Urolithin A's Role in Alleviating Severe Acute Pancreatitis via Endoplasmic Reticulum-Mitochondrial Calcium Channel Modulation.

Severe acute pancreatitis (SAP), characterized by pancreatic acinar cell death, currently lacks effective targeted therapies. Ellagic acid (EA), rich in pomegranate, shows promising anti-inflammatory and antioxidant effects in SAP treatment. However, the roles of other forms of EA, such as plant extracellular vesicles (EVs) extracted from pomegranate, and Urolithin A (UA), converted from EA through gut microbiota metabolism in vivo, have not been definitively elucidated. Our research aimed to compare the effects of pomegranate-derived EVs (P-EVs) and UA in the treatment of SAP to screen an effective formulation and to explore its mechanisms in protecting acinar cells in SAP. By comparing the protective effects of P-EVs and UA on injured acinar cells, UA showed superior therapeutic effects than P-EVs. Subsequently, we further discussed the mechanism of UA in alleviating SAP inflammation. In vivo animal experiments found that UA could not only improve the inflammatory environment of pancreatic tissue and peripheral blood circulation in SAP mice but also revealed that the mechanism of UA in improving SAP might be related to mitochondria and endoplasmic reticulum (ER) through the results including pancreatic tissue transcriptomics and transmission electron microscopy. Further research found that UA could regulate ER-mitochondrial calcium channels and reduce pancreatic tissue necroptosis. In vitro experiments of mouse pancreatic organoids and acinar cells also confirmed that UA could improve pancreatic inflammation by regulating the ER-mitochondrial calcium channel and necroptosis pathway proteins. This study not only explored the therapeutic effect of plant EVs on SAP but also revealed that UA could alleviate SAP by regulating ER-mitochondrial calcium channel and reducing acinar cell necroptosis, providing insights into the pathogenesis and potential treatment of SAP.

Urolithin A protects severe acute pancreatitis-associated acute cardiac injury by regulating mitochondrial fatty acid oxidative metabolism in cardiomyocytes.

Severe acute pancreatitis (SAP) often develops into acute cardiac injury (ACI), contributing to the high mortality of SAP. Urolithin A (UA; 3,8-dihydroxy-6H-dibenzopyran-6-one), a natural polyphenolic compound, has been extensively studied and shown to possess significant anti-inflammatory effects. Nevertheless, the specific effects of UA in SAP-associated acute cardiac injury (SACI) have not been definitively elucidated. Here, we investigated the therapeutic role and mechanisms of UA in SACI using transcriptomics and untargeted metabolomics analyses in a mouse model of SACI and in vitro studies. SACI resulted in severely damaged pancreatic and cardiac tissues with myocardial mitochondrial dysfunction and mitochondrial metabolism disorders. UA significantly reduced the levels of lipase, amylase and inflammatory factors, attenuated pathological damage to pancreatic and cardiac tissues, and reduced myocardial cell apoptosis and oxidative stress in SACI. Moreover, UA increased mitochondrial membrane potential and adenosine triphosphate production and restored mitochondrial metabolism, but the efficacy of UA was weakened by the inhibition of CPT1. Therefore, UA can attenuate cardiac mitochondrial dysfunction and reduce myocardial apoptosis by restoring the balance of mitochondrial fatty acid oxidation metabolism. CPT1 may be a potential target. This study has substantial implications for advancing our understanding of the pathogenesis and drug development of SACI.

Extracellular Vesicle ITGAM and ITGB2 Mediate Severe Acute Pancreatitis-Related Acute Lung Injury.

