A microRNA checkpoint for Ca2+ signaling and overload in acute pancreatitis.

Acute pancreatitis (AP) is a common digestive disease without specific treatment, and its pathogenesis features multiple deleterious amplification loops dependent on translation, triggered by cytosolic Ca2+ ([Ca2+]i) overload; however, the underlying mechanisms in Ca2+ overload of AP remains incompletely understood. Here we show that microRNA-26a (miR-26a) inhibits pancreatic acinar cell (PAC) store-operated Ca2+ entry (SOCE) channel expression, Ca2+ overload, and AP. We find that major SOCE channels are post-transcriptionally induced in PACs during AP, whereas miR-26a expression is reduced in experimental and human AP and correlated with AP severity. Mechanistically, miR-26a simultaneously targets Trpc3 and Trpc6 SOCE channels and attenuates physiological oscillations and pathological elevations of [Ca2+]i in PACs. MiR-26a deficiency increases SOCE channel expression and [Ca2+]i overload, and significantly exacerbates AP. Conversely, global or PAC-specific overexpression of miR-26a in mice ameliorates pancreatic edema, neutrophil infiltration, acinar necrosis, and systemic inflammation, accompanied with remarkable improvements on pathological determinants related with [Ca2+]i overload. Moreover, pancreatic or systemic administration of an miR-26a mimic to mice significantly alleviates experimental AP. These findings reveal a previously unknown mechanism underlying AP pathogenesis, establish a critical role for miR-26a in Ca2+ signaling in the exocrine pancreas, and identify a potential target for the treatment of AP.

Disruption of the primary salicylic acid hydroxylases in rice enhances broad-spectrum resistance against pathogens.

Salicylic acid (SA) is a crucial hormone involved in plant immunity. Rice (Oryza sativa) maintains high SA levels that are not induced by pathogens. However, the roles of SA in rice immunity and yield remain largely unknown. Here, we identified SA 5-hydroxylases 1 (OsS5H1) and 2 (OsS5H2) as the primary enzymes engaged in catalysing SA to 2,5-dihydroxybenzoic acid (2,5-DHBA) in rice. SA levels were significantly increased in the oss5h mutants, while they were dramatically decreased in the OsS5H1 and OsS5H2 overexpression lines. The mutants were resistant, whereas the overexpression lines were susceptible to Pyricularia oryzae and Xanthomonas oryzae pv. Oryzae. Moreover, the pathogen-associated molecular patterns-triggered immunity responses, including reactive oxygen species burst and callose deposition, were enhanced in all the mutants and compromised in the overexpression lines. Quantification of the agronomic traits of the oss5h mutants grown in the paddy fields demonstrated that the grain number per panicle was decreased as the SA levels increased; however, the tiller number and grain size were enhanced, resulting in no significant yield penalty. Collectively, we reveal that mildly increasing SA content in rice can confer broad-spectrum resistance without yield penalty and put new insights into the roles of SA in immunity and growth.

Stress Hyperglycemia Is Independently Associated with Persistent Organ Failure in Acute Pancreatitis.

BACKGROUND/AIMS: Stress hyperglycemia is common in critical illness but it has not been clearly studied in patients with acute pancreatitis (AP). This study aimed to investigate the specific blood glucose (BG) level that defines stress hyperglycemia and to determine the impact of stress hyperglycemia on clinical outcomes in AP patients. METHODS: AP patients admitted ≤ 48 h after abdominal pain onset were retrospectively analyzed. Patients were stratified by pre-existing diabetes and stress hyperglycemia was defined using stratified BG levels for non-diabetes and diabetes with clinical outcomes compared. RESULTS: There were 967 non-diabetic and 114 diabetic (10.5%) patients met the inclusion criteria and the clinical outcomes between these two groups were not significantly different. In non-diabetes, the cut-off BG level of ≥ 180 mg/dl was selected to define stress hyperglycemia with an 8.8-fold higher odds ratio for persistent organ failure (POF) (95% CI 5.4-14.3; P < 0.001). For diabetes, ≥ 300 mg/dl was selected with a 7.5-fold higher odds ratio for POF (95% CI 1.7-34.3; P = 0.009). In multivariable logistic regression, stress hyperglycemia was independently associated with POF, acute necrotic collection, major infection and mortality. The combination of BG and systemic inflammatory response syndrome (SIRS) score in predicting POF was better than SIRS or Glasgow score alone. CONCLUSIONS: This study identifies a cut-off BG level of ≥ 180 mg/dl and ≥ 300 mg/dl was optimal to define stress hyperglycemia for non-diabetic and diabetic AP patients, respectively. There was a significant relationship between stress hyperglycemia and adverse clinical outcomes.

Predicting persistent organ failure on admission in patients with acute pancreatitis: development and validation of a mobile nomogram.

BACKGROUND: Early prediction of persistent organ failure (POF) is important for triage and timely treatment of patients with acute pancreatitis (AP). METHODS: All AP patients were consecutively admitted within 48 h of symptom onset. A nomogram was developed to predict POF on admission using data from a retrospective training cohort, validated by two prospective cohorts. The clinical utility of the nomogram was defined by concordance index (C-index), decision curve analysis (DCA), and clinical impact curve (CIC), while the performance by post-test probability. RESULTS: There were 816, 398, and 880 patients in the training, internal and external validation cohorts, respectively. Six independent predictors determined by logistic regression analysis were age, respiratory rate, albumin, lactate dehydrogenase, oxygen support, and pleural effusion and were included in the nomogram (web-based calculator: https://shina.shinyapps.io/DynNomapp/). This nomogram had reasonable predictive ability (C-indexes 0.88/0.91/0.81 for each cohort) and promising clinical utility (DCA and CIC). The nomogram had a positive likelihood ratio and post-test probability of developing POF in the training, internal and external validation cohorts of 4.26/31.7%, 7.89/39.1%, and 2.75/41%, respectively, superior or equal to other prognostic scores. CONCLUSIONS: This nomogram can predict POF of AP patients and should be considered for clinical practice and trial allocation.

Ethyl pyruvate and analogs as potential treatments for acute pancreatitis: A review of in vitro and in vivo studies.

Ethyl pyruvate (EP) has been shown to improve outcomes from multiple organ dysfunction syndrome (MODS) in experimental animal models of critical illness. This review aimed to summarise in vitro and in vivo effects of EP analogs on acute pancreatitis (AP) with the objective of proposing medicinal chemistry modifications of EP for future research. In vitro studies showed that both sodium pyruvate and EP significantly reduced pancreatic acinar necrotic cