Epithelial dedifferentiation and myofibroblast activation are critical drivers of chronic kidney disease (CKD) progression. Elevated levels of IGFBP6 have been linked to decreased renal function in CKD patients, but its precise role and underlying mechanisms remain unclear. In this study, we observed significantly increased IGFBP6 expression in the kidney tissues of both renal fibrosis patients and animal models. Global or tubule-specific IGFBP6 knockout attenuated renal cellular senescence and fibrosis development in mice. In vitro, IGFBP6 deficiency preserved epithelial cell phenotype and inhibited fibroblast activation. Additionally, anti-IGFBP6 treatment demonstrated promising therapeutic effects in alleviating renal cellular senescence and fibrosis. Mechanistically, IGFBP6, acting as an adaptor protein, could bind to thrombospondin 1 (THBS1) and prevent its ubiquitination-mediated degradation, thereby activating the THBS1-CD47 cellular pathway in epithelial cells, which contributed to renal cellular senescence and fibrosis. Notably, both genetic and neutralizing antibody-mediated inhibition of IGFBP6 alleviated renal cellular senescence and fibrosis, suggesting that the IGFBP6/THBS1/CD47 axis represents a potential therapeutic target for chronic kidney injury.

Acute lung injury (ALI), characterized by dysregulated inflammation and oxidative stress, remains a major therapeutic challenge due to limited treatment options. Inspired by the anti-inflammatory activity of the in-house synthetic intermediate A1, we designed and synthesized a series of chalcone derivatives through structural optimization, preserving the phenylthioether moiety while modifying the aromatic ring on one side of the chalcone olefin. Among these, compound B15 emerged as the lead candidate, exhibiting potent nitric oxide (NO) inhibition (IC50 = 2.6 ± 1.4 μM) in LPS-stimulated RAW264.7 macrophages with minimal cytotoxicity. Mechanistic studies demonstrated that B15 concentration-dependently inhibited MAPK phosphorylation (p38, JNK, and ERK), suppressed transcription levels of TNF-α and IL-6, and reduced ROS production in vitro, via direct binding to p38 MAPK. In an LPS-induced murine model of ALI, B15 significantly attenuated pathological damage and inflammatory response. Collectively, these finding establish B15 as a promising anti-inflammatory lead for ALI therapy via p38 MAPK inhibition, meriting further pharmacological development.

Cyclin-dependent kinase 6 (CDK6) has been reported to exert tumor-suppressive functions through its nonenzymatic activity and promotes the degradation of the oncogenic driver EYA2. This suggests a potential strategy for targeted protein degradation by exploiting endogenous protein-protein interactions. Here, we report the design and synthesis of a bifunctional small molecule, EC21, capable of simultaneously engaging CDK6 and EYA2. Structure-guided design based on the reported binding modes of palbociclib and the EYA2 inhibitor ETC616 enabled the identification of EC21 as a lead compound. Biological studies demonstrated that EC21 effectively reduced EYA2 protein levels in breast cancer cells in a time- and concentration-dependent manner through a ubiquitin-proteasome-dependent process. Mechanistic investigations indicated that EC21 enhances the interaction between CDK6 and EYA2, thereby promoting EYA2 degradation. Functional analyses further revealed that EC21 disrupts DNA damage repair pathways and significantly suppresses breast cancer cell proliferation. Importantly, EC21 exhibited pronounced antitumor activity in a syngeneic breast cancer mouse model and reduced EYA2 protein levels in tumor tissues without apparent toxicity. Collectively, these findings demonstrate that pharmacological stabilization of the CDK6-EYA2 interaction represents a feasible strategy for inducing EYA2 degradation and provides a potential therapeutic approach for EYA2-driven breast cancer.

