Nanometer material for treating endometriosis and application thereof

Abstract

The invention discloses a nanometer material for treating endometriosis and application thereof, and belongs to the technical field of biological medicine and nanometer medicine delivery. The nano material is ACS14 (at) BSA (bovine serum albumin) nano particles; the ACS14-coated BSA nanoparticles take BSA as a carrier, and are loaded with hydrogen sulfide release type aspirin ACS14. According to the present invention, the BSA is adopted as the carrier, the selective enrichment of the lesion can be achieved through the neutrophil carrying mechanism, the ACS14 can simultaneously provide proliferation inhibition and anti-inflammatory effects at the lesion so as to reduce the recurrence risk, and the hydrogen sulfide released at the lesion can act on the anterior clasp cortex through blood circulation, can up-regulate the GLT-1, can reduce the extracellular glutamic acid, and can inhibit the neuronal over-excitation; the related anxiety is relieved. According to the invention, focus targeted delivery, proliferation inhibition and anti-inflammatory treatment can be realized, related anxiety can be improved synchronously, and comprehensive treatment of endometriosis and complications such as related anxiety can be realized.

Description

Technical Field

[0001] This invention relates to the fields of biomedicine and nanomedicine delivery technology, and in particular to a nanomaterial for treating endometriosis and its application. Background Technology

[0002] Endometriosis (EMs) is a common chronic inflammatory gynecological disease, mainly characterized by the ectopic growth of endometrial-like tissue. It can cause dysmenorrhea, pelvic pain, and infertility, severely reducing patients' quality of life. The occurrence and development of endometriosis are closely related to abnormal proliferation of ectopic cells, persistent activation of inflammatory responses, and dysregulation of the immune microenvironment.

[0003] Currently, the main treatments for endometriosis are surgery and hormone therapy. While surgical removal can reduce the lesion burden in the short term, it has a high recurrence rate and may even affect fertility. Hormone therapy can inhibit lesion progression, but recurrence is very common after discontinuation, and it may also cause side effects such as weight gain and mood swings. Traditional anti-inflammatory and analgesic drugs mostly provide symptomatic relief and are difficult to control lesions at the level of proliferation and inflammation. Therefore, there is an urgent need for a new, non-surgical, non-hormonal treatment method for endometriosis that can fundamentally control lesions and reduce side effects.

[0004] In addition, endometriosis is often accompanied by psychological comorbidities such as anxiety. The incidence of anxiety and depression in patients is significantly higher than in healthy people. However, existing anti-anxiety drugs mainly target mental symptoms and provide symptomatic treatment, which is difficult to control the lesions. Moreover, they may cause adverse reactions, such as drowsiness and dependence, and cannot achieve comprehensive management that treats both the mind and body.

[0005] Hydrogen sulfide, a gaseous signaling molecule, possesses certain anti-inflammatory and neuromodulation potential. However, its rapid release and metabolism under physiological conditions lead to uncontrollable release and poor targeted distribution in clinical applications, severely limiting its clinical use. Nanotechnology offers a new solution for drug delivery, but most existing nanodelivery systems emphasize local targeting or single-drug efficacy, making it difficult to simultaneously achieve unified improvement of different symptoms.

[0006] Therefore, in order to simultaneously address the issues of lesion treatment and anxiety relief for patients with endometriosis, it is of great significance to develop a new treatment method for endometriosis with a low recurrence rate and few side effects. Summary of the Invention

[0007] This invention provides a nanomaterial for treating endometriosis and its application, which can solve the problems of high recurrence rate, large side effects, and difficulty in simultaneously treating lesions and alleviating anxiety in the existing technology for treating endometriosis.

[0008] In a first aspect, the present invention provides a nanomaterial for treating endometriosis, the nanomaterial being ACS14@BSA nanoparticles; the ACS14@BSA nanoparticles use BSA as a carrier and are loaded with hydrogen sulfide-releasing aspirin ACS14.

[0009] Preferably, the raw materials for ACS14@BSA nanoparticles include BSA and hydrogen sulfide-releasing aspirin ACS14 in a mass ratio of 10:(2.5-3.5).

