Application of IP receptor stimulant in preparation of medicine for treating lung injury

Abstract

The invention provides application of an IP receptor stimulant in preparation of a medicine for treating lung injury, and relates to the technical field of biological medicine. Transdifferentiation of alveolar II-type epithelial cells to I-type epithelial cells after lung injury is realized by utilizing selepag and other IP receptor agonists, and the scheme not only can relieve lung tissue inflammation and fibrosis degree induced by bleomycin and other factors, but also can reduce the death rate of model animals and improve lung function prognosis. Meanwhile, the used IP receptor stimulant part is a clinically mastered drug, so that the safety is higher, the selectivity is good, and a new effective way and a new drug development direction are provided for the treatment of lung injury and related fibrosis diseases.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, and in particular to the application of IP receptor agonists in the preparation of drugs for treating lung injury. Background Technology

[0002] Lung injury is a serious clinical condition caused by various factors, including infection, inhalation of harmful substances, and chest trauma. Its pathological process involves acute inflammatory response, disruption of the alveolar epithelial barrier, and subsequent fibrotic changes, which can lead to respiratory failure and even death in severe cases. Current clinical treatments for lung injury mainly include respiratory support, anti-inflammatory drugs, and symptomatic management; however, there is a lack of targeted drugs that can effectively promote alveolar structural regeneration and prevent or reverse fibrosis progression. Especially in the critical stage of lung injury repair—alveolar epithelial regeneration—the transdifferentiation of type II alveolar epithelial cells (AT2) into type I epithelial cells (AT1) is a core link in restoring alveolar structure and function. However, the molecular regulatory mechanisms of this transdifferentiation process are not yet fully elucidated, hindering the discovery of relevant repair targets and drug development.

[0003] Existing research indicates that AT2 cells possess stem cell characteristics, enabling them to proliferate and differentiate into AT1 cells after lung injury, thereby participating in the repair and reconstruction of alveolar epithelium. Blockage of this process can easily lead to abnormal repair, alveolar structural disorder, and pulmonary fibrosis. Although some signaling pathways (such as Notch, Wnt, and HIPpo) and transcription factors (such as p53 and FoxM1) have been reported to participate in the regulation of AT2 cell fate, their upstream regulatory mechanisms and key druggable targets remain unclear, limiting the development of targeted therapeutic strategies.

[0004] In terms of drug intervention, existing non-selective cyclooxygenase inhibitors may cause side effects in multiple systems such as the gastrointestinal tract and kidneys because they affect the synthesis of various prostaglandins, and may even aggravate the condition in certain infectious backgrounds. Therefore, they are not ideal treatment strategies for lung injury.

[0005] In summary, the field of lung injury repair currently lacks drug targets with clearly defined mechanisms that can specifically promote alveolar epithelial regeneration. Elucidating the regulatory network of AT2 cell transdifferentiation and identifying key receptors and downstream pathways that can be intervened are of great significance for the development of novel lung repair drugs.

[0006] Therefore, this invention is proposed. Summary of the Invention

[0007] To address the aforementioned technical problems, this invention provides the application of IP receptor agonists in the preparation of drugs for treating lung injury. This is significant for elucidating the regulatory network of AT2 cell transdifferentiation and identifying key modifiable receptors and downstream pathways.

[0008] In order to achieve the objective of this invention, the following technical solution is adopted: This invention provides the use of an IP receptor agonist in the preparation of a medicament for treating lung injury, wherein the IP receptor agonist is any one or more of selexipag, cicarboprost, iloprost, or beraprost.

[0009] Furthermore, the IP receptor agonist is selepag.

[0010] Furthermore, the effective dose of the IP receptor agonist is 1 mg / kg body weight / day to 10 mg / kg body weight / day.

[0011] Furthermore, the effective dose of the IP receptor agonist is 3 mg / kg body weight / day.

[0012] Furthermore, the lung injury includes: chemical lung injury, infectious lung injury, mechanical lung injury, radiation lung injury, or pulmonary fibrosis resulting from the above-mentioned injuries.

