Application of nuciferine in preparation of medicine for relieving pyroptosis of trophoblasts in preeclampsia

Abstract

The invention belongs to the technical field of biology, and particularly relates to application of nuciferine in preparation of a medicine for relieving pyroptosis of trophoblasts in preeclampsia. Nuciferine inhibits activation of NLRP3 inflammasomes and reduces caspase-1 cleavage and GSDMD mediated pyroptosis, so that placenta inflammation and dysfunction are relieved, and preeclampsia symptoms are improved. The nuciferine remarkably improves the symptoms of fetal growth limitation and placenta weight reduction of preeclampsia mice, and reduces placenta oxidative stress and inflammation level. In a cell model, nuciferine effectively reverses the pyroptosis phenotype of trophoblast cells, and the protein expression of NLRP3, cleaved-caspase-1, GSDMD-NT and IL-1beta is remarkably reduced. The application potential of nuciferine as a promising natural medicine in preeclampsia treatment is clarified, and the understanding of the relationship between inflammation and pyroptosis in preeclampsia pathogenesis is deepened.

Description

Technical Field

[0001] This invention belongs to the field of biotechnology, specifically relating to the application of lotus leaf alkaloids in the preparation of drugs to alleviate pyroptosis of trophoblast cells in preeclampsia. Background Technology

[0002] Preeclampsia (PE) is a multisystem dysfunction specific to pregnancy, characterized by hypertension and proteinuria after 20 weeks of gestation, and is one of the leading causes of maternal and perinatal morbidity and mortality worldwide. Placental dysfunction is considered the core of the pathophysiology of PE, with excessive inflammation and programmed cell death of placental trophoblasts playing a key role. Pyroptosis, a pro-inflammatory programmed cell death pathway mediated by Gasdermin family proteins, classically depends on the assembly of the NLRP3 inflammasome, activation of caspase-1, and release of mature inflammatory factors such as interleukin-1β and IL-18. Recent studies have shown significant pyroptosis in the placenta of patients with PE.

[0003] Lotus leaf alkaloid, a natural compound extracted from lotus leaves, is an apophene-type alkaloid and has been reported to possess various pharmacological activities, including anti-inflammatory and antioxidant effects. However, whether lotus leaf alkaloid plays a protective role in preeclampsia remains unclear. Summary of the Invention

[0004] To address the aforementioned technical challenges, this invention investigates the therapeutic effects of lotus leaf alkaloids by constructing an in vivo mouse model of preeclampsia and an in vitro trophoblast pyroptosis model, and further explores its target and signaling pathways. This invention is the first to discover that lotus leaf alkaloids can inhibit the activation of the NLRP3 inflammasome, thereby alleviating trophoblast pyroptosis and improving preeclampsia symptoms, providing a new candidate drug and theoretical target for the treatment of preeclampsia.

[0005] The first objective of this invention is to provide the use of lotus leaf alkaloids in the preparation of drugs to alleviate trophoblast pyroptosis in preeclampsia.

[0006] Furthermore, lotus leaf alkaloids inhibit NLRP3 inflammasome activation, reduce caspase-1 cleavage and GSDMD-mediated pyroptosis, thereby alleviating placental inflammation and dysfunction and improving preeclampsia symptoms.

[0007] Furthermore, lotus leaf alkaloids significantly improved the symptoms of fetal growth restriction and placental weight loss in preeclampsia mice, and reduced placental oxidative stress and inflammation levels.

[0008] Furthermore, in cell models, lotus leaf alkaloids effectively reversed the pyroptosis phenotype of trophoblast cells and significantly downregulated the protein expression of NLRP3, cleaved-caspase-1, GSDMD-NT, and IL-1β.

[0009] Furthermore, the administration strategy for the lotus leaf alkaloid is by gavage.

[0010] A second objective of this invention is to provide the use of lotus leaf alkaloids as a natural drug in the preparation of drugs for treating preeclampsia.

[0011] This invention establishes a lipopolysaccharide (LPS)-induced pyroptosis (PE) mouse model to observe the effects of lotus leaf alkaloids on fetal growth restriction, placental weight, oxidative stress, and inflammatory factor levels in mice. In vitro, a pyroptosis model is established by treating human trophoblast cell lines with LPS in combination with ATP. Cell morphology is observed using microscopy and transmission electron microscopy, and the expression of key pyroptosis proteins such as NLRP3, cleaved-caspase-1, GSDMD-NT, and IL-1β is detected by Western blotting.

[0012] The beneficial effects of this invention are as follows: First, this invention demonstrated in a mouse model of preeclampsia that lotus leaf alkaloids can improve core symptoms such as fetal growth restriction and placental dysfunction. This discovery links the anti-inflammatory and antioxidant properties of lotus leaf alkaloids with the clinical pathology of preeclampsia. Subsequently, the research focus was placed on the emerging mechanism of pyroptosis. In vitro data clearly showed that lotus leaf alkaloids can inhibit pyroptosis of trophoblast cells at the morphological and molecular levels, which provides a direct cellular basis for explaining its protective effect in vivo.

[0013] Secondly, this invention proposes a novel signaling pathway for the role of lotus leaf alkaloids in preeclampsia: lotus leaf alkaloids inhibit NLRP3 inflammasome activation, reduce caspase-1 cleavage, and decrease GSDMD-mediated pyroptosis. This alleviates placental inflammation and dysfunction, thereby improving preeclampsia symptoms.