[0010] Preferably, the loading rate of hydrogen sulfide-releasing aspirin ACS14 is 8.9–11.5%.

[0011] Preferably, the particle size of ACS14@BSA nanoparticles is 200–250 nm.

[0012] Preferably, ACS14@BSA nanoparticles are prepared according to the following method: S1. Add BSA to deionized water and stir to dissolve at room temperature to obtain an aqueous BSA solution; add ACS14 to dimethyl sulfoxide and stir to dissolve to obtain an ACS14 solution; S2. Add ACS14 solution to BSA aqueous solution and stir at room temperature to obtain a mixed solution; S3. Add anhydrous ethanol dropwise to the mixed solution and stir until homogeneous; then add a crosslinking agent solution with a mass fraction of 2-3% and stir for 5-6 hours to obtain the reaction solution; S4. Centrifuge the reaction solution at 4-5℃ for 10-15 min, collect the precipitate, resuspend it in ultrapure water, vortex mix it, and then centrifuge and wash it 2-3 times; finally, add it to the buffer solution for redispersion to obtain the final product.

[0013] Preferably, the crosslinking agent includes glutaraldehyde.

[0014] Preferably, the mass-to-volume ratio of BSA, anhydrous ethanol, and crosslinking agent solution is 10 mg: (3-3.5) mL: (8-15) μL.

[0015] More preferably, the buffer solution includes PBS buffer.

[0016] By adopting the above technical solution, the nanomaterial provided by this invention for treating endometriosis is an ACS14@BSA nanoparticle. The ACS14@BSA nanoparticle uses BSA (bovine serum albumin) as a carrier and is loaded with a novel hydrogen sulfide-releasing aspirin, ACS14. Hydrogen sulfide-releasing aspirin ACS14 is an aspirin derivative loaded with hydrogen sulfide and capable of releasing hydrogen sulfide. It uses aspirin as the parent drug and is linked with a hydrogen sulfide-releasing group, namely a dithiol thione. Its chemical structural formula is shown below:

[0017] In the preparation of ACS14@BSA nanoparticles, BSA aqueous solution and ACS14 solution are mixed to achieve initial dispersion of ACS14 on the BSA support. Then, anhydrous ethanol is added for alcohol precipitation, which reduces the hydration layer around BSA molecules, increases the hydrophobic region, and enhances the binding between BSA and ACS14 molecules. Through strong hydrophobic interactions, hydrogen bonds, and intermolecular forces, ACS14 gradually and spontaneously assembles into nanoscale particles, effectively encapsulating ACS14 inside the particles formed by the hydrophobic core of BSA. Then, a crosslinking agent is added, which can react with the groups on the surface of BSA molecules to further form a dense covalent crosslinking network, thereby improving the stability and continuous release characteristics of the nanoparticles. Finally, after centrifugation, washing, and redispersion, the ACS14@BSA nanoparticle dispersion is obtained.

[0018] ACS14@BSA nanoparticles use bovine serum albumin as a carrier material and can achieve targeted delivery in vivo through a neutrophil loading mechanism. This allows the nanoparticles to selectively accumulate at ectopic lesions in endometriosis, improving the targeting of the nanomaterials.

[0019] The addition of ACS14 can inhibit ectopic cell proliferation by regulating the PI3K / Akt signaling pathway, and reduce local inflammatory response by inhibiting the NF-κB pathway. Therefore, it can control lesions from two different mechanism levels, namely proliferation and inflammation, to precisely and efficiently inhibit lesion progression and greatly reduce the risk of recurrence.

[0020] Meanwhile, the hydrogen sulfide released by ACS14 at the lesion site can enter the bloodstream and be delivered to the anterior cingulate cortex, which is closely related to anxiety regulation, upregulate the expression of glutamate transporter 1 (GLT-1), reduce extracellular glutamate levels, and inhibit abnormal overexcitation of glutamate neurons, thereby improving anxiety symptoms associated with endometriosis.