Liver fibrosis is a passive and irreversible wound healing process caused by chronic liver injury. Research has shown that the upregulation of hypoxia inducible factor-1 alpha (HIF-1α) is closely related to the occurrence and development of liver fibrosis and HIF-1 α may be a promising target for the treatment of liver fibrosis. AMSP-30 m is a newly developed novel HIF-1α inhibitor by our group, which has strong anti-tumor and anti-inflammatory effects. In this study, we described the therapeutic effect and specific mechanism of AMSP-30 m on carbon tetrachloride (CCl4) induced liver fibrosis in mice. Liver fibrosis induced by CCl4 in mice and liver fibrosis induced by cobalt dichloride (CoCl2) in LX-2 cells (human hepatic stellate cell (HSC) line) were studied. Hematoxylin & eosin (H&E)and Masson's trichrome staining were used to observe pathological conditions. Western Blot, immunofluorescence and immunohistochemistry were used to detect protein expression and localization in cells, and quantitative real-time PCR analysis (qRT-PCR) was used to detect mRNA expression. Biochemical detection kits were used to detect alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels. The results demonstrated that AMSP-30 m significantly alleviated pathological symptoms, reduced ALT and AST levels, and inhibited the expression of alpha-smooth muscle actin (α-SMA) and collagen type I (COL1α1) in CCl4-induced liver fibrosis in mice. AMSP-30 m could significantly reduce the expression of HIF-1α and sonic hedgehog (Shh) pathway related proteins (Smoothened (Smo), Shh, and glioma-associated oncogene-1 (Gli-1)) in CCl4 induced liver fibrosis mice. AMSP-30 m also played a similar role in the CoCl2-induced anoxic liver fibrosis model of LX-2 cells. Further experiments showed that Cyclopamine (a Shh inhibitor) could significantly inhibit the increase of α-SMA and COL1α1 resulting from HIF-1α but not significantly inhibit HIF-1α induced by CoCl2 in LX-2 cells. And the combination of Cyclopamine and AMSP-30 m further reduced the expression of α-SMA and COL1α1 induced by HIF-1α. In summary, this study demonstrates that the HIF-1α inhibitor AMSP-30 m exerts a robust anti-fibrotic effect by inhibiting the Shh pathway, which is identified as a critical underlying mechanism. These findings suggest a promising therapeutic strategy for the treatment of liver fibrosis.

In this study, we employed a structure-based approach and step-by-step structural optimization to identify a series of quinazoline derivatives as potent receptor interacting protein kinase 3 (RIPK3) inhibitors. Among these, compound 32 emerged as the most effective inhibitor, with strong inhibition of RIPK3 (IC50 = 27 nM) and necroptosis (EC50 = 0.45 μM). Biological evaluation showed that compound 32 binds directly to RIPK3, inhibiting the phosphorylation of both RIPK3 and its downstream substrate, MLKL, thus suppressing necroptosis. Additionally, compound 32 induces Caspase-8/3-mediated apoptosis, resulting in moderate anti-proliferative effects. By converting inflammatory necroptosis to non-inflammatory apoptosis, compound 32 not only exerts anti-inflammatory effects but also reduces inflammatory hyperplasia. More importantly, compared to the known RIPK3 inhibitor HS-1371, compound 32 significantly lower toxicity in vivo in mice. In an IMQ-induced mouse model of psoriasis, compound 32 significantly alleviates skin inflammation, scaling, and hyperkeratosis, without inducing notable toxicity. This study highlights a promising therapeutic strategy for inflammatory proliferative diseases, such as psoriasis, by inhibiting RIPK3 and shifting the mode of cell death from necroptosis to apoptosis.

Gremlin 1 is a highly conserved secretory glycoprotein of the DAN family that plays significant roles in human and mouse follicular development and ovulation through both BMP-dependent and BMP-independent pathways. However, its function and mechanisms in bovine follicular development and atresia remain unclear. This study investigated its specific molecular mechanism during follicular development in bovines. Gremlin 1 protein is expressed in ovarian follicles at various developmental stages, with both its mRNA and protein levels increasing in correlation with the progression of follicular atresia. Following Si-RNA mediated knockdown of Gremlin 1 expression, a reduction in apoptosis and an enhancement in proliferative activity were observed in granulosa cells (GCs). In contrast, recombinant human Gremlin 1 (Rh-Gremlin 1) protein significantly induced apoptosis of GCs and inhibited cell proliferation. Previous studies have shown that Gremlin 1 can mediate apoptosis in a variety of cells through the TGF-β/SMAD signaling pathway. In our study, we observed that Gremlin 1 was associated with the phosphorylation levels of SMAD-2/3, and treatment with the TGF-β signaling pathway inhibitor SB431542 partially reversed Gremlin 1-induced apoptosis of GCs. Moreover, SB431542 significantly restored the proliferative viability and cell cycle progression of GCs. The results showed that Gremlin 1 was involved in follicular atresia-related processes by regulating the apoptosis and proliferation of GCs, and partially mediated this process through the TGF-β/SMAD signaling pathway. This provides a new avenue for further exploration of its role in follicular development.