Integrins expressed on extracellular vesicles (EVs) secreted by various cancers are reported to mediate the organotropism of these EVs. Our previous experiment found that pancreatic tissue of mice with severe cases of acute pancreatitis (SAP) overexpresses several integrins and that serum EVs of these mice (SAP-EVs) can mediate acute lung injury (ALI). It is unclear if SAP-EV express integrins that can promote their accumulation in the lung to promote ALI. Here, we report that SAP-EV overexpress several integrins and that preincubation of SAP-EV with the integrin antagonist peptide HYD-1 markedly attenuates their pulmonary inflammation and disrupt the pulmonary microvascular endothelial cell (PMVEC) barrier. Further, we report that injecting SAP mice with EVs engineered to overexpress two of these integrins (ITGAM and ITGB2) can attenuate the pulmonary accumulation of pancreas-derived EVs and similarly decrease pulmonary inflammation and disruption of the endothelial cell barrier. Based on these findings, we propose that pancreatic EVs can mediate ALI in SAP patients and that this injury response could be attenuated by administering EVs that overexpress ITGAM and/or ITGB2, which is worthy of further study due to the lack of effective therapies for SAP-induced ALI.

Emodin Ameliorates Acute Pancreatitis-Associated Lung Injury Through Inhibiting the Alveolar Macrophages Pyroptosis.

Objective: To investigate the protective effect of emodin in acute pancreatitis (AP)-associated lung injury and the underlying mechanisms. Methods: NaT-AP model in rats was constructed using 3.5% sodium taurocholate, and CER+LPS-AP model in mice was constructed using caerulein combined with Lipopolysaccharide. Animals were divided randomly into four groups: sham, AP, Ac-YVAD-CMK (caspase-1 specific inhibitor, AYC), and emodin groups. AP-associated lung injury was assessed with H&E staining, inflammatory cytokine levels, and myeloperoxidase activity. Alveolar macrophages (AMs) pyroptosis was evaluated by flow cytometry. In bronchoalveolar lavage fluid, the levels of lactate dehydrogenase and inflammatory cytokines were measured by enzyme-linked immunosorbent assay. Pyroptosis-related protein expressions were detected by Western Blot. Results: Emodin, similar to the positive control AYC, significantly alleviated pancreas and lung damage in rats and mice. Additionally, emodin mitigated the pyroptotic process of AMs by decreasing the level of inflammatory cytokines and lactate dehydrogenase. More importantly, the protein expressions of NLRP3, ASC, Caspase1 p10, GSDMD, and GSDMD-NT in AMs were significantly downregulated after emodin intervention. Conclusion: Emodin has a therapeutic effect on AP-associated lung injury, which may result from the inhibition of NLRP3/Caspase1/GSDMD-mediated AMs pyroptosis signaling pathways.

Role of neutrophil extracellular traps in inflammatory evolution in severe acute pancreatitis.

ABSTRACT: Severe acute pancreatitis (SAP) is a life-threatening acute abdominal disease with two peaks of death: the first in the early stage, characterized by systemic inflammatory response-associated organ failure; and the second in the late stage, characterized by infectious complications. Neutrophils are the main immune cells participating in the whole process of SAP. In addition to the traditional recognition of neutrophils as the origination of chemokine and cytokine cascades or phagocytosis and degranulation of pathogens, neutrophil extracellular traps (NETs) also play an important roles in inflammatory reactions. We reviewed the role of NETs in the occurrence and development of SAP and its fatal complications, including multiple organs injury, infected pancreatic necrosis, and thrombosis. This review provides novel insights into the involvement of NETs throughout the entire process of SAP, showing that targeting NETs might be a promising strategy in SAP treatment. However, precision therapeutic options targeting NETs in different situations require further investigation.

Effect of Sheng-Jiang Powder on Gut Microbiota in High-Fat Diet-Induced NAFLD.