[0021] Furthermore, the ACS14@BSA nanoparticles of this invention have a nanometer-scale particle size, exhibiting excellent stability and sustained release characteristics. The obtained ACS14@BSA nanoparticles can not only effectively inhibit the proliferation of endometriosis lesions, reduce the possibility of symptom recurrence and side effects, but also alleviate abnormal neuronal activity and improve related anxiety symptoms, thus providing comprehensive treatment for endometriosis and its related complications such as anxiety.

[0022] Secondly, the present invention provides an application of a nanomaterial for treating endometriosis, based on the above-mentioned nanomaterial for treating endometriosis, which is used to treat endometriosis or to prepare a drug for treating endometriosis.

[0023] Preferably, the nanomaterials are used to treat psychological comorbidities such as anxiety caused by endometriosis or to prepare drugs for treating psychological comorbidities such as anxiety caused by endometriosis.

[0024] Preferably, the nanomaterials are used to simultaneously treat endometriosis and psychological comorbidities such as anxiety caused by endometriosis, or to prepare drugs for simultaneously treating endometriosis and psychological comorbidities such as anxiety caused by endometriosis.

[0025] The beneficial effects of this invention are: 1. The nanomaterials for treating endometriosis provided by this invention can combine lesion-targeted delivery, inhibition of proliferation and anti-inflammatory treatment. Furthermore, through the "lesion-brain axis" mechanism, it can simultaneously improve related anxiety. It is a non-hormonal nanomedicine system that can solve the problem of difficulty in simultaneously controlling lesion progression and psychological comorbidity, and achieve comprehensive treatment of endometriosis and its related complications such as anxiety.

[0026] 2. The nanomaterial provided by this invention for treating endometriosis is ACS14@BSA nanoparticles. Using BSA as a carrier, ACS14@BSA nanoparticles can achieve selective enrichment of lesions through a neutrophil loading mechanism, improving effective exposure. ACS14 can simultaneously exert inhibitory proliferation and anti-inflammatory effects at the lesion site, reducing the risk of recurrence. Furthermore, through the "lesion-brain axis" linkage, hydrogen sulfide released from the lesion can act on the anterior cingulate cortex via blood circulation, upregulating GLT-1, reducing extracellular glutamate, and inhibiting neuronal overexcitation, mechanistically alleviating related anxiety and reducing drug side effects. The nanomaterials have good stability and sustained release characteristics, which are beneficial for maintaining therapeutic efficacy. Attached Figure Description

[0027] Figure 1 This invention provides a schematic diagram of the synthesis of nanomaterials for treating endometriosis and a schematic diagram of the mechanism of action for treating endometriosis in Example 1. Figure 2 , Figure 3 and Figure 4 A schematic diagram showing the physicochemical characterization results of ACS14@BSA nanoparticles in Example 1 of this invention; Figure 5-1 and Figure 5-2 This is a schematic diagram of the targeted delivery capability evaluation results of Embodiment 2 of the present invention; Figure 6-1 , Figure 6-2 , Figure 6-3 , Figure 7-1 , Figure 7-2 and Figure 8 This is a schematic diagram illustrating the verification results of the volume assessment and control mechanism of lesion proliferation and inflammation in Embodiment 3 of the present invention; Figure 9 and Figure 10 This is a schematic diagram illustrating the verification results of the psychological comorbidity mechanism of anxiety and other psychological disorders in Embodiment 3 of the present invention. Figure 11 and Figure 12 This is a schematic diagram illustrating the verification results of the mechanism regulating the excitability of glutamate neurons in the ACC region during the treatment of endometriosis in Example 4 of the present invention. Figure 13 , Figure 14 and Figure 15 This is a schematic diagram illustrating the verification results of the improvement of anxiety-like behaviors in endometriosis in Embodiment 5 of the present invention. Figure 16 and Figure 17 This is a schematic diagram of the safety evaluation results of Embodiment 6 of the present invention. Detailed Implementation

[0028] The specific embodiments of the present invention will be described in detail below, but it should be understood that the scope of protection of the present invention is not limited to the specific embodiments.