Receptor-interacting serine/threonine protein kinase 2 (RIPK2) and RIPK3 have been demonstrated to be promising targets for treating multiple inflammatory diseases, including inflammatory bowel diseases (IBDs). Due to the complexity of IBD pathogenesis, on the basis of synergy strategies, we herein describe the discovery and optimization of a series of N,7-diaryl-quinazolin-4-amine derivatives as dual RIPK2 and RIPK3 inhibitors. Based on a step-by-step process involving three rounds of structural modifications, compound 29 was identified as the most one, exhibiting balanced potency against RIPK2 (IC50 = 12 nM) and RIPK3 (IC50 = 18 nM), as well as demonstrating good selectivity over other kinase targets. Further biological evaluation confirmed that compound 29 could bind directly to RIPK2 and RIPK3, effectively suppressing NOD-induced cytokine production and cellular necroptosis. Notably, compound 29 displayed significant therapeutic effects in a DSS-induced colitis mouse model, with no detectable toxicity, indicating its promising therapeutic potential as RIPK2/RIPK3 dual inhibitors for treatment of IBD.

Key PointsIGF-binding protein 7 (IGFBP7) expression was elevated in kidney and liver tissues of mice subjected to chronic cadmium exposure.IGFBP7 deficiency protected against cadmium-induced hepatorenal dysfunction and fibrosis.Inhibition of the IGFBP7/alpha-enolase/H3K18la axis may be a potential therapeutic intervention for cadmium-induced hepatorenal fibrosis.BackgroundChronic cadmium exposure can induce the onset and progression of hepatorenal fibrosis; however, its molecular basis is unclear. IGF-binding protein 7 (IGFBP7) is not only a biomarker of AKI but also plays a functional role in promoting kidney injury and inflammation. Abnormal repair of AKI causes kidney fibrosis and CKD. IGFBP7 has also been reported as a more sensitive biomarker for liver fibrosis. However, its role in hepatorenal fibrosis requires further investigation.MethodsIGFBP7 global and conditional knockout mice were used to determine the role of IGFBP7 in cadmium-induced hepatorenal fibrosis. Then, liquid chromatography-mass spectrometry, truncated mutants, coimmunoprecipitation, and microscale thermophoresis were used to unravel the downstream mechanisms.ResultsIGFBP7 expression was significantly elevated in kidney and liver tissues of mice subjected to chronic cadmium exposure. IGFBP7 deficiency attenuated cadmium-induced hepatorenal dysfunction and fibrosis, whereas restoration of IGFBP7 expression in IGFBP7-deficient mice reproduced hepatorenal fibrosis. Mechanistically, IGFBP7 interacted with alpha-enolase (ENO1) and inhibited its ubiquitination and degradation. Upregulated ENO1 further promoted glucose metabolic reprogramming and lactate accumulation. Conversely, lactate accumulation enhanced IGFBP7 transcription and expression through histone H3K18 lactylation. Importantly, therapy targeting IGFBP7 significantly ameliorated cadmium-induced hepatorenal fibrosis.ConclusionsIGFBP7 promoted cadmium-induced hepatorenal fibrosis by enhancing ENO1-driven abnormal glycolysis and lactate accumulation.