[0013] Furthermore, the chemical lung injury is pulmonary fibrosis induced by drugs or toxic substances.

[0014] Furthermore, the drug or toxic substance is bleomycin.

[0015] Furthermore, the drug is used to upregulate the expression of p53 in alveolar type II epithelial cells and promote the transdifferentiation of alveolar type II epithelial cells into alveolar type I epithelial cells.

[0016] The present invention also provides a pharmaceutical composition comprising: a first active ingredient and a second active ingredient; The first active ingredient includes: an IP receptor agonist and a pharmaceutically acceptable carrier or excipient; The second active ingredient includes any one or more of the following: MAP3K5 inhibitor, JNK inhibitor, JUN inhibitor, nintedanib, or pirfenidone.

[0017] Furthermore, the IP receptor agonist is any one or more of selepag, cicarboprost, iloprost, or beraprost.

[0018] Furthermore, the pharmaceutical composition comprises: First active ingredient: Selepag; The second active ingredient is either a JNK inhibitor or a JUN inhibitor.

[0019] The present invention has the following technical effects: This invention effectively overcomes the problem of the lack of specific targets for promoting alveolar epithelial regeneration in the prior art by providing the application of IP receptor agonists in the preparation of drugs for treating lung injury.

[0020] This technique utilizes IP receptor agonists such as selexipag and cicarboprost to significantly promote the transdifferentiation of type II alveolar epithelial cells (AT2) into type I epithelial cells (AT1) after lung injury, thereby accelerating alveolar structural repair and functional reconstruction. Its mechanism of action involves activating the IP receptor, which then inhibits the abnormal activation of the transcription factor JUN through the PKA / MAP3K5 / JNK signaling pathway, thereby relieving its inhibition of p53 expression and ultimately upregulating p53 levels and driving AT2 cells to differentiate into AT1 cells. Experiments show that this approach not only alleviates lung tissue inflammation and fibrosis induced by bleomycin and other factors, but also reduces mortality in model animals and improves lung function prognosis.

[0021] Compared with the limitations of existing antifibrotic drugs, which mainly delay the progression but cannot promote regeneration, this invention provides a treatment strategy that promotes active repair of alveolar epithelium from the source. Furthermore, the IP receptor agonist used is already a clinically approved drug with high safety and good selectivity, providing a new and effective approach and direction for drug development in the treatment of lung injury and related fibrotic diseases. Attached Figure Description