[0029] Example 1: A nanomaterial for treating endometriosis was prepared according to the following method: S1. Weigh 10 mg of BSA and add it to 1 mL of deionized water and stir at room temperature to dissolve it to obtain a BSA aqueous solution; Weigh 3.0 mg of ACS14 (average drug loading rate of 10.5%) and add it to 100 μL of dimethyl sulfoxide and stir to dissolve to obtain an ACS14 solution; S2. Add ACS14 solution to BSA aqueous solution and stir at 600 rpm for 1 h at room temperature to obtain a mixed solution; S3. Add 3 mL of anhydrous ethanol to the mixed solution and stir until homogeneous; then add 10 μL of 2.5% glutaraldehyde solution and stir at 600 rpm for 6 h at room temperature to obtain the reaction solution; S4. Centrifuge the reaction solution at 4℃ and 12000rpm for 10min, discard the supernatant, collect the precipitate, resuspend it in ultrapure water, vortex mix, and then centrifuge and wash three times to remove residual organic solvents and free substances; finally, add PBS buffer to 1mL and redisperse to obtain the final product. The average particle size of the obtained ACS14@BSA nanoparticles is 220nm.

[0030] The physicochemical properties of the obtained ACS14@BSA nanoparticles were tested: Transmission electron microscopy (TEM) images of the ACS14@BSA nanoparticles are shown below. Figure 2 As shown; Fourier transform infrared spectrum as Figure 3 As shown; the in vitro cumulative release curves of hydrogen sulfide and aspirin in PBS buffer at pH 5.5 over 48 hours are shown in the figure. Figure 4 As shown.

[0031] Example 2: Evaluation of the targeted delivery capability of a nanomaterial for treating endometriosis: In vivo distribution assays were conducted, with separate experimental and control groups. The experimental group used ACS14@BSA nanoparticles prepared in Example 1 labeled with FITC (fluorescein isothiocyanate), denoted as FITC-ACS14@BSA; the control group used free FITC dye. During the experiment, the fluorescence intensity of the fluorescence signal at different locations in vivo was measured 16 hours after intraperitoneal injection, as detailed below. Figure 5-1 , Figure 5-2 As shown.

[0032] from Figure 5-1 and Figure 5-2 As can be seen, FITC-ACS14@BSA exhibited significantly stronger fluorescence signals in endometriosis lesions compared to other organs, while there was no significant difference in fluorescence between FITC-ACS14@BSA and the control group of free FITC in major organs. This indicates that ACS14@BSA nanoparticles have excellent targeted delivery properties to endometriosis lesions in vivo.

[0033] Example 3: Animal experiment to verify the therapeutic mechanism of a nanomaterial for treating endometriosis: Establishment of an endometriosis model: Six- to eight-week-old female Balb / c mice were selected as test subjects and divided into donor and recipient groups. Donor mice received a single subcutaneous injection of estradiol benzoate (3 μg per mouse). One week after injection, the donor mice were sacrificed, and the uterine horns were harvested. The uterine horns were then longitudinally cut open using surgical scissors and fragmented into small tissue pieces. The tissue fragments obtained from the uterine horns of three donor mice were then intraperitoneally injected into four recipient mice, thereby establishing an endometriosis model.

[0034] Experimental Procedure: Mice with endometriosis were randomly divided into four groups for comparing the efficacy of different formulations. The different formulations selected were PBS buffer, BSA-NPs (bovine serum albumin nanoparticles), free ACS14, and ACS14@BSA nanoparticles prepared in Example 1, respectively. Each group received an intraperitoneal injection of 100 μL of the formulation every other day. Mouse weight was recorded every other day during the treatment period, and the results are as follows: Figure 6-1 As shown; after 3 weeks of continuous drug administration, the animals were sacrificed and samples were collected. The ectopic lesions were excised for volume assessment and subsequent histological examination. The excised ectopic lesion samples are shown below. Figure 6-2 As shown.