Background: Chronic cadmium exposure can induce the onset and progression of hepatorenal fibrosis; however, its molecular basis is unclear. Insulin-like growth factor-binding protein 7 (IGFBP7) is not only a biomarker of acute kidney injury (AKI), but also plays a functional role in promoting kidney injury and inflammation. Abnormal repair of AKI causes kidney fibrosis and chronic kidney disease. IGFBP7 has also been reported as a more sensitive biomarker for liver fibrosis. However, its role in hepatorenal fibrosis requires further investigation. Methods: IGFBP7 global and conditional knockout mice were used to determine the role of IGFBP7 in cadmium-induced hepatorenal fibrosis. Then, liquid chromatography-mass spectrometry, truncated mutants, co-immunoprecipitation, and microscale thermophoresis were employed to unravel the downstream mechanisms. Results: IGFBP7 expression was significantly elevated in kidney and liver tissues of mice subjected to chronic cadmium exposure. IGFBP7 deficiency attenuated cadmium-induced hepatorenal dysfunction and fibrosis, whereas restoration of IGFBP7 expression in IGFBP7-deficient mice reproduced hepatorenal fibrosis. Mechanistically, IGFBP7 interacted with alpha-enolase (ENO1) and inhibited its ubiquitination and degradation. Upregulated ENO1 further promoted glucose metabolic reprogramming and lactate accumulation. Conversely, lactate accumulation enhanced IGFBP7 transcription and expression through histone H3K18 lactylation. Importantly, therapy targeting IGFBP7 significantly ameliorated cadmium-induced hepatorenal fibrosis. Conclusions: IGFBP7 promoted cadmium-induced hepatorenal fibrosis by enhancing ENO1-driven abnormal glycolysis and lactate accumulation.

The role of N 6 -methyladenosine (m6A) modifications in colonic inflammation is poorly understood. In this study, we describe the role of fat mass and obesity-associated protein (FTO), whose expression is elevated in Crohn’s disease (CD) models as well as in human colitis samples. In a trinitrobenzene sulfonic acid (TNBS)-induced inflammatory bowel diseases mouse model, inflammation was reduced by FTO knockdown. BACH2 was determined to be a direct substrate of FTO by m6A methylated RNA immunoprecipitation and RNA sequencing analyses. BACH2 is a key regulator of CD4 + T cells differentiation, alleviating inflammatory diseases by controlling the balance between tolerance and immunity, FTO affects the differentiation of CD4 + T cells (CD25, CD44 and CD69) by reducing the expression of transcription factor BACH2, thus aggravating colon inflammation. Therefore, anti-inflammatory compounds targeting FTO are expected to alleviate damage in Crohn's disease. On basis of this finding, structure-based screening was used to identify anti-inflammatory activity licochalcone B as a potential FTO inhibitor that directly binds to FTO and attenuates colitis through inhibiting of FTO-BACH2- CD4 + T axis.

Renal fibrosis serves as a critical pathological mechanism driving the progression of chronic kidney disease (CKD). However, the pathogenesis and therapeutic targets involved in this process remain unclear. Interestingly, we currently found that IGFBP7 is highly expressed in tubular epithelial cells (TECs) from the fibrotic kidneys of human patients and animal models. However, their functional roles in abnormal kidney repair and renal fibrosis remain unclear. Here, we report that IGFBP7 knockout (KO) or TEC conditional KO (cKO) attenuated renal fibrosis in multiple mouse models, whereas IGFBP7 knock-in or restoration in IGFBP7-KO mice enhanced renal fibrosis. These in vivo findings were verified using cultured TECs and organoids generated from IGFBP7-cKO mice. Mechanistically, we found that IGFBP7 bound to pyruvate kinase M2 (PKM2) to promote the acetylation of PKM2 at the K433 site, thereby enhancing PKM2 dimerization and nuclear translocation, and subsequently accelerating lipid production and renal fibrosis via SREBP1-dependent mechanisms. Notably, through drug screening, we identified salmeterol (an asthma medication) as an IGFBP7 antagonist that effectively reduced fibrosis. Our findings reveal the IGFBP7/PKM2/SREBP1 axis as a central regulator of lipogenic fibrosis, offering genetic and pharmacological inhibition of IGFBP7 as promising therapeutic strategies for CKD.