BACKGROUND AND AIMS: Nonalcoholic fatty liver disease (NAFLD) is an alarming global health problem that is predicted to be the major cause of cirrhosis, hepatocellular carcinoma, and liver transplantation by next decade. Gut microbiota have been revealed playing an important role in the pathogenesis of NAFLD. Sheng-Jiang Powder (SJP), an empirical Chinese medicine formula to treat NAFLD, showed great hepatoprotective properties, but the impact on gut microbiota has never been identified. Therefore, we performed this study to investigate the effect of SJP on gut microbiota in NAFLD mice. METHODS: NAFLD was induced by 12 weeks' high-fat diet (HFD) feeding. Mice were treated with SJP/normal saline daily for 6 weeks. Blood samples were obtained for serum biochemical indices and inflammatory cytokines measurement. Liver tissues were obtained for pathological evaluation and oil red O staining. The expression of lipid metabolism-related genes was quantified by RT-PCR and Western blotting. Changes in gut microbiota composition were analyzed by the 16s rDNA sequencing technique. RESULTS: HFD feeding induced significant increase in bodyweight and serum levels of TG, TC, ALT, and AST. The pathological examination revealed obvious hepatic steatosis in HFD feeding mice. Coadministration of SJP effectively protected against bodyweight increase and lipid accumulation in blood and liver. Increased expression of PPARγ mRNA was observed in HFD feeding mice, but a steady elevation of PPARγ protein level was only found in SJP-treated mice. Meanwhile, the expression of FASN was much higher in HFD feeding mice. Microbiome analysis revealed obvious changes in gut microbiota composition among diverse groups. SJP treatment modulated the relative abundance of short-chain fatty acids (SCFAs) producing bacteria, including norank-f-Erysipelotrichaceae and Roseburia. CONCLUSIONS: SJP is efficient in attenuating HFD-induced NAFLD, and it might be partly attributed to the regulation of gut microbiota.

Targeting neutrophil extracellular traps in severe acute pancreatitis treatment.

Severe acute pancreatitis (SAP) is a critical abdominal disease associated with high death rates. A systemic inflammatory response promotes disease progression, resulting in multiple organ dysfunction. The functions of neutrophils in the pathology of SAP have been presumed traditionally to be activation of chemokine and cytokine cascades accompanying the inflammatory process. Recently, since their discovery, a new type of antimicrobial mechanism, neutrophil extracellular traps (NETs), and their role in SAP, has attracted widespread attention from the scientific community. Significantly different from phagocytosis and degranulation, NETs kill extracellular microorganisms by releasing DNA fibers decorated with granular proteins. In addition to their strong antimicrobial functions, NETs participate in the pathophysiological process of many noninfectious diseases. In SAP, NETs injure normal tissues under inflammatory stress, which is associated with the activation of inflammatory cells, to cause an inflammatory cascade, and SAP products also trigger NET formation. Thus, due to the interaction between NET generation and SAP, a treatment targeting NETs might become a key point in SAP therapy. In this review, we summarize the mechanism of NETs in protecting the host from pathogen invasion, the stimulus that triggers NET formation, organ injury associated with SAP involving NETs, methods to interrupt the harmful effects of NETs, and different therapeutic strategies to preserve the organ function of patients with SAP by targeting NETs.

Effect of Sheng-jiang powder on multiple-organ inflammatory injury in acute pancreatitis in rats fed a high-fat diet.

BACKGROUND: Obesity worsens inflammatory organ injury in acute pancreatitis (AP), but there is no effective preventive strategy. Sheng-jiang powder (SJP) has been shown to alleviate multiple-organ inflammatory injury in rats with high-fat diet-induced obesity. Hence, SJP is supposed to have an effect on multiple-organ inflammatory injury in AP in rats fed a high-fat diet. AIM: To explore how obesity may contribute to aggravating inflammatory organ injury in AP in rats and observe the effect of SJP on multiple-organ inflammatory injury in AP in rats fed a high-fat diet. METHODS: Rats were randomly assigned to a control group (CG), an obese group (OG), and an SJP treatment group (SG), with eight rats per group. The rats in the OG and SG were fed a high-fat diet. From the third week, the rats in the SG were given oral doses of SJP (5 g/kg of body weight). After 12 wk, AP was induced in the three groups. Serum amylase level, body weight, Lee's index, serum biochemistry parameters, and serum inflammatory cytokine and tissue cytokine levels were assessed, and the tissue histopathological scores were evaluated and compared. RESULTS: Compared with the CG, serum triglyceride, total cholesterol, interleukin-6, and interleukin-10 levels were significantly higher in the OG, and serum high-density lipoprotein cholesterol level was significantly lower in the OG. Moreover, enhanced oxidative damage was observed in the pancreas, heart, spleen, lung, intestine, liver, and kidney. Evidence of an imbalanced antioxidant defense system, especially in the pancreas, spleen, and intestine, was observed in the obese AP rats. Compared with the OG, serum high-density lipoprotein cholesterol, interleukin-10, and superoxide dismutase expression levels in the pancreas, spleen, and intestine were increased in the SG. Additionally, SJP intervention led to a decrease in the following parameters: body weight; Lee's index; serum triglyceride levels; serum total cholesterol levels; malondialdehyde expression levels in the pancreas, heart, spleen, lung, and liver; myeloperoxidase expression levels in the lung; and pathological scores in the liver. CONCLUSION: Obesity may aggravate the inflammatory reaction and pathological multiple-organ injury in AP rats, and SJP may alleviate multiple-organ inflammatory injury in AP in rats fed a high-fat diet.

