mm was made along the midline of the neck, and the muscles and fascia were peeled off layer by layer until the Common Carotid Artery (CCA), internal Carotid Artery (ICA) and External Carotid Artery (ECA) were completely exposed. The distal end of ECA was ligated using a suture, and two non-invasive arterial clamps were placed in sequence at the proximal end of CCA and the distal end of ICA, temporarily clamping the vessel. A "V" shaped incision was made at about 3mm at the start of the ECA and a 15mm length nylon suture was inserted along the incision into the CCA. Ligature at the bifurcation of ECA, fix nylon suture, prevent it from sliding out. The ECA is then completely severed and the clip of ICA is released. Nylon sutures were advanced to ICA about 8-10mm until resistance was felt. After further advancing about 1mm, the nylon suture was pulled out and tied after feeling the sense of breakthrough, and the residual ECA was completely ligated. The CCA was confirmed to be clear prior to suturing the skin and re-sterilizing. After the operation, the mice were placed on a constant temperature pad at 37 ℃ to recover the state, and then returned to the animal room.

Sham groups performed the same procedure as SAH and sah+vehicle groups, but did not puncture blood vessels.

4. Neural function scoring

The experiment adopts two evaluation methods: modified Garcia scoring and balance beam experiments, neural behavioral defect levels 24h and 72h after SAH were evaluated blindly.

The modified Garcia score included six tests aimed at assessing various aspects of neurological function, with a total of 18 points assigned to assess the severity of neurological impairment. These tests included assessment of spontaneous activity, spontaneous limb movement, forelimb extension, climbing, proprioception, and response to beard stimulation. The score range for each individual test was 3-18 points. Higher scores in this range indicate improved neurological function for the corresponding test.

The specific flow of the balance beam experiment is as follows: mice were placed on the balance beam and their distance to travel through the balance beam was measured within 1 min. Subsequently, the distance traveled by the mice was scored on a scale of 0 to 5, with higher scores indicating better motor coordination and balance.

SAH hierarchical assessment

The present study used an 18 point grading system to assess the severity of SAH. The skull base of the mice was divided into six areas and the extent of bleeding in each area was assessed using a 0 to 3 grading standard. A score of 0 indicates no bleeding and a score of 1 indicates little bleeding. A score of 2 indicates bleeding with a clearly visible vascular morphology, while a score of 3 indicates massive bleeding with an unclear vascular morphology. The overall severity score for bleeding is obtained by summing the scores of all six regions. Scores from 0 to 7 represent mild bleeding, scores from 8 to 12 represent moderate bleeding, and scores from 13 to 18 represent severe bleeding. Mice with SAH score below 8 score were discharged.

6. Brain water content

Mice were euthanized using the cervical method, and the brain tissue was then placed in an oven at 105 ℃ for 24 hours to completely remove moisture. After drying was completed, the weight of the brain was measured again to determine its dry weight. The formula for calculating the cerebral water content is as follows:

brain water content = (wet weight-dry weight)/wet weight x 100%

7. Evan blue permeability assessment

Evan blue dye permeation was used to evaluate disruption of the blood brain barrier.

24H after SAH, mice were intravenously injected with 2% concentration of Evans blue dye (4 ml/kg) via the left femoral vein under 1% sodium pentobarbital anesthesia, followed by a 60min cycle. To clear residual dye from the blood vessels, mice were then perfused with PBS buffer. Brain tissue was collected and homogenates were prepared in saline solution. Next, the brain tissue homogenate was centrifuged at 15000rpm for 30min using a refrigerated centrifuge, and the supernatant above the supernatant precipitation was mixed with an equal amount of trichloroacetic acid to induce protein precipitation. After overnight incubation at 4 ℃, the mixture was centrifuged again at 15000rpm to collect the final supernatant. The concentration of Evan's blue dye present in the supernatant was measured using a spectrophotometer, and absorbance at a wavelength of 620nm was measured.