Since synovial hypoxic microenvironment significantly promotes the pathological progress of rheumatoid arthritis (RA), hypoxia-inducible factor 1 (HIF-1) has been emerged as a promising target for the development of novel therapeutic agents for RA treatment. In this study, we designed and synthesized a series of diaryl substituted isoquinolin-1(2H)-one derivatives as HIF-1 signaling inhibitors using scaffold-hopping strategy. By modifying the substituents on N-atom and 6-position of isoquinolin-1-one, we discovered compound 17q with the most potent activities against HIF-1 (IC50 = 0.55 μM) in a hypoxia-reactive element (HRE) luciferase reporter assay. Further pharmacological studies revealed that 17q concentration-dependently blocked hypoxia-induced HIF-1α protein accumulation, reduced inflammation response, inhibited cellular invasiveness and promoted VHL-dependent HIF-1α degradation in human RA synovial cell line. Moreover, 17q improved the pathological injury of ankle joints, decreased angiogenesis and attenuated inflammation response in the adjuvant-induced arthritis (AIA) rat model, indicating the promising therapeutic potential of compound 17q as an effective HIF-1 inhibitor for RA therapy.

Paraquat (PQ) causes fatal poisoning that leads to systemic multiple organ fibrosis, and transforming growth factor (TGF)-β1 plays a critical role in this process. In this study, we aimed to investigate the effects of AZ12601011 (a small molecular inhibitor of TGFβRI) on PQ-induced multiple organ fibrosis. We established a mouse model of PQ in vivo and used PQ-treated lung epithelial cell (A549) and renal tubular epithelial cells (TECs) in vitro. Haematoxylin-eosin and Masson staining revealed that AZ12601011 ameliorated pulmonary, hepatic, and renal fibrosis, consistent with the decrease in the levels of fibrotic indicators, alpha-smooth muscle actin (α-SMA) and collagen-1, in the lungs and kidneys of PQ-treated mice. In vitro data showed that AZ12601011 suppressed the induction of α-SMA and collagen-1 in PQ-treated A549 cells and TECs. In addition, AZ12601011 inhibited the release of inflammatory factors, interleukin (IL)-1β, IL-6, and tumour necrosis factor-α. Mechanistically, TGF-β and TGFβRI levels were significantly upregulated in the lungs and kidneys of PQ-treated mice. Cellular thermal shift assay and western blotting revealed that AZ12601011 directly bound with TGFβRI and blocked the activation of Smad3 downstream. In conclusion, our findings revealed that AZ12601011 attenuated PQ-induced multiple organ fibrosis by blocking the TGF-β/Smad3 signalling pathway, suggesting its potential for PQ poisoning treatment.

Background: Shikonin (SKN), the main bioactive component isolated from Lithospermum erythrorhizon Sieb et Zucc, has multiple activities including anti-rheumatic effect, but its specific roles and the precise mechanisms in regulating biological properties of rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS) are unclear and need further clarification. Purpose: This study explored the therapeutic roles of SKN on rat adjuvant-induced arthritis (AIA) and cellular inflammation, migration and invasion of TNF-alpha-induced RA FLS (MH7A cells), and further demonstrated the involved mechanisms. Methods: SKN was intraperitoneally given to AIA rats and its therapeutic role was valued. The effects of SKN in vivo and in vitro on the production of pro-inflammatory factors were examined by ELISA and western blot. Wound-healing, transwell and phalloidin staining assay were carried out to evaluate the effects of SKN on TNF-alpha-induced migration and invasion in RA FLS. The involvement of Wnt/beta-catenin pathway was checked by immunohistochemistry or immunofluorescence assay for beta-catenin and western blot for pathway-related proteins. Results: SKN treatment in AIA rats reduced paw swelling, arthritis index and pathological damage of ankle joints, indicating its anti-arthritic effect in vivo. SKN had anti-inflammatory roles in vivo and in vitro, evidenced by inhibiting the production of pro-inflammatory factors (like IL-1 beta, IL-6, IL-8, TNF-alpha, MMP-2 and MMP-9) in sera and synovium of AIA rats, and in TNF-a-induced MH7A cells. Gelatin zymography result revealed the suppression of SKN on TNF-alpha-induced MMP-2 activity in vitro. Moreover, SKN inhibited TNF-alpha-induced migration, invasion and cytoskeletal reorganization in MH7A cells. Mechanistically, SKN suppressed the activation of Wnt/beta-catenin signaling in AIA rat synovium and in TNF-a-induced MH7A cells, indicated by the reduced protein levels of Wnt1, p-GSK-3 beta (Ser9) and beta-catenin, the raised protein level of GSK-3 beta and the decreased nuclear translocation of beta-catenin. Interestingly, the combination of LiCl (Wnt/beta-catenin agonist) canceled the therapeutic functions of SKN on cellular inflammation, migration and invasion in TNF-a-induced MH7A cells, whereas XAV939 (Wnt/beta-catenin inhibitor) enhanced the therapeutic roles of SKN. Conclusion: SKN showed therapeutic effects on rat AIA and cellular inflammation, migration and invasion of TNF-alpha-stimulated RA FLS via interrupting Wnt/beta-catenin pathway.