[0022] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0023] Figure 1 The results of suppressing IP receptors and reducing the proportion of AT2 cells differentiating into AT1 cells are presented as follows: A) Analysis of spatial transcriptomic data (S-BSST1410) from distal lung tissues of healthy controls and patients with idiopathic pulmonary fibrosis (IPF); B) Cell clustering UMAP diagram of spatial transcriptomic data from IPF and healthy control lung tissues; C) Proportional distribution of AT2, AT1, and transitional AT2 cells in healthy control and IPF samples; D) Proportional network of prostaglandin receptor-AT1 related gene interactions inferred in transitional AT2 cells through mutual information analysis; node size represents the number of AT1 genes linked to prostaglandin receptors; E) In situ expression signals of PTGIS and IP on IPF lung sections; the upper figure shows PTGIS expression in neighboring cells, and the lower figure shows the AUCell score of the AT1 gene set in transitional AT2 cells; FG) IP + With IP -Correlation analysis of AUCell score of AT1 gene set in transitional AT2 cells with PTGIS expression in neighboring cells; H represents the effect of nine PG receptor inhibitors on AT2 cell differentiation (detected by RT-qPCR). Pdpn mRNA (AT1) and Sftpc mRNA (AT2) levels, the ratio of which represents the differentiation rate of AT2 to AT1; EP1 receptor inhibitor: SC-51322, EP2 receptor inhibitor: PF-04418948, EP3 receptor inhibitor: L-798,106, EP4 receptor inhibitor: L-161,982, DP1 receptor inhibitor: BWA868C; DP2 receptor inhibitor: CAY10595, IP receptor inhibitor: Cay10441, TP receptor inhibitor: SQ29548, FP receptor inhibitor: AL8810, n=5-6; I is for immunofluorescence staining detection of WT and IP receptor systemic knockout mice (IP - / - Differentiation of AT2 cells in organoids; AT2 (Pro-SPC), AT1 (PDPN), cell nucleus (DAPI); scale bar is 20 μm; J is the PDPN / pro-SPC staining area ratio obtained according to I, a quantitative analysis of AT2 to AT1 transdifferentiation (n = 23); K. RT-qPCR detection of the proportion of AT2 to AT1 cell differentiation, n=5, *** indicates p<0.001; Figure 2 : Results of constructing AT2 cell-mediated IP receptor knockout mice, where A represents Sftpc -CreER; Rosa26 -mTmG; IP F / F A) Mouse construction diagram; B) DNA electrophoresis diagram for mouse gene identification; C) Knockout efficiency under RT-qPCR detection conditions, n=4, P<0.05, * indicates p<0.05; Figure 3 The results of AT2 conditional knockout of the IP receptor exacerbating bleomycin-induced lung injury in mice are shown in the following figures: A shows the lung injury model induced by tamoxifen-induced conditional knockout of IP receptor AT2 cells and nebulized administration of bleomycin; B shows the survival rate curve of mice after lung injury, n=21-22; C shows a representative image of HE staining of mouse lung tissue, with the scale bar of the lung lobe at 1 mm and the scale bar of the magnified field of view at 20 μm; D shows a representative image of Masson staining of mouse lung tissue; E shows the collagen area statistics based on Masson staining, n=3-8; F shows the Ashcroft score based on Masson staining, n=3-8. * indicates p<0.05, *** indicates p<0.001. Figure 4Conditional knockout of the IP receptor in AT2 exacerbates bleomycin-induced pulmonary fibrosis in mice; A shows the hydroxyproline content in mouse lung tissue (n=3-8); B shows the protein expression levels of α-SMA and Vimentin in mouse lung tissue; *** indicates p<0.001. Figure 5 : Results of decreased AT2 cell differentiation capacity in AT2 conditional knockout IP mice after bleomycin-induced lung injury; A: Immunofluorescence showing morphological changes of AT2 cells differentiating into AT1 cells after bleomycin injury; GFP: AT2 cells, PDPN: AT1 cells, DAPI: cell nucleus; arrows point to GFP and PDPN double-positive cells; scale bar is 10 μm; B: Statistical representation of the proportion of GFP+PDPN+ cells to total GFP+ cells, n=3-8; *** indicates p<0.001; Figure 6 The receptor agonist Selexipag alleviated bleomycin-induced lung injury in mice. A is a flowchart of Selexipag treatment for mouse lung injury; mice were administered bleomycin via tracheal nebulization two weeks after tamoxifen induction, followed by Selexipag 8 days later, continuing until day 18 post-surgery, with samples collected 21 days later; B is the mouse survival rate curve (n=22-25); C is a representative image of HE staining in mouse lung tissue, with a scale bar of 1 mm for lung lobes and 20 μm for magnification; D is a representative image of Masson staining in mouse lung tissue; E is the collagen area statistics based on Masson staining (n=8); F is the Ashcroft score based on Masson staining (n=8). * indicates p < 0.05, *** indicates p < 0.001. Figure 7 The IP receptor agonist Selexipag alleviated bleomycin-induced pulmonary fibrosis in mice. A represents the hydroxyproline content in mouse lung tissue (n=8); B represents the protein expression levels of α-SMA and Vimentin in mouse lung tissue as determined by Western blot. ** indicates p < 0.01. Figure 8 Results of the IP receptor agonist Selexipag promoting the transdifferentiation of AT2 cells to AT1 cells after lung injury. A represents the immunofluorescence staining of GFP (lineage-labeled AT2) and PDPN (AT1) in mouse lung tissue; arrows point to GFP and PDPN double-positive cells; DAPI (nucleus); scale bar is 10 μm; B represents GFP. + PDPN + Cellular percentage of total GFP + The cell proportion statistics, n=8, *** indicates p<0.001; Figure 9The study investigated the inhibitory effect of Jun siRNA knockdown on AT2 transdifferentiation. A represents the effect of immunofluorescence on the AT2-to-AT1 differentiation of organoids by the JUN inhibitor T5224, with AT2 (Pro-SPC), AT1 (PDPN), and cell nucleus (DAPI); scale bar is 20 μm. B represents the effect of RT-qPCR on the AT2-to-AT1 differentiation ratio in organoids; n=5-6. C represents the validation of Jun siRNA knockdown efficiency; n=5-6. D represents the effect of RT-qPCR on the AT2-to-AT1 differentiation ratio in organoids; n=6. ** indicates p < 0.01, *** indicates p < 0.001.