[0035] Test and verification detection: 1. Volume Assessment: The length and width of the lesions were measured using calipers; the severity of the disease was assessed by the total volume of all lesions in each mouse; the volume of a single lesion was calculated using the following formula: Lesion volume V = 0.5 × length × width 2 The volume assessment results are as follows: Figure 6-3 As shown, from Figure 6-1 , Figure 6-2 and Figure 6-3 As can be seen, the ACS14@BSA nanoparticles provided by this invention can significantly reduce the volume of ectopic lesions after continuous administration. Furthermore, there was no significant change in body weight in mice administered with different dosage forms during the experimental treatment period.

[0036] 2. Verification of the mechanism for controlling lesion proliferation and inflammation: (1) mRNA sequencing and pathway analysis: Total RNA was extracted from ectopic lesion tissue prepared with ACS14@BSA nanoparticles and PBS buffer, respectively. The purity and concentration were detected using NanoDrop 2000, and the integrity was assessed using Agilent 2100. A library was constructed using the VAHTS Universal V6 kit. 150 bp paired-end sequencing was performed using Illumina NovaSeq 6000. The genome was aligned to a reference genome and the expression level was calculated. HTSeq-count was used to obtain the count. PCA was performed using R software to assess sample consistency. Differentially expressed genes were screened using DESeq2, and hierarchical clustering and KEGG enrichment analysis were performed on the differentially expressed genes. The results of the above experiments are as follows: Figure 7-1 and Figure 7-2 As shown in the figure, differentially expressed genes are significantly enriched in pathways such as PI3K / Akt and NF-κB. (2) Histological and mechanism verification: The ectopic lesions of mice treated with different preparations in the above four groups were embedded in paraffin, sectioned after embedding, and then Ki67 immunohistochemistry and IL-6 immunofluorescence staining were performed. Then, scanning was performed using a Zeiss LSM880 confocal microscope and an Olympus IX71 fluorescence microscope. The results are as follows: Figure 8 As shown, where Figure 8 A and Figure 8 C represents the fluorescence scan result. Figure 8 B and Figure 8 D represents the fluorescence signal intensity results of different immunohistochemical staining methods.

[0037] from Figure 8 As can be seen, the ectopic lesion tissue treated with the ACS14@BSA nanoparticle formulation showed a reduction in Ki67-positive cells and a significant decrease in the pro-inflammatory factor IL-6, indicating that the ACS14@BSA nanoparticles provided by this invention mainly exert their therapeutic effect by inhibiting PI3K / Akt-related proliferation and NF-κB-related inflammation.

[0038] 3. Verification of comorbid psychological mechanisms such as anxiety: (1) Blood-mediated delivery verification: The above-mentioned endometriosis model mice were divided into two groups. The experimental group was injected intraperitoneally with the ACS14@BSA nanoparticle dispersion prepared in Example 1, while the control group was injected intramuscularly with PBS buffer. Then, the Washington State Probe-1 (WSP-1, hydrogen sulfide probe) was used to image hydrogen sulfide in the ectopic lesions and the in situ endometrium. The hydrogen sulfide signal showed a bright green fluorescence. The detection results and signal intensity were compared as follows: Figure 9 As shown.

[0039] from Figure 9 As can be seen, mice treated with the ACS14@BSA nanoparticles of this invention exhibited strong fluorescence in ectopic lesions, while the endometrium of mice in the control group treated with PBS buffer showed only a very weak signal. This indicates that hydrogen sulfide is specifically released in ectopic lesions. The intraperitoneally administered ACS14@BSA nanoparticles can achieve targeted delivery to endometriotic lesions and can release hydrogen sulfide locally.

[0040] (2) Stereotyping of the brain and fixation of the drug delivery cannula: After anesthetizing the endometriosis model mice by intraperitoneal injection of sodium pentobarbital (20 mg / kg), they were fixed in a stereotyping instrument (RWD Life Science). The core body temperature was maintained at about 36°C throughout the process using a heating pad. The skull was exposed and leveled by cutting along the midline of the mouse scalp. A dental drill was used to drill holes in the ACC region, i.e., the anterior cingulate cortex: with the anterior fontanelle (bregma) as a reference, the anterior-posterior (AP) diameter was +1.09 mm, the lateral (ML) diameter was ±0.30 mm, and the dorsoventral (DV) diameter was -1.75 mm.