8. Western blot analysis (Western blotting)

Mouse brain tissue was extracted, and protein was extracted from the left brain hemisphere, and protein analysis was performed by Western blotting. 40. Mu.g of protein was added to each gel well and gel electrophoresis was performed. Subsequently, the proteins were transferred to nitrocellulose membranes and blocked with blocking liquid for 60min at room temperature. After blocking, nitrocellulose membranes were incubated overnight at 4 ℃ with a properly diluted primary antibody. Then, the secondary antibody was incubated with appropriate dilution at room temperature for 2h. The primary secondary antibodies included goat anti-rabbit IgG/HRP and goat anti-mouse IgG/HRP. Imaging was performed using a chemiluminescent imaging system and densitometric analysis was performed using ImageJ software.

9. Immunofluorescent staining

Immunofluorescent staining was performed using fixed frozen sections. After anesthetizing the mice with 1% sodium pentobarbital, heart perfusion was performed with PBS buffer (4 mgPFA/100ml PBS) containing a mass percentage concentration of 4%. Brains were immediately collected, soaked in 4% PFA, and stored at 4 ℃ for 24h. Subsequently, the tissue was dehydrated in a sucrose solution (30 mg sucrose/100 ml PBS) with a mass fraction of 30% for 24 hours, flash frozen at-35℃and then 8 μm coronal brain sections were prepared using a microtome. To render the sections permeable, they were perforated with a 0.3% Triton X-100 solution, treated at room temperature for 30min and then blocked with BSA at the same temperature for 60min. Then, incubation with primary antibody was performed at 4 ℃. Subsequently, the sections were incubated with a fluorescent secondary antibody corresponding to the primary antibody source at room temperature for 60min. The main secondary fluorescent antibodies include goat anti-mouse AF555 and goat anti-rabbit AF555. Apoptotic cells were then detected using TUNEL apoptosis detection kit according to the manufacturer's instructions. Finally, the cells were blocked with an anti-fluorescence quencher containing DAPI. The sections were observed using a fluorescence microscope to analyze molecular localization.

10. Statistics and analysis

Data are presented as mean ± Standard Deviation (SD). All data were checked for normalization and variance alignment (Brown-Forsythe test). Results between the two groups were compared using independent t-test, and a comparison was made between the groups to select Bonferroni corrected one-way analysis of variance (ANOVA). Statistical analysis was performed using GRAPHPAD PRISM 9.0.0, with statistical significance defined as P less than 0.05.

2. Experimental results

1. Polymers of ferrocenyl thiodihydropyrimidinone structure can improve neurological function after SAH

The neurological function of each group of mice was assessed by modified Garcia score (fig. 1) and balance bar test (fig. 2). As can be seen from the bar graphs of fig. 1 and 2, the SAH and sah+vehicle groups showed significantly reduced modified Garcia score and balance beam experimental score, respectively, at 24h and 72h after SAH compared to sham group, indicating significant nerve damage in mice after SAH.

It can also be seen in figures 1 and 2 that mice treated with DHPMs were significantly improved in 24h and 72h post SAH modified Garcia score and balance test results compared to SAH and sah+vehicle groups, with the most significant improvement observed at 300mg/kg dose.

Meanwhile, we have determined the antioxidant efficacy of DHPMs by comparison with the known antioxidants EDA. There were no statistically significant differences between 24h and 72h modified Garcia scores and balance beam test results for the sah+ DHPMs group compared to the sah+eda group.

From these results, we conclude that DHPMs can alleviate the neurological impairment following SAH, with an effect comparable to that of the well-known antioxidant EDA, and with an optimal effect at a dose of 300 mg/kg.

Therefore, we selected DHPMs mg/kg as the optimal dose for the subsequent experiments.

2. The ferrocenyl thio dihydropyrimidinone structural polymer can improve blood brain barrier injury after SAH

To explore the potential ameliorating effect of DHPMs on BBB injury after SAH, we measured brain water content 24h after SAH induction in each group of mice using a dry-wet method (fig. 3) and assessed the permeability of evans blue (fig. 4). These measurements reflect changes in severity of cerebral edema and BBB permeability.

As shown in FIG. 3, the brain water content of mice in SAH+Vehicle group was significantly higher than that in sham group and the brain water content of mice in SAH+ DHPMs group of 300mg/kg was significantly lower than that in SAH+Vehicle group at 24h post SAH.

As shown in FIG. 4, the SAH+Vehicle group of mice showed a significant increase in Evan blue permeability, while 300mg/kg of SAH+ DHPMs group of mice showed a decrease in Evan blue permeability over the SAH+Vehicle group.