[0024] Figure 10 The study investigated the effect of Jun knockdown on the aggravation of bleomycin-induced lung injury in mice by inhibiting IP deficiency in AT2 cells. A represents the tamoxifen-induced conditional knockout and nebulized bleomycin-induced lung injury models in mice. B shows the survival rate curves of mice after lung injury, n=24-25. C is a representative image of HE staining in mouse lung tissue, with a scale bar of 1 mm for lung lobes and 20 μm for magnification. D is a representative image of Masson staining in mouse lung tissue. E shows the collagen area statistics based on Masson staining, n=8. F shows the Ashcroft score based on Masson staining, n=8. * indicates p<0.05, ** indicates p<0.01, *** indicates p<0.001. Detailed Implementation

[0025] To make the objectives, technical solutions, and advantages of this invention clearer, the technical solutions of this invention will be clearly and completely described below. Obviously, the described embodiments are only a part of the embodiments of this invention, and not all of them. Based on the embodiments of this invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this invention.

[0026] In a first aspect, the present invention provides the use of an IP receptor agonist in the preparation of a medicament for treating lung injury, wherein the IP receptor agonist is any one or more of selexipag, cicarboprost, iloprost, or beraprost.

[0027] In some embodiments, the IP receptor agonist is selepag.

[0028] In some embodiments, the effective amount of the IP receptor agonist is 1 mg / kg body weight / day to 10 mg / kg body weight / day.

[0029] In some embodiments, the effective amount of the IP receptor agonist is 3 mg / kg body weight / day.

[0030] In some embodiments, the lung injury includes: chemical lung injury, infectious lung injury, mechanical lung injury, radiation lung injury, or pulmonary fibrosis resulting from the above-mentioned injuries.

[0031] In some embodiments, the chemical lung injury is drug- or toxic induced pulmonary fibrosis.

[0032] In some embodiments, the drug or toxic substance is bleomycin.

[0033] In some embodiments, the drug is used to upregulate the expression of p53 in alveolar type II epithelial cells, thereby promoting the transdifferentiation of alveolar type II epithelial cells into alveolar type I epithelial cells.

[0034] In a second aspect, this application provides a pharmaceutical composition comprising: a first active ingredient and a second active ingredient; The first active ingredient includes: an IP receptor agonist and a pharmaceutically acceptable carrier or excipient; The second active ingredient includes any one or more of the following: MAP3K5 inhibitor, JNK inhibitor, JUN inhibitor, nintedanib, or pirfenidone.

[0035] In some embodiments, the IP receptor agonist is any one or more of selepag, cicarboprost, iloprost, or beraprost.

[0036] In some embodiments, the pharmaceutical composition comprises: First active ingredient: Selepag; The second active ingredient is either a JNK inhibitor or a JUN inhibitor.

