Use of sitosterol in the preparation of a medicament for preventing and treating necrotizing enterocolitis in newborns

Abstract

The application discloses application of sitosterol in preparation of a medicine for preventing and treating necrotizing enterocolitis of a newborn, and belongs to the technical field of biological medicine. The technical scheme uses sitosterol as an active ingredient to play a role through an enteral administration route in view of the problems of hidden onset and lack of effective prevention and treatment means of NEC. Experiments in vitro and in vivo prove that sitosterol can significantly inhibit intestinal inflammatory response, down-regulate expression of proinflammatory factors, and effectively block polarization of macrophages to M1 type, thereby improving pathological damage of the intestine. The application provides a new medicine selection for clinical prevention and treatment of NEC, and has important application value.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, and in particular relates to the application of sitosterol in the preparation of drugs for the prevention and treatment of neonatal necrotizing enterocolitis. Background Technology

[0002] Necrotizing enterocolitis (NEC) in newborns is one of the most fatal gastrointestinal emergencies in the neonatal intensive care unit (NICU), posing a serious threat to the lives and health of premature infants. The disease is characterized by its insidious onset and rapid progression, presenting significant challenges to early clinical diagnosis and timely intervention.

[0003] Early clinical manifestations of neonatal sepsis (NEC), such as feeding intolerance, gastric retention, and apnea, lack specificity. These symptoms are highly similar to those of other common critical illnesses like neonatal sepsis, easily leading to clinical confusion and delaying crucial intervention. By the time typical symptoms like abdominal distension and bloody stools appear, the condition has often progressed to a severe stage such as intestinal perforation. Data shows that the intestinal perforation rate in premature infants with NEC can be as high as 30%, drastically increasing the difficulty and risk of treatment.

[0004] Currently, clinical prevention and treatment measures for NEC are very limited. Once the condition progresses to the point where surgical intervention is required, surgery becomes the primary treatment. However, surgical treatment is not only highly invasive, but its prognosis is often unsatisfactory. Many children face a series of serious long-term sequelae after surgery, such as short bowel syndrome and neurodevelopmental disorders. These complications will severely affect the child's growth and development and long-term quality of life, placing a heavy burden on families and society.

[0005] Even more serious is the fact that, to date, the medical community lacks effective preventative and specific treatments for NEC. Faced with this disease with an extremely high mortality rate, clinicians are often in a reactive position. Therefore, there is an urgent need to explore and develop safe and effective new prevention and treatment strategies to fill the current gaps in clinical treatment, improve the prognosis of children with NEC, and reduce their mortality and disability rates. This is precisely the key scientific problem and technical challenge that urgently needs to be solved in this field. Summary of the Invention

[0006] To address the aforementioned technical problems, this invention proposes the application of gramustol in the preparation of drugs for the prevention and treatment of neonatal necrotizing enterocolitis.

[0007] To achieve the above objectives, the present invention provides the following technical solution: The use of gram-sitosterol in the preparation of drugs for the prevention and treatment of necrotizing enterocolitis in newborns, wherein the structural formula of the gram-sitosterol is as follows: .

[0008] Optionally, the drug may be administered via enteral administration.

[0009] Optionally, the dose of serotonin equivalent to a single administration of the drug is 20 mg / kg.

[0010] Optionally, the dosage form of the drug is selected from oral liquid, suspension, granules or capsules.

[0011] Optionally, the drug may also contain pharmaceutically acceptable excipients.

[0012] Optionally, the drug is used to reduce the concentration of [unclear - possibly referring to a specific substance or ingredient] in the ileum tissue of neonatal necrotizing enterocolitis model animals. Il-1β , Il-6 or Tnf-α The level of expression.

[0013] A drug for the prevention and treatment of neonatal necrotizing enterocolitis, with graminol as the active ingredient.

[0014] Compared with the prior art, the present invention has the following advantages and technical effects: This invention demonstrates, through in vitro cell experiments and in vivo animal experiments, that scotosterol has significant technical effects in preventing and treating neonatal necrotizing enterocolitis (NEC).

[0015] At the cellular level, this invention was validated using an LPS-induced mouse primary bone marrow macrophage (BMDM) inflammation model. Experimental results showed that glutathione significantly inhibited pro-inflammatory factors (such as...). Il-1β , Il-6 , Tnf-α ) and M1 macrophage markers (such as cd86 , inos The expression of ) indicates that glutathione can effectively inhibit macrophage polarization towards the pro-inflammatory M1 phenotype, thereby exerting an anti-inflammatory effect at the cellular level.