Furthermore, DHPMs was found to exhibit more pronounced effects in reducing brain water content and evans blue extravasation when compared to the sah+eda group.

The Claudin-5, occludin and ZO-1 tight junctions play a key role in the structure and function of the BBB. Changes in the expression levels of these tight junction proteins are associated with changes in the integrity and function of the BBB. Thus, we used Western blotting to quantitatively measure the expression levels of Claudin-5, occludin and ZO-1 proteins (as shown in FIG. 5).

As can be seen from FIGS. 5 and 6, the expression levels of the Claudin-5, occludin and ZO-1 proteins were reduced in the mice of the SAH+Vehicle group compared to the sham group. In contrast, the expression levels of the mouse Claudin-5, occludin and ZO-1 proteins were significantly higher in the DHPMs group at 300mg/kg than in the Vehicle group. These findings indicate DHPMs are able to alleviate blood brain barrier damage in SAH mice.

3. The ferrocenyl thio-dihydropyrimidinone structure polymer can reduce activation of glial cells and apoptosis of nerve cells after SAH

After SAH, excessive ROS generated by oxidative stress induces activation of glial cells and apoptosis of neural cells, significantly affecting early pathophysiological changes and adverse consequences. SAH generally triggers inflammation and microglial activation, resulting in increased numbers of glial cells, altered phenotypes, and release of inflammatory mediators, further exacerbating oxidative stress and inflammatory responses. Iba-1 is a calbindin expressed exclusively in microglia, whereas GFAP is exclusively present in the cell body of astrocytes. Thus, astrocyte GFAP immunofluorescent staining and microglial Iba-1 immunofluorescent staining were performed, and the results are shown in fig. 13 and 15.

Meanwhile, as can be seen from the results of the ratio of GFAP-positive cells to normal cells in FIG. 14 and the ratio of Iba-1-positive cells to normal cells in FIG. 16, 24 hours after SAH, both microglia and astrocytes were activated as compared with sham group. However, activation of glial cells was reduced in mice treated with DHPMs mg/kg.

Furthermore, we studied the effect of DHPMs on SAH mouse neuronal apoptosis using Tunel staining method (fig. 17). As can be seen from the results of fig. 17, 18, the number of Tunel positive neurons increased after SAH, indicating increased neuronal apoptosis, whereas in mice treated with DHPMs mg/kg, the number of Tunel positive neurons was significantly reduced.

We further evaluated the level of apoptotic proteins using Western blotting. As shown in the results of fig. 9, 10, bax, caspase3 and Chop1 levels were significantly increased in the sah+veccle group, while Bcl-2 levels were significantly decreased, compared to the sham group, indicating increased neuronal apoptosis after SAH. However, bax, caspase3 and Chop1 levels were significantly reduced and Bcl-2 levels were significantly increased after receiving DHPMs dry at 300 mg/kg. Together, these findings indicate that DHPMs can reduce glial activation and apoptosis.

4. The ferrocenyl thio-dihydropyrimidinone structure polymer can reduce inflammatory reaction after SAH

The levels of inflammatory cytokines IL-1. Beta. And IL-18 were measured in brain tissue following SAH using Western blotting (FIG. 11). From the results in FIG. 12, it is shown that IL-1β and IL-18 levels were significantly increased in the SAH+Vehicle group compared to the sham group, indicating an increased inflammatory response after SAH. In contrast, the levels of IL-1β and IL-18 were significantly reduced in SAH+ DHPMs300mg/kg group. These findings demonstrate that DHPMs at 300mg/kg can significantly reduce the SAH-induced inflammatory response.