[0037] The following is a detailed explanation using specific embodiments: Example 1: Inhibition of IP receptor weakens the transdifferentiation ability of AT2 cells to AT1 cells To investigate the role of prostaglandin receptors in AT2 cell differentiation, we first analyzed publicly available spatial transcriptome data including samples from patients with idiopathic pulmonary fibrosis and healthy controls. Figure 1 (As shown in A). Based on marker genes, cells were divided into AT1 cells, AT2 cells, and transitional AT2 cells (as shown in A). Figure 1 (As shown in B). Compared with healthy controls, the proportion of transitional AT2 cells in IPF lung tissue was significantly increased ( Figure 1(As shown in C). To assess whether prostaglandin receptor-mediated signaling regulates AT1 gene expression, we used mutual information methods to specifically evaluate the association between prostaglandin receptors and the AT1 gene in transitional AT2 cells. Among all prostaglandin receptors, the prostacyclin receptor showed the strongest association with AT1 gene expression, suggesting it may act as a key regulator of AT2 to AT1 transdifferentiation. Figure 1 (As shown in D in the figure). We further used the AUCell scoring method to quantify the in situ enrichment signal of the AT1 gene in transitional AT2 cells within fibrotic regions of different severities, and assessed the gene expression level of prostacyclin synthase (PTGIS) in neighboring cells within a 200 μm radius around each transitional AT2 cell. The results showed that higher PTGIS expression in neighboring cells was associated with higher AT1 gene scores and milder fibrotic lesions in transitional AT2 cells, while lower PTGIS gene expression corresponded to lower AT1 gene scores and more severe fibrosis (as shown in D in the figure). Figure 1 (As shown in E in the figure). To further assess whether higher local PGI2 availability in the microenvironment promotes AT2 to AT1 transdifferentiation via IP signaling, we performed Spearman rank correlation analysis and found that PTGIS gene expression in neighboring cells was positively correlated with AT1 gene enrichment in IP-positive (but not IP-negative) transitional AT2 cells. Figure 1 (As shown in F and G in the figure), this indicates that the PGI2 / IP axis plays a specific role in promoting the transdifferentiation of AT2 to AT1. Further flow cytometry sorting of wild-type mouse (WT) AT2 cells (CD31-, CD45-, CD34-, EPCAM+, Sca1-, CD24-) and MRC5 human lung fibroblasts were used for in vitro co-culture of alveolar organoids, using the AT1 marker gene. Pdpn mRNA levels and AT2 cell marker genes Sftpc The ratio of mRNA levels indicates the transdifferentiation capacity of AT2 cells. The results showed that the IP receptor inhibitor Cay10441 significantly inhibited the transdifferentiation of AT2 cells into AT1 cells. Figure 1 (H in the figure). Further, AT2 cells were isolated from WT mice and IP receptor systemic knockout mice (IP KO). Immunofluorescence and RT-qPCR results after organoid co-culture showed that IP knockout significantly reduced the expression of AT1 marker genes within the organoids, and significantly decreased the transdifferentiation ability of AT2 cells to AT1 cells. Figure 1 (As shown in I, J, and K).

[0038] Example 2: Conditional knockout of IP receptor AT2 exacerbates bleomycin-induced lung injury in mice use Sftpc -CreER; Rosa26 -mTmG mice and IPF / F Fluorescent tracer mice with conditional knockout of IP receptor in AT2 cells were constructed. Sftpc -CreER; Rosa 26-mTmG; IP F / F ), Sftpc Specifically expressed in AT2 cells, CreER is formed by the fusion of the recombinase cre and the estrogen receptor ER. Rosa26 -mTmG / + This is a red-green fluorescent reporter system. In uninduced mice, the cell membrane expresses red fluorescent protein. Tamoxifen induction activates AT2 cells, causing them to switch from expressing red fluorescent protein to expressing green fluorescent protein. Simultaneously, it specifically knocks out the IP receptor in AT2 cells. Figure 2 (A and B in the text). Two weeks after tamoxifen injection, samples were separated by flow cytometry. Sftpc -CreER; Rosa26 -mTmG mice and Sftpc -CreER; Rosa26 -mTmG; IP F / F The mRNA level of the IP receptor in GFP+ AT2 cells from mouse lung tissue was detected by RT-qPCR, and the results showed... Sftpc -CreER; Rosa26 -mTmG; IP F / F The IP receptor knockdown efficiency in AT2 cells of mice reached approximately 80%. Figure 2 (C in the middle).