[0041] Next, the drug delivery cannula was vertically inserted above the injection site, and an optical fiber was inserted at an angle below the cannula to monitor the fluorescence signal. Super-Bond C&B dental acrylic resin was used for fixation. After ≥14 days of postoperative recovery, the hydrogen sulfide fluorescent probe WSP-1 (1 mM) was injected into the ACC through the microinjection cannula, with the tip of the injection cannula extending 0.5 mm beyond the guide cannula. Subsequently, a fiber optic photometry system was used to record real-time changes in hydrogen sulfide fluorescence within the ACC to assess the dynamic level of hydrogen sulfide in the ACC after drug delivery.

[0042] Hydroxycobalamin was pre-injected via the tail vein before drug administration. This substance is a hydrogen sulfide scavenger that does not easily cross the blood-brain barrier and can be used to remove circulating hydrogen sulfide. After pretreatment, the ACS14@BSA nanoparticle dispersion prepared in Example 1 was administered, and changes in serum hydrogen sulfide and fluorescence signals within the ACC were detected, as detailed below. Figure 10 As shown.

[0043] from Figure 10 As can be seen, hydroxycobalamin can eliminate the increase in serum hydrogen sulfide caused by ACS14@BSA nanoparticle dispersion and prevent the hydrogen sulfide signal in the ACC from rising again, thus indicating that hydrogen sulfide is transported from the lesion to the ACC via blood circulation.

[0044] Example 4: Verification of the mechanism by which a nanomaterial for treating endometriosis regulates the excitability of glutamate neurons in the ACC region during the treatment of endometriosis: 1. Stereotactic localization and viral injection: Mice with the endometriosis model from Example 3 were anesthetized by intraperitoneal injection of sodium pentobarbital (20 mg / kg) and fixed in a stereotactic unit (RWD Life Science). A heating pad was used throughout to maintain the core body temperature at approximately 36°C. The skull was exposed and leveled by incision along the midline of the scalp. A hole was drilled in the ACC region using a dental drill: using the anterior fontanelle as a reference, the anterior-posterior (AP) diameter was +1.09 mm, the lateral (ML) diameter was ±0.30 mm, and the dorsoventral (DV) diameter was -1.75 mm. AAV2 / 9-CaMKIIα-GCaMP6m was injected into the ACC using a microinjection pump (RWD Life Science) connected to a pull-type glass micropipette. Approximately 200–300 nL of viral solution was delivered at a rate of 50 nL / min, depending on the viral titer and expression efficiency. After injection, the microtube was left in place for several minutes to reduce reflux.

[0045] Immediately after viral injection, an optical fiber was vertically inserted above the injection site and fixed using Super-Bond C&B dental acrylic resin. Fiber photometry was recorded starting three weeks later.

[0046] 2. Short-term treatment: The mice treated as described above were randomly divided into four groups: normal group, PBS group (mice with endometriosis were given PBS buffer), ASA@BSA group (mice with endometriosis were given aspirin-loaded BSA nanoparticles), and ACS14@BSA group (mice with endometriosis were given the ACS14@BSA nanoparticles obtained in Example 1).

[0047] 3. Fiber Optic Recording and Result Analysis: A fiber optic photometric record was performed before short-term treatment administration as a baseline value; after administration, a second record was made under the same conditions, and the changes in ΔF / F and the area under the curve were compared between the two records. Figure 11 A and Figure 11 As shown in C; and the event amplitude evoked by the stimulus as shown in Figure C. Figure 11 As shown in B.

[0048] from Figure 11 As can be seen, compared with the PBS group, the ACS14@BSA group, i.e., the group treated with the ACS14@BSA nanoparticles of this invention, can significantly reduce Glu. ACC The enhanced calcium activity induced by neuronal stimulation indicates that the ACS14@BSA nanoparticles provided in this invention can inhibit endometriosis-related Glu. ACC Over-excitement.