[0014] Third, this invention not only elucidates the application potential of lotus leaf alkaloid as a promising natural drug in the treatment of preeclampsia, but also deepens the understanding of the relationship between inflammation and pyroptosis in the pathogenesis of preeclampsia.

[0015] Fourth, this invention provides a research foundation for further optimizing the administration strategy of lotus leaf alkaloids and exploring the possibility of its combined use with other drugs, so as to promote its transformation to clinical application. Attached Figure Description

[0016] Figure 1 A diagram illustrating how lotus leaf alkaloids improve symptoms in a mouse model of LPS-induced preeclampsia. Figure 1 Image A shows representative images of fetuses from the control group (n=3), the LPS group (n=3), and the LPS + lotus leaf alkaloid group (n=3). Figure 1Figure B shows the placental and fetal weight statistics for the control group (n=3), the LPS group (n=3), and the LPS + lotus leaf alkaloid group (n=3). Figure 1 In the middle C, there are statistical images of placental MDA, GSH, IL-1β, and IL-6 in different groups. Figure 1 Representative images of immunofluorescence staining of IL-1β and IL-6 in placenta from different groups in the middle D group. Data in the figure are expressed as mean ± standard deviation (SD). One-way ANOVA was performed first, followed by Tukey–Kramer multiple comparison test based on at least three independent experiments. *p<0.05, ***p<0.001, ****p<0.0001. NS: no statistical significance. Figure 2 The diagram illustrates the inhibition of LPS / ATP-induced pyroptosis in trophoblast cells by lotus leaf alkaloids. Figure 2 Image A shows representative light microscopic images of cells from the control group (n=3), LPS group (n=3), and LPS + lotus leaf alkaloid group (n=3). Figure 2 Image B shows representative electron micrographs of cells from different groups. Figure 2 The CD image shows representative immunoblot images and statistical graphs of NLRP3, caspase-1, GSDMD, and IL-1β in different groups of placenta. Data are expressed as mean ± standard deviation (SD), first analyzed by one-way ANOVA, followed by Tukey–Kramer multiple comparison tests based on at least three independent experiments. *p<0.05, ***p<0.001, ****p<0.0001. NS: No statistical significance. Detailed Implementation

[0017] Example 1 Lotus leaf alkaloids improve symptoms in a mouse model of LPS-induced preeclampsia. To evaluate the therapeutic effect of lotus leaf alkaloids on preeclampsia, this invention established an LPS-induced mouse model of preeclampsia.

[0018] On day 11 of pregnancy, all mice were injected intravenously with 1 μg / kg LPS solution via the tail vein to induce a preeclampsia model. On day 12, after measuring blood pressure, the mice were randomly divided into two groups: the LPS group and the LPS + lotus leaf alkaloid group (40 mg / kg / ig).

[0019] from Figure 1 Studies A and B concluded that, compared with the control group, the LPS model group mice exhibited typical preeclampsia symptoms, including fetal growth restriction and a significant decrease in placental weight.

[0020] from Figure 1 As shown in C and D, the levels of oxidative stress and the concentration of inflammatory factors in the placental tissue of the LPS model group were also significantly increased. However, after intervention with lotus leaf alkaloids during LPS modeling, the above pathological symptoms were significantly reversed: fetal weight and placental weight were restored, and oxidative stress and inflammatory response in placental tissue were significantly reduced.

[0021] These results indicate that lotus leaf alkaloids have the potential to improve the pathological process of preeclampsia in vivo.

[0022] Example 2 Lotus leaf alkaloids effectively inhibit the pyroptosis process of trophoblasts. In in vitro experiments, we successfully constructed a pyroptosis model by treating human trophoblast cell lines with LPS in combination with ATP.

[0023] Cells were treated with 200 ng / mL LPS for 24 hours, followed by the addition of 5 mmol ATP for 45 minutes.

[0024] Figure 2 Microscopic and transmission electron microscopy observations in A and B revealed that cells in the LSP / ATP model group exhibited typical pyroptosis morphological features, such as cell swelling, membrane pore formation and rupture, while lotus leaf alkaloid treatment significantly improved these cell morphologies.

[0025] Figure 2 Figures C and D show that, at the molecular level, Western blotting results indicate that LPS / ATP stimulation significantly upregulated the expression of key pyroptosis proteins NLRP3, cleaved-caspase-1, GSDMD-NT, and IL-1β. However, after intervention with lotus leaf alkaloids, the expression levels of these key proteins were significantly reversed, confirming that lotus leaf alkaloids can effectively inhibit the pyroptosis process in trophoblast cells.

Claims

1. Application of lotus leaf alkaloids in the preparation of drugs to alleviate pyroptosis of trophoblast cells in preeclampsia.

2. In the application described in claim 1, the lotus leaf alkaloid inhibits NLRP3 inflammasome activation, reduces caspase-1 cleavage and GSDMD-mediated pyroptosis, thereby alleviating placental inflammation and dysfunction and improving preeclampsia symptoms.

3. As described in claim 1, lotus leaf alkaloids improved the symptoms of fetal growth restriction and placental weight loss in preeclampsia mice, and reduced placental oxidative stress and inflammation levels.

4. In the application described in claim 1, lotus leaf alkaloid reversed the pyroptosis phenotype of trophoblast cells and downregulated the protein expression of NLRP3, cleaved-caspase-1, GSDMD-NT and IL-1β.

5. In the application as described in claim 1, the administration strategy of the lotus leaf alkaloid is by gavage.

6. The application of lotus leaf alkaloids as a natural drug in the preparation of drugs for the treatment of preeclampsia.

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