[0039] For 8-week-old Sftpc -CreER; Rosa26 -mTmG mice and Sftpc -CreER; Rosa26 -mTmG; IP F / F Mice were induced with tamoxifen, and two weeks later were given PBS or bleomycin via tracheal nebulization. Lung tissue samples were collected 21 days later. Figure 3 (A) Compared with the control group, the mortality rate of Sftpc-CreER;Rosa26-mTmG;IPF / F mice was higher ( Figure 3 (B in the text). HE staining results showed that after administration of PBS, the control group mice and... Sftpc -CreER; Rosa26 -mTmG; IP F / F The alveolar structure of the mouse lung tissue was intact, with no inflammatory cell infiltration observed. The alveolar septa were relatively thin, and there was no significant difference in morphology between the two groups of lung tissue. Figure 3 (C and D in the text); after intratracheal administration of bleomycin, compared with control mice, Sftpc -CreER; Rosa26- mTmG; IP F / F The lung tissue structure of mice was severely damaged, the alveolar septa were significantly thickened, and the infiltration of inflammatory cells was increased. Figure 3 (C). Masson staining results showed that, compared with control mice, after administration of bleomycin... Sftpc -CreER; Rosa26 -mTmG; IP F / F An increase in the area of ​​blue-stained regions in the lungs of mice indicates increased deposition of fibrous collagen. Figure 3 The results of the Ashcroft score showed that the degree of fibrosis in the lung tissue of Sftpc-CreER;Rosa26-mTmG;IPF / F mice was significantly more severe than that in the control group after administration of bleomycin (D and E). Figure 3 (F in the text).

[0040] The results of hydroxyproline detection showed that after administration of PBS, the control group mice and Sftpc -CreER; Rosa26 -mTmG; IP F / F There was no significant difference in hydroxyproline content in mouse lung tissue; however, after administration of bleomycin... Sftpc -CreER; Rosa26 -mTmG; IP F / F The content of hydroxyproline in the lung tissue of mice was significantly increased compared with that in the control group. Figure 4 (A) Western blot analysis of lung fibrosis markers revealed that, compared to the control group, after bleomycin administration... Sftpc -CreER; Rosa26 -mTmG; IP F / F The expression of fibrosis markers α-SMA and Vimentin in mouse lung tissue was significantly increased. Figure 4 (B in the text). The above data indicate that AT2 conditional knockout of IP exacerbates bleomycin-induced lung injury and subsequent fibrosis in mice.

[0041] Morphological changes in AT2 cells from the lung tissue of traced mice were observed. Immunofluorescence staining showed that GFP-labeled AT2 cells were cuboidal in the resting state, and no significant morphological changes were observed after IP knockout. After administration of bleomycin, some green AT2 cells became elongated and simultaneously showed positivity for the AT1 cell marker PDPN, indicating that they had transdifferentiated into AT1 cells. Compared with the control group, Sftpc -CreER; Rosa26 -mTmG; IP F / F Long strips of GFP in mouse lung tissue + PDPN + The proportion of cells decreased significantly. Figure 5 The results (A and B in the text) indicate that knocking out IP prevents the transdifferentiation of AT2 cells to AT1 cells during bleomycin-induced lung injury, thereby leading to severe fibrosis.