[0049] 4. Glutamate content and GLT-1 detection: ACC tissues from the normal group, PBS group, ASA@BSA group, and ACS14@BSA group were collected, and glutamate levels were quantitatively detected by HPLC. The results are as follows: Figure 12 As shown in Figure A; simultaneously, GLT-1 protein expression was detected by Western blot, and the results are as follows. Figure 12 B and Figure 12 As shown in C.

[0050] from Figure 12 The results show that ACC glutamate levels were elevated in the PBS and ASA@BSA groups, while ACC glutamate concentration was decreased and approached normal in the ACS14@BSA group. Furthermore, GLT-1 expression within the ACC was upregulated in the ACS14@BSA group, indicating that the ACS14@BSA nanoparticles of this invention may alleviate Glutamate deficiency by enhancing extracellular glutamate uptake and maintaining glutamate homeostasis. ACC Neurons are abnormally highly active.

[0051] Example 5: Validation of the effect of a nanomaterial for treating endometriosis on improving anxiety-like behaviors associated with endometriosis: 1. Short-term treatment: After living normally for 3 weeks, the mice with endometriosis model in Example 3 were randomly divided into three groups: PBS group, ASA@BSA group and ACS14@BSA group. They were treated with PBS buffer, ASA@BSA nanoparticles and ACS14@BSA nanoparticles prepared in Example 1 of this invention, respectively, and the treatment was carried out for three consecutive days. 2. Behavioral tests: A control group was added, consisting of normal mice. Behavioral tests were conducted on both groups. (1) Elevated Cross Maze (EPM): Mice were placed in the experimental environment for acclimatization 1 hour before the test. The EPM device consists of two open arms, two closed arms, and a central platform, and is used to assess anxiety-like behaviors. Mice were placed on the central platform facing one open arm and allowed to explore freely for 5 minutes. The device was cleaned with 70% ethanol after each test. The number of times mice entered the open arms and the dwell time in the open arms were recorded and analyzed using Any-Maze software. The results are as follows: Figure 13 A, Figure 13 B and Figure 13 As shown in C; (2) Open Field Test (OFT): After EPM, OFT was performed to assess anxiety-like behavior and spontaneous activity. The setup was a 50 cm × 50 cm × 40 cm open field. Mice were placed in the central area and allowed to explore freely for 5 minutes. The setup was cleaned after the test. Any-Maze software was used to record total walking distance, number of times the mice entered the central area, and time spent in the central area. The results are as follows: Figure 13 D、 Figure 13 E and Figure 13 As shown in F.

[0052] From the above experiments and Figure 13 As can be seen, compared with the normal group, the PBS group showed a significant reduction in the proportion of time spent in the open arm of the EPM and the number of times the mice entered the open arm. In the OFT, the number of times the mice entered the central region and the time spent in the central region were also reduced, indicating that mice with endometriosis exhibit significant anxiety-like behaviors. The ACS14@BSA group, treated with the ACS14@BSA nanoparticles of this invention, showed a significant improvement in the above behavioral indicators, while the ASA@BSA group showed no significant improvement, indicating that the anti-anxiety effect mainly comes from the hydrogen sulfide released by the ACS14@BSA nanoparticles.

[0053] 3. Serum corticosterone (ELISA): Mice were anesthetized after the behavioral test, and blood was collected from the inner canthus vein. The blood samples were centrifuged at 12,000 rpm for 20 min to separate the serum, which was then stored at -80℃. Serum corticosterone levels were detected using an ELISA kit (CSB-E07969m, CUSABIO). The results are as follows: Figure 14As shown in the figure, the corticosterone level in mice with endometriosis was significantly increased in the PBS group, while it was significantly decreased in the ACS14@BSA group; no significant decrease was observed in the corticosterone level in the ASA@BSA group, indicating that ACS14@BSA nanoparticles have a significant effect on reducing corticosterone levels.

[0054] 4. Lesion exclusion verification: To verify whether the anti-anxiety effect is independent of lesion progression, the size of ectopic lesions was compared between mice in the PBS group and the ACS14@BSA group during short-term treatment. The results are as follows: Figure 15 As shown in the figure, no change in lesion size was observed after three consecutive days of administration, indicating that the anti-anxiety effect exhibited by ACS14@BSA nanoparticles was not due to a reduction in the volume of ectopic lesions.