5. The ferrocenyl thio dihydropyrimidinone structure polymer can relieve oxidative stress reaction after SAH through Nrf2-HO-1 signal path

Nrf2 is a transcription factor that is critical for cellular antioxidant stress response. Upon receipt of oxidative stress or other damage signals, nrf2 is transferred from the cytoplasm to the nucleus and activates transcription of a range of antioxidant stress genes, including HO-1.HO-1 is taken as an important downstream target gene of Nrf2, participates in metabolism of heme and plays roles in protecting against oxidization, inflammation, apoptosis and the like. The Nrf2/HO-1 signaling pathway plays a key regulatory role in cellular responses to oxidative stress, inflammation and other damaging stimuli, and is thought to be one of the key mechanisms to maintain cellular homeostasis and promote cellular survival. Therefore, it is of great importance to study the mechanisms and potential therapeutic value of the Nrf2/HO-1 signaling pathway in a variety of diseases. To elucidate the mechanism of action of DHPMs in SAH, we used Western blotting to determine protein levels of Nrf2 and HO-1 (FIG. 7). The results in FIG. 8 show that SAH+Vehicle group showed increased levels of Nrf2 protein expression and decreased HO-1 compared to sham group. In contrast, in the DHPMs mg/kg group, the Nrf2 protein level was significantly reduced and HO-1 was significantly increased compared to the SAH+Vehicle group. These results indicate that DHPMs therapeutic effects on SAH are mediated through the Nrf2/HO-1 signaling pathway.

Experimental results show that DHPMs reduces oxidative stress injury of mice after SAH through Nrf2/HO-1 signaling pathway. DHPMs exhibits protective effects in the mouse nervous system following SAH by improving neurological function, protecting blood brain barrier integrity, inhibiting glial cell activation, reducing neuronal apoptosis, and reducing inflammatory response. Furthermore, DHPMs is superior to EDA in terms of reducing cerebral oedema and reducing blood brain barrier damage. Our studies have determined DHPMs to be useful as a drug to play a neuroprotective role in SAH therapy.

It should be understood that the foregoing detailed description of the present invention is provided for illustration only and is not limited to the technical solutions described in the embodiments of the present invention, and those skilled in the art should understand that the present invention may be modified or substituted for the same technical effects; as long as the use requirement is met, the invention is within the protection scope of the invention.

Claims

1. A ferrocenyl thio dihydropyrimidinone structure polymer, the method is characterized in that: the polymer is poly (2- (methacryloyloxy) 6-methyl-4-ferrocenyl-dihydropyrimidine-2 (1H) -thioketone-5-carboxylic acid ethyl ester-polyethylene glycol methacrylate), and the molecular formula is (C₂₄H₃₀FeN₂O₄S)_x(C₅H₈O₂(C₂H₄O)_m)_y,, wherein x and y are integers more than 20, x: y has a value of 0.5-2 and m is an integer between 4-44.

2. A method for preparing a ferrocenyl thiodihydropyrimidinone structured polymer according to claim 1, wherein: ferrocene formaldehyde, AEMA, thiourea, anhydrous magnesium chloride in a ratio of 1:1: adding the mixture into acetic acid in a molar ratio of 1.5:0.2, mixing, oscillating at 100 ℃, reacting for 4 hours to obtain a reaction solution, wherein the concentration of ferrocene formaldehyde in the acetic acid is 2mol/L, pouring the reaction solution into deionized water for precipitation, filtering to obtain first filter residues, ultrasonically washing, filtering to obtain second filter residues, and drying the second filter residues for 24 hours to obtain an Fc-S-AEMA monomer;

3. A method for preparing a ferrocenyl thiodihydropyrimidinone structured polymer according to claim 2, wherein: the average molecular weight of the PEGMA is 950g/mol.

4. A method for preparing a ferrocenyl thiodihydropyrimidinone structured polymer according to claim 2, wherein: the ultrasonic washing is to wash for 3 times respectively by deionized water and a mixed solvent, wherein the washing time is 15min each time, and the mixed solvent is composed of diethyl ether and petroleum ether in a volume ratio of 1:4.

5. A method for preparing a ferrocenyl thiodihydropyrimidinone structured polymer according to claim 2, wherein: the post-treatment process comprises the following steps: the polymerization solution was cooled, diethyl ether was added dropwise with stirring until no more precipitate was formed, the mixture was dissolved in 5ml of THF, diethyl ether was added dropwise until no more precipitate was formed, and the mixture was filtered and dried for 24 hours to obtain a ferrocenyl thiodihydropyrimidinone-structured polymer.

6. A medicament for alleviating early brain injury after subarachnoid hemorrhage, which is characterized in that the composition of the medicament is the ferrocenyl thiodihydropyrimidinone structure polymer of claim 1.