有机化学是药学类专业的基础课程,相对于其它专业基础课,该门课程的知识点繁杂、理解困难、熟练掌握并灵活运用不易,因此学习难度颇大.通过立足于具体教学实践,对有机化学的课程教学进行了一系列混合式教学探索,创新教学模式,包括优化教学内容、拓展线上教学、注重实践教学和优化评价体系等,希望能为药学类专业有机化学课程的教学提供一些有益的借鉴.

Signal transducer and activator of transcription 3 (STAT3) is a cell-signal transcription factor that has attracted considerable attention in recent years. The stimulation of cytokines and growth factors can result in the transcription of a wide range of genes that are crucial for several cellular biological processes involved in pro- and anti-inflammatory responses. STAT3 has attracted considerable interest as a result of a recent upsurge in study because of their role in directing the innate immune response and sustaining inflammatory pathways, which is a key feature in the pathogenesis of many diseases, including renal disorders. Several pathological conditions which may involve STAT3 include diabetic nephropathy, acute kidney injury, lupus nephritis, polycystic kidney disease, and renal cell carcinoma. STAT3 is expressed in various renal tissues under these pathological conditions. To better understand the role of STAT3 in the kidney and provide a theoretical foundation for STAT3-targeted therapy for renal disorders, this review covers the current work on the activities of STAT3 and its mechanisms in the pathophysiological processes of various types of renal diseases.

BACKGROUND:Acute kidney injury (AKI) has high morbidity and mortality, which is manifested by inflammation and apoptosis. Effective treatment methods for AKI are currently lacking. OBJECTIVE:This study demonstrated the protecting effects of Madecassoside (MA) in the cisplatin- and hypoxia-reoxygenation-induced renal tubular epithelial cells in vitro and AKI mice in vivo. METHODS:In vivo AKI mouse models were established by inducing them with cisplatin and renal ischemia-reperfusion. In vitro injury models of mouse renal tubular epithelial cells were established by inducing them with cisplatin and hypoxia and reoxygenation, respectively. The mechanism of MA effects was further explored using molecular docking and RNA-sequencing. RESULTS:MA could significantly reduce kidney injury in the cisplatin-and renal ischemia-reperfusion (IRI)-induced AKI. Further validation in the two cellular models also showed that MA had protect effects. MA can alleviate AKI in vitro and in vivo by inhibiting inflammation, cell apoptosis, and oxidative stress. MA exhibited high permeability across the Caco-2 cell, can enter cells directly. Through RNA-seq and molecular docking analysis, this study further demonstrated that MA inhibits its activity by directly binding to JNK kinase, thereby inhibiting c-JUN mediated cell apoptosis and improving AKI. In addition, MA has better renal protective effects compared to curcumin and JNK inhibitor SP600125. CONCLUSION:The results demonstrate that MA might be a potential drug for the treatment of AKI and act through the JNK/c-JUN signaling pathway.