Sheng-Jiang Powder Ameliorates High Fat Diet Induced Nonalcoholic Fatty Liver Disease via Inhibiting Activation of Akt/mTOR/S6 Pathway in Rats.

BACKGROUND AND AIMS: Nonalcoholic fatty liver disease (NAFLD) is an alarming public health problem that directly contributes to increased prevalence of liver cirrhosis and hepatic cell cancer, but without any specific pharmacological option. Sheng-jiang powder (SJP), an empirical Chinese medicine formula to treat NAFLD, showed great efficacy but the specific mechanisms have never been reported. Therefore, we performed this study to explore the effect of SJP on NAFLD and the potential mechanism. METHODS: NAFLD was induced by high fat diet (HFD) feeding. Rats were treated with SJP/normal saline daily for 10 weeks and all rats were euthanized after 12 weeks' feeding. Liver tissue samples were obtained for biochemistry test and pathological evaluation. Additionally, oleic acid induced LO2 cells were employed to simulate a cell model of NAFLD. Cells were subjected to western blotting for Akt, mTOR, S6, SREBP1-c, and FASN detection after coincubated with SJP, LY294002 (a selective PI3K inhibitor), or the combination for 24h. RESULTS: HFD significantly induced hepatic steatosis. Plenty of lipid droplets were observed under transmission electron microscope. The ultrastructure of liver cells showed distinct changes with obvious endoplasmic reticulum expansion, mitochondrial contraction, and cell matrix solidification. Although no difference was detected in serum hepatic enzymes and tissue proinflammatory cytokines, the tissue level of SOD and GSH-px was much lower and the pathologic injuries were much severe in HFD feeding rats. However, SJP treatment significantly attenuated the ultrastructure changes and protected rat liver against inflammatory injury. Abundant of lipid droplets and high expression of pAkt, pmTOR, pS6, and FASN were observed in oleic acid treated LO2 cells, while these changes were restored by SJP treatment. CONCLUSIONS: SJP is efficient in attenuating HFD induced NAFLD in rats and this effect might be partly related to the inhibition of Akt/mTOR/S6 pathway.

Effect of Da-Cheng-Qi decoction for treatment of acute kidney injury in rats with severe acute pancreatitis.