[0042] Example 3: The IP receptor agonist selexipag significantly alleviated bleomycin-induced lung injury in mice. The absence of IP receptors in AT2 cells inhibits AT2 cells from differentiating into AT1 cells, thus exacerbating bleomycin-induced lung injury in mice. However, whether activation of IP receptors has a protective effect against bleomycin-induced lung injury requires further investigation. Selexipag, a stable and potent IP receptor agonist, was approved by the FDA in 2015. Selexipag is characterized by activating only IP receptors without binding to other PG receptors, demonstrating good efficacy in treating pulmonary hypertension. To further investigate the role of Selexipag in lung injury repair, an 8-week-old study was conducted... Sftpc -CreER; Rosa26 -mTmG and Sftpc -CreER; Rosa26 -mTmG; IP F / F Mice were induced with tamoxifen, and two weeks later, bleomycin was administered via intratracheal nebulization to induce lung injury. On day 8 post-modeling, [the mice were then...]. Sftpc -CreER; Rosa26 -mTmG and Sftpc -CreER; Rosa26 -mTmG; IP F / F Mice were randomly divided into a corn oil group and a group given the selective IP receptor agonist selepag. Figure 6 (A) Survival curve results showed that, compared with the corn oil group, Seluxipag treatment resulted in... Sftpc -CreER; Rosa26 The survival rate of -mTmG mice was significantly improved, but for Sftpc -CreER; Rosa26 -mTmG; IP F / F The survival rate of mice did not improve significantly. Figure 6 (B in the text). HE staining results showed that, compared with the corn oil group, Seluxipag treatment... Sftpc -CreER; Rosa26 -mTmG mice showed reduced inflammatory cell infiltration and less alveolar structural damage in their lung tissue, while Sftpc -CreER; Rosa26 -mTmG; IP F / F The degree of inflammatory cell infiltration and alveolar structural damage in mouse lung tissue was not significantly altered. Figure 6 (C). Masson staining results showed that, compared with the corn oil group, Selexipag treatment... Sftpc -CreER; Rosa26 The blue-stained areas in the lung tissue of -mTmG mice were significantly reduced, and fibrotic collagen deposition was significantly reduced. Sftpc -CreER; Rosa26 -mTmG; IP F / F No significant changes were observed in fibrotic collagen deposition in mouse lung tissue. Figure 6 (D, E in the text). Ashcroft scoring results showed that Selexipag treatment alleviated... Sftpc -CreER; Rosa26 The degree of fibrosis in the lung tissue of -mTmG mice, but for Sftpc -CreER; Rosa26 -mTmG; IP F / F The degree of fibrosis in mouse lung tissue was not significantly affected. Figure 6 (F in the text). The above results indicate that Selexipag treatment can alleviate bleomycin-induced lung injury in mice, but cannot alleviate bleomycin-induced... Sftpc -CreER; Rosa26 -mTmG; IP F / F Lung injury in mice indicates that the anti-injury effect of Selexipag is achieved by promoting the transdifferentiation of AT2 cells.

[0043] Example 4: The receptor agonist Selexipag alleviates bleomycin-induced pulmonary fibrosis in mice. Furthermore, the hydroxyproline detection results showed that, compared with the corn oil group, Seluxipag treatment resulted in lower levels of hydroxyproline. Sftpc -CreER; Rosa26 The hydroxyproline content in the lung tissue of -mTmG mice was significantly reduced. Figure 7 (A) Western blot analysis of fibrosis markers revealed that, compared to the corn oil group, Seluxipag treatment resulted in... Sftpc -CreER; Rosa26 The expression of fibrosis markers α-SMA and Vimentin in the lung tissue of -mTmG mice was significantly reduced. Figure 7 (B in the text), but Selexipag is correct. Sftpc -CreER; Rosa26 -mTmG; IP F / F The degree of fibrosis in mouse lung tissue was not significantly improved. These results indicate that Selexipag treatment alleviates bleomycin-induced pulmonary fibrosis in mice by promoting AT2 cell transdifferentiation.

[0044] Finally, the differentiation of AT2 cells in mice treated with the IP receptor agonist Selexipag was examined. Immunofluorescence staining revealed that, compared with the corn oil group, the differentiation of AT2 cells in mice treated with Selexipag was significantly lower. Sftpc -CreER; Rosa26 GFP in -mTmG mice + PDPN + The number of cells increased significantly. Figure 8 (A, B), but Selexipag cannot be restored. Sftpc -CreER; Rosa26 -mTmG; IP F / F The differentiation rate of mouse AT2 cells into AT1 cells.

[0045] Experiment Example 5: Verification of JUN Functionality Flow cytometry sorting of WT mice and IP mice - / - Mouse AT2 cells were used for in vitro organoid culture. Immunofluorescence and RT-qPCR results showed that treatment with the JUN inhibitor T5224 promoted the transdifferentiation of AT2 cells into AT1 cells and reversed the inhibitory effect of IP knockout on AT2 transdifferentiation (e.g., Figure 9 (A and B in the original text). Knockdown of Jun using siRNA yielded results consistent with those of JUN inhibitors.