[0055] Example 6: Safety evaluation of a nanomaterial for treating endometriosis: The endometriosis model mice from Example 3 were randomly divided into four groups and administered the drugs in the following formulations: PBS buffer, BSA-NPs (bovine serum albumin nanoparticles), free ACS14, and ACS14@BSA nanoparticles prepared in Example 1. These were designated as the PBS group, BSA group, ACS14 group, and ACS14@BSA group, respectively. The drugs were administered three times a week for a total of nine weeks. Blood routine and blood biochemistry tests were performed after administration, and major organs were stained with hematoxylin and eosin (HE). The results are as follows: Figure 16 and Figure 17 As shown in the figure, the mice treated with the ACS14@BSA nanoparticles of this invention showed no significant toxic side effects.

[0056] The above-disclosed embodiments are merely a few specific examples of the present invention. However, the embodiments of the present invention are not limited thereto, and any variations that can be conceived by those skilled in the art should fall within the protection scope of the present invention.

Claims

1. A nanomaterial for treating endometriosis, characterized in that, The nanomaterial is ACS14@BSA nanoparticles; the ACS14@BSA nanoparticles use BSA as a carrier and are loaded with hydrogen sulfide-releasing aspirin ACS14.

2. The nanomaterial for treating endometriosis according to claim 1, characterized in that, The raw materials for the ACS14@BSA nanoparticles include BSA and hydrogen sulfide-releasing aspirin ACS14 in a mass ratio of 10:(2.5-3.5).

3. The nanomaterial for treating endometriosis according to claim 1, characterized in that, The drug loading rate of the hydrogen sulfide-releasing aspirin ACS14 was 8.9–11.5%.

4. The nanomaterial for treating endometriosis according to claim 1, characterized in that, The particle size of the ACS14@BSA nanoparticles is 200–250 nm.

5. The nanomaterial for treating endometriosis according to claim 1, characterized in that, The ACS14@BSA nanoparticles were prepared according to the following method: S1. Add BSA to deionized water and stir to dissolve at room temperature to obtain an aqueous BSA solution; add ACS14 to dimethyl sulfoxide and stir to dissolve to obtain an ACS14 solution; S2. Add ACS14 solution to BSA aqueous solution and stir at room temperature to obtain a mixed solution; S3. Add anhydrous ethanol dropwise to the mixed solution and stir until homogeneous; then add a crosslinking agent solution with a mass fraction of 2-3% and stir for 5-6 hours to obtain the reaction solution; S4. Centrifuge the reaction solution at 4-5℃ for 10-15 min, collect the precipitate, resuspend it in ultrapure water, vortex mix it, and then centrifuge and wash it 2-3 times; finally, add it to the buffer solution for redispersion to obtain the final product.

6. The nanomaterial for treating endometriosis according to claim 5, characterized in that, The crosslinking agent includes glutaraldehyde.

7. The nanomaterial for treating endometriosis according to claim 5, characterized in that, The mass-to-volume ratio of BSA, anhydrous ethanol, and crosslinking agent solution is 10 mg: (3–3.5) mL: (8–15) μL.

8. The application of a nanomaterial for treating endometriosis, based on the nanomaterial for treating endometriosis according to any one of claims 1 to 7, characterized in that, The nanomaterials are used to treat endometriosis or to prepare drugs for treating endometriosis.

9. The application of the nanomaterial for treating endometriosis according to claim 8, characterized in that, The nanomaterials are used to treat psychological comorbidities such as anxiety caused by endometriosis or to prepare drugs for treating psychological comorbidities such as anxiety caused by endometriosis.

10. The application of the nanomaterial for treating endometriosis according to claim 8, characterized in that, The nanomaterials are used to simultaneously treat endometriosis and psychological comorbidities such as anxiety caused by endometriosis, or to prepare drugs for simultaneously treating endometriosis and psychological comorbidities such as anxiety caused by endometriosis.

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