[0016] At the animal level, this invention further validated its efficacy by establishing a neonatal rat NEC model. Experiments revealed that after intervention with glutathione, the expression levels of pro-inflammatory factors in the ileum tissue of NEC model rats were significantly reduced, and the proportion of M1 macrophages in the intestinal lamina propria was also significantly decreased. Histopathological evaluation (H&E staining and NEC score) confirmed that glutathione can significantly improve NEC-induced intestinal tissue damage.

[0017] In summary, this invention achieves the technical effect of effectively preventing and treating neonatal necrotizing enterocolitis by inhibiting intestinal inflammatory response and M1 polarization of macrophages. Attached Figure Description

[0018] The accompanying drawings, which form part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an undue limitation of the invention. In the drawings: Figure 1 The results are from the experiment of daily gavage administration of gram-sitosterol in a neonatal rat NEC model. Figure 2 To investigate the effects of sitosterol on an LPS-induced mouse primary bone marrow macrophage inflammation model in in vitro cell experiments; Figure 3 To investigate the multifaceted effects of graminosterol on intestinal tissue in a neonatal rat NEC model. Detailed Implementation

[0019] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0020] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Every smaller range between any stated value or intermediate value within a stated range, and any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0021] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0022] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0023] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0024] Unless otherwise specified, the term "parts" in this invention refers to parts by weight.

[0025] All raw materials used in this invention were purchased commercially. Gramisterol, a sterol compound derived from almond oil, was purchased from MCE.

[0026] The technical solution of the present invention will be further illustrated by the following embodiments.

[0027] Example 1 The cells used in the experiments of this invention were primary mouse bone marrow macrophages (BMDM). The conditions for establishing the NEC cell model were as follows: 20 ng / mL M-CSF induced M0 macrophages, followed by LPS (10 ng / mL) induction of the NEC inflammation model. The cells were incubated at 37°C with simultaneous LPS and Gramisterol (16 μM) for 24 h. Grouping: CTL group (blank control group), LPS group, LPS + Gramisterol group, with all other culture conditions being the same. Inflammatory factors in BMDM were detected by qPCR. Il-6 , Il-1β , Tnf-α ) and macrophage M1 markers ( cd86 , inos The level of expression of ).

[0028] Example 2 The animals used in the experiments of this invention were newborn SD rat pups. The conditions for establishing the NEC animal model were as follows: Newborn SD rat pups were fed hypertonic formula (15 g Similac 60 / 40 (Ross Pediatrics, Columbus, Ohio) added to 75 mL Esbilac (Pet-Ag, New Hampshire, IL), providing 836.8 kJ / kg daily. Initially, 0.1 mL was administered by gavage each time, increasing by 0.1 mL daily, gradually increasing to a maximum of 0.4 mL per feeding. The model was then subjected to 10 min of 5% O2 hypoxia followed by 10 min of cold stimulation at 4℃ for 3 days. Gramisterol (20 mg / kg) was administered by gavage once daily. The rats were divided into three groups: CTL group, NEC group, and NEC + Gramisterol group, with all other conditions being the same.

[0029] Ileal tissue was collected, and the level of intestinal damage was assessed using an HE assay; inflammatory factors were detected using qPCR. Il-6 , Il-1 β , Tnf-α The expression level of CD86 was measured; the proportion of CD86-labeled macrophages was detected by flow cytometry.

[0030] In summary, this invention first induced mouse bone marrow macrophages (BMDM) to differentiate into unstimulated M0 type using M-CSF in Example 1, and then used LPS (lipopolysaccharide) stimulation to construct an NEC inflammatory cell model. Based on this, by setting up a control group (CTL), a model group (LPS), and a drug intervention group (LPS + Gramisterol), it was observed and verified whether 16 μM gram-sitosterol could effectively inhibit macrophage polarization towards the pro-inflammatory M1 type and downregulate related inflammatory factors ( ) within 24 hours. Il-6 , Il-1β , Tnf-α ) and M1 type marker ( cd86 , inos The expression of ) was then verified in Example 2, which examined the therapeutic effect of glutathione in a live animal setting. Specifically, a neonatal SD rat NEC model was successfully established using the classic combined intervention method of "hyperosmolar formula feeding + hypoxia + cold stimulation." During the 3-day modeling process, the model rats were administered glutathione at a dose of 20 mg / kg daily via gavage. The efficacy was evaluated using three detection methods at the end of the experiment: HE staining: a direct assessment of the degree of pathological damage to intestinal tissues; qPCR experiment: Detection of pro-inflammatory factors in ileal tissue ( Il-6, Il-1β, Tnf-α The level of expression; Flow cytometry: Quantitatively detects the proportion of CD86-positive (i.e. M1 type) macrophages in intestinal tissue.