BACKGROUND: The traditional Chinese formula Da-Cheng-Qi-decoction (DCQD) has been used to treat acute pancreatitis for decades. DCQD could ameliorate the disease severity and the complications of organ injuries, including those of the liver and lungs. However, the pharmacological effects in the kidney, a target organ, are still unclear. This study aimed to investigate the herbal tissue pharmacology of DCQD for acute kidney injury (AKI) in rats with severe acute pancreatitis (SAP). METHODS: Rats were randomly divided into the sham-operation group (SG), the model group (MG) and the low-, medium- and high-dose treatment groups (LDG, MDG, and HDG, respectively). Sodium taurocholate (3.5%) was retrogradely perfused into the biliopancreatic duct to establish the model of SAP in rats. Different doses of DCQD were administered to the treatment groups 2 h after the induction of SAP. The major components of DCQD in kidney tissues were detected by HPLC-MS/MS. Inflammatory mediators in the kidney tissues, as well as serum creatinine (Scr), blood urea nitrogen (BUN) and pathologic scores, were also evaluated. RESULTS: Ten components of DCQD were detected in the kidneys of the treatment groups, and their concentrations increased dose-dependently. Compared with the SG, the levels of inflammatory mediators, Scr, BUN and pathological scores in the MG were obviously increased (p < 0.05). The high dose of DCQD showed a maximal effect in downregulating the pro-inflammatory mediators interleukin-6 (IL)-6 and tumour necrosis factor-α (TNF-α), upregulating anti-inflammatory mediators IL-4 and IL-10 in the kidney and alleviating the pathological damages. DCQD decreased the pancreas and kidney pathological scores of rats with SAP, especially in the HDG (p < 0.05). Compared with the MG, the level of Scr in the HDG was significantly decreased (p < 0.05). CONCLUSIONS: DCQD ameliorated AKI in rats with SAP via regulating the inflammatory response, which might be closely related to the distribution of its components in the kidney.

Effect of Sheng-Jiang Powder on Obesity-Induced Multiple Organ Injuries in Rats.

BACKGROUND AND AIMS: Obesity has become the main public health issue nowadays with poor control and has been associated with increased risk of multiorgan disease, but the specific mechanism and effective medication are still to be addressed. Sheng-jiang powder (SJP) showed great potential in preventing obesity in Chinese researches but has no trace in English reports. This study was designed to investigate the effect of SJP on obesity and obesity-mediated multiorgan injuries. METHODS: Rats were randomized into normal group (NG), obese group (OG), and SJP treatment group (SG). Obesity was induced by high-fat diet feeding. Rats were gavaged with SJP/normal saline daily from the third week and all rats were sacrificed after 12 weeks' feeding. Tissues were obtained for cytokines tests. RESULTS: Firstly, high-fat diet feeding led to significant obesity. Compared to NG, the level of SOD in the liver, spleen, lung, and kidney was much lower in OG (p < 0.05), while the pathological scores of pancreas, liver, spleen, lung, and kidney were much higher. SJP significantly increased SOD level in the liver, spleen, and lung and reduced the pathological scores of pancreas, liver, spleen, lung, and kidney correspondingly (p < 0.05). CONCLUSIONS: SJP ameliorates inflammatory response and mitigates obesity-induced multiple organ injuries.

Optimal timing for the oral administration of Da-Cheng-Qi decoction based on the pharmacokinetic and pharmacodynamic targeting of the pancreas in rats with acute pancreatitis.