[0046] Example 6: Knockout of Jun alleviates the aggravating effect of IP loss in AT2 cells on lung injury in mice. To further verify whether the AT2 cell IP receptor participates in the lung injury repair process through JUN, we constructed AT2 cell conditionally knocked-out IP receptor and Jun mice ( Sftpc -CreER; Rosa26 -mTmG; IP F / F ; Jun F / F ) and AT2 cell conditional knockout Jun mice ( Sftpc -CreER; Rosa26 -mTmG; Jun F / F Two weeks after tamoxifen injection, bleomycin was administered via nebulization to the lungs of mice to induce lung injury. Figure 10 A in the text). Compared with control mice ( Sftpc -CreER; Rosa26 Compared to bleomycin-treated AT2 cell IP receptor-deficient mice (-mTmG), bleomycin treatment for 50 years resulted in the reduction of IP receptor-deficient mice in AT2 cells. Sftpc -CreER; Rosa26 -mTmG; IP F / F The mortality rate of AT2 cell Jun-deficient mice was significantly increased, while that of AT2 cell Jun-deficient mice was significantly increased. Sftpc -CreER; Rosa26 -mTmG; Jun F / F The mortality rate was significantly reduced, and knocking out Jun significantly improved the mortality rate in AT2 cell IP receptor-deficient mice. Sftpc -CreER; Rosa26 -mTmG; IP F / F Survival rate () Figure 10 (B in the text). HE and Masson staining showed that, compared with the control group, AT2 cell IP receptor-deficient mice had more severe lung tissue structural damage, significantly thickened alveolar septa, and increased inflammatory cell infiltration and fibrosis area; while AT2 cell Jun-deficient mice showed significantly reduced lung tissue structural damage, decreased inflammatory cell infiltration and fibrosis area, and Jun knockout significantly alleviated the lung injury and increased fibrosis caused by AT2 cell IP receptor deficiency in mice. Figure 10 (CF in the middle).

[0047] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the technical solutions of the embodiments of the present invention.

Claims

1. The application of IP receptor agonists in the preparation of drugs for treating lung injury, characterized in that, The IP receptor agonist is any one or more of selepag, cicarboprost, iloprost, or beraprost.

2. The use of the IP receptor agonist according to claim 1 in the preparation of a medicament for treating lung injury, characterized in that, The IP receptor agonist is selepag.

3. The use of the IP receptor agonist according to claim 1 in the preparation of a medicament for treating lung injury, characterized in that, The effective dose of the IP receptor agonist is 1 mg / kg body weight / day to 10 mg / kg body weight / day.

4. The use of the IP receptor agonist according to claim 1 in the preparation of a medicament for treating lung injury, characterized in that, The lung injury includes: chemical lung injury, infectious lung injury, mechanical lung injury, radiation lung injury, or pulmonary fibrosis resulting from the above injuries.

5. The use of the IP receptor agonist according to claim 1 in the preparation of a medicament for treating lung injury, characterized in that, The drug is used to upregulate the expression of p53 in alveolar type II epithelial cells and promote the transdifferentiation of alveolar type II epithelial cells into alveolar type I epithelial cells.

6. A pharmaceutical composition for treating lung injury, characterized in that, The pharmaceutical composition comprises: a first active ingredient and a second active ingredient; The first active ingredient includes: an IP receptor agonist and a pharmaceutically acceptable carrier or excipient; The second active ingredient includes any one or more of the following: MAP3K5 inhibitor, JNK inhibitor, JUN inhibitor, nintedanib, or pirfenidone.

7. The pharmaceutical composition according to claim 6, characterized in that, The IP receptor agonist is any one or more of selepag, cicarboprost, iloprost, or beraprost.

8. The pharmaceutical composition according to any one of claims 6-7, characterized in that, The pharmaceutical composition comprises: First active ingredient: Selepag; The second active ingredient is either a JNK inhibitor or a JUN inhibitor.

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