[0031] Specific effects are as follows Figures 1-3 As shown: 1. Inhibits NEC intestinal inflammatory response Figure 1 This study presents the results of daily gramicular administration of gramisterol in a neonatal rat NEC model; specifically, neonatal rats were administered gramisterol once daily during the NEC model establishment period. N=6. A portion of the ileum was collected for H&E staining and NEC score assessment; the other portion was used for qPCR experiments. The results showed that gramisterol could inhibit the expression of pro-inflammatory intestinal factors and significantly improve intestinal damage in NEC.

[0032] 2. Suppress BMDM M1 type polarization Figure 2 To investigate the effects of gramisterol on an LPS-induced mouse primary bone marrow macrophage (BMDM) inflammation model in in vitro cell experiments, BMDM cells were pretreated with a specified concentration of gramisterol for 1 hour, followed by LPS stimulation for 6 hours. Subsequently, inflammatory factors were assessed using qPCR. Il-1β , Il-6 and Tnf-α ) and macrophage M1 markers ( cd86 and inos The results showed that Gramisterol significantly inhibited the expression of macrophage pro-inflammatory factors and M1 markers.

[0033] 3. Inhibits M1 polarization of intestinal macrophages Figure 3 To investigate the multifaceted effects of gramisterol on intestinal tissue in a neonatal rat NEC model, newborn rats were administered Gramisterol once daily by gavage during the NEC model establishment period. Mononuclear cells from the lamina propria of the ileum were collected and analyzed by flow cytometry. Results showed that Gramisterol significantly inhibited M1 polarization of macrophages.

[0034] In summary, in cell experiments, Gramisterol significantly reduced the expression of pro-inflammatory factors and M1 markers in BMDM; in animal experiments, Gramisterol significantly reduced pro-inflammatory factors in ileal tissue, inhibited macrophage M1 polarization, and improved ileal tissue damage.

[0035] The above are merely preferred embodiments of the present invention, but the scope of protection of the present invention is not limited thereto. Any variations or substitutions that can be easily conceived by those skilled in the art within the scope of the technology disclosed in the present invention should be included within the scope of protection of the present invention.

Claims

1. The application of phytosterol in the preparation of drugs for the prevention and treatment of neonatal necrotizing enterocolitis, characterized in that, The structural formula of the graminol is as follows: 。 2. The use of sitosterol according to claim 1 in the preparation of a medicament for the prevention and treatment of neonatal necrotizing enterocolitis, characterized in that, The drug is administered via enteral administration.

3. The use of sitosterol according to claim 1 in the preparation of a medicament for the prevention and treatment of neonatal necrotizing enterocolitis, characterized in that, The dose of serotonin equivalent to a single dose of the drug is 20 mg / kg.

4. The use of sitosterol according to claim 1 in the preparation of a medicament for the prevention and treatment of neonatal necrotizing enterocolitis, characterized in that, The dosage form of the drug is selected from oral liquid, suspension, granules or capsules.

5. The use of sitosterol according to claim 4 in the preparation of a medicament for the prevention and treatment of neonatal necrotizing enterocolitis, characterized in that, The drug also contains pharmaceutically acceptable excipients.

6. The use of sitosterol according to claim 1 in the preparation of a medicament for the prevention and treatment of neonatal necrotizing enterocolitis, characterized in that, The drug is used to reduce the levels of [unclear - possibly referring to a specific substance or ingredient] in the ileum tissue of neonatal necrotizing enterocolitis model animals. Il- 1β , Il-6 or Tnf-α The level of expression.

7. A drug for the prevention and treatment of necrotizing enterocolitis in newborns, characterized in that, The active ingredient is strychnol as described in claim 1.

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