AIM: To identify the optimal oral dosing time of Da-Cheng-Qi decoction (DCQD) in rats with acute pancreatitis (AP) based on the pharmacokinetic and pharmacodynamic parameters. METHODS: First, 24 male Sprague-Dawley rats were divided into a sham-operated group [NG(a)] and three model groups [4hG(a), 12hG(a) and 24hG(a)]. The NG(a) and model groups were administered DCQD (10 g/kg.BW) intragastrically at 4 h, 4 h, 12 h and 24 h, respectively, after AP models induced by 3% sodium taurocholate. Plasma samples were collected from the tails at 10 min, 20 min, 40 min, 1 h, 2 h, 4 h, 8 h, 12 h and 24 h after a single dosing with DCQD. Plasma and pancreatic tissue concentrations of the major components of DCQD were determined by high-performance liquid chromatography tandem mass spectroscopy. The pharmacokinetic parameters and serum amylase were detected and compared. Second, rats were divided into a sham-operated group [NG(b)] and three treatment groups [4hG(b), 12hG(b) and 24hG(b)] with three corresponding control groups [MG(b)s]. Blood and pancreatic tissues were collected 24 h after a single dosing with DCQD. Serum amylase, inflammatory cytokines and pathological scores of pancreatic tissues were detected and compared. RESULTS: The concentrations of emodin, naringin, honokiol, naringenin, aloe-emodin, chrysophanol and rheochrysidin in the 12hG(a) group were higher than those in the 4hG(a) group in the pancreatic tissues (P < 0.05). The area under the plasma concentration-time curve from time 0 to the time of the last measurable concentration values (AUC0→t) for rhein, chrysophanol, magnolol and naringin in the 12hG(a) group were larger than those in the 4hG(a) or 24hG(a) groups. The 12hG(a) group had a higher Cmax than the other two model groups. The IL-10 levels in the 12hG(b) and 24hG(b) groups were higher than in the MG(b)s (96.55 ± 7.84 vs 77.46 ± 7.42, 251.22 ± 16.15 vs 99.72 ± 4.7 respectively, P < 0.05), while in the 24hG(b) group, the IL-10 level was higher than in the other two treatment groups (251.22 ± 16.15 vs 154.41 ± 12.09/96.55 ± 7.84, P < 0.05). The IL-6 levels displayed a decrease in the 4hG(b) and 12hG(b) groups compared to the MG(b)s (89.99 ± 4.61 vs 147.91 ± 4.36, 90.82 ± 5.34 vs 171.44 ± 13.43, P < 0.05). CONCLUSION: Late-time dosing may have higher concentrations of the most major components of DCQD, with better pharmacokinetics and pharmacodynamics of anti-inflammation than early-time dosing, which showed the late time to be the optimal dosing time of DCQD for AP.

Pharmacokinetics and pharmacodynamics of Shengjiang decoction in rats with acute pancreatitis for protecting against multiple organ injury.

AIM: To explore the pharmacokinetics and pharmacodynamics of Shengjiang decoction (SJD) in rats with acute pancreatitis (AP) for protecting against multiple organ injury. METHODS: An AP model was established by retrograde perfusion of 3.5% sodium taurocholate into the biliopancreatic duct, and a control group (CG) received 0.9% sodium chloride instead. Twelve male Sprague-Dawley rats were randomly divided into a CG treated with SJD (CG + SJD) and a model group treated with SJD (MG + SJD), both of which were orally administered with SJD (5 g/kg) 2 h after surgery. Blood samples were collected via the tail vein at 10, 20, and 40 min and 1, 2, 3, 4, 6, 8, and 12 h after a single dose of SJD to detect its main components using high-performance liquid chromatography-tandem mass spectrometry. The pharmacokinetic parameters were compared. In the pharmacodynamic experiment, 18 male Sprague-Dawley rats were randomly divided into a CG, an AP model group (MG), and an SJD treated AP group (SJDG). Serum amylase, lipase, and inflammatory cytokines were measured, and heart, lung, liver, spleen, pancreas, kidney, and intestine tissues were collected for pathological examination. RESULTS: The MG + SJD displayed significantly shorter mean residence time (MRT) and higher clearance (CL) for emodin and aloe-emodin; significantly shorter time of maximum concentration and T1/2 and a lower area under curve (AUC) for aloe-emodin; a significantly higher AUC and lower CL for rhein; and longer MRT and lower CL for chrysophanol than the CG + SJD. In the pharmacodynamic experiment, the amylase, interleukin (IL)-6, IL-10, and tumor necrosis factor (TNF)-α levels in the MG were higher than those in the CG (P < 0.05). After the herbal decoction treatment, the SJDG had higher IL-10 and lower TNF-α levels than the MG (P < 0.05). The MG had the highest pathological scores, and the pathological scores of the lung, pancreas, kidney, and intestine in the SJDG were significantly lower than those in the MG (P < 0.05). CONCLUSION: AP may have varying effects on the pharmacokinetics of the major SJD components in rats. SJD might alleviate pathological injuries of the lung, pancreas, kidney, and intestine in rats with AP via regulating pro- and anti- inflammatory responses, which might guide the clinical application of SJD for AP treatment.