BACKGROUND AND PURPOSE:Necroptosis plays an essential role in acute kidney injury and is mediated by receptor-interacting protein kinase 1 (RIPK1), receptor-interacting protein kinase 3 (RIPK3), and mixed lineage kinase domain-like pseudokinase (MLKL). A novel RIPK3 inhibitor, compound 42 (Cpd-42) alleviates the systemic inflammatory response. The current study was designed to investigate whether Cpd-42 exhibits protective effects on acute kidney injury and reveal the underlying mechanisms.EXPERIMENTAL APPROACH:The effects of Cpd-42 were determined in vivo through cisplatin- and ischaemia/reperfusion (I/R)-induced acute kidney injury and in vitro through cisplatin- and hypoxia/re-oxygenation (H/R)-induced cell damage. Transmission electron microscopy and periodic acid-Schiff staining were used to identify renal pathology. Cellular thermal shift assay and RIPK3-knockout mouse renal tubule epithelial cells were used to explore the relationship between Cpd-42 and RIPK3. Molecular docking and site-directed mutagenesis were used to determine the binding site of RIPK3 with Cpd-42.KEY RESULTS:Cpd-42 reduced human proximal tubule epithelial cell line (HK-2) cell damage, necroptosis and inflammatory responses in vitro. Furthermore, in vivo, cisplatin- and I/R-induced acute kidney injury was alleviated by Cpd-42 treatment. Cpd-42 inhibited necroptosis by interacting with two key hydrogen bonds of RIPK3 at Thr94 and Ser146, which further blocked the phosphorylation of RIPK3 and mitigated acute kidney injury.CONCLUSION AND IMPLICATIONS:Acting as a novel RIPK3 inhibitor, Cpd-42 reduced kidney damage, inflammatory response and necroptosis in acute kidney injury by binding to sites Thr94 and Ser146 on RIPK3. Cpd-42 could be a promising treatment for acute kidney injury.

Gastrin-releasing peptide (GRP) binds to its receptor (GRP receptor [GRPR]) to regulate multiple biological processes, but the function of GRP/GRPR axis in acute kidney injury (AKI) remains unknown. In the present study, GRPR is highly expressed by tubular epithelial cells (TECs) in patients or mice with AKI, while histone deacetylase 8 may lead to the transcriptional activation of GRPR. Functionally, we uncovered that GRPR was pathogenic in AKI, as genetic deletion of GRPR was able to protect mice from cisplatin- and ischemia-induced AKI. This was further confirmed by specifically deleting the GRPR gene from TECs in GRPRFlox/Flox//KspCre mice. Mechanistically, we uncovered that GRPR was able to interact with Toll-like receptor 4 to activate STAT1 that bound the promoter of MLKL and CCL2 to induce TEC necroptosis, necroinflammation, and macrophages recruitment. This was further confirmed by overexpressing STAT1 to restore renal injury in GRPRFlox/Flox/KspCre mice. Concurrently, STAT1 induced GRP synthesis to enforce the GRP/GRPR/STAT1 positive feedback loop. Importantly, targeting GRPR by lentivirus-packaged small hairpin RNA or by treatment with a novel GRPR antagonist RH-1402 was able to inhibit cisplatin-induced AKI. In conclusion, GRPR is pathogenic in AKI and mediates AKI via the STAT1-dependent mechanism. Thus, targeting GRPR may be a novel therapeutic strategy for AKI.

Naoluoxintong (NLXT) has been used to treat ischemic stroke (IS) in China for more than two hundred years. However, the pharmacodynamic material basis of NLXT has not been fully studied. Under the guidance of the former network pharmacological analysis, a rapid and reliable method combining UPLC-Q-TOF-MSE with the novel informatics UNIFI™ platform was established which was used to study the composition of NLXT and its prototype components and metabolites in vivo. A total of 102 compounds were identified. 13 compounds were sourced from "Monarch herb", mainly involving flavonoids and their glycosides. 54 compounds were sourced from "Minister herb", mainly involving triterpenoid saponins, organic acids and lactones. 11 compounds were from the "Assistant herb", mostly containing citric acid and esters of citric acid. 24 compounds were from the "Guide herb", mostly including flavonoids and their glycosides, organic acids and lactones. Moreover, 24 prototype components and 30 metabolites were detected, and in vivo transformation pathways for different types of chemical components were provided. This is a comprehensive report on the identification of major chemical components in NLXT and metabolic components in rats by UPLC-Q-TOF-MS combined with UNIFI platform under the guidance of network pharmacology, which is helpful for the quality control of NLXT and the study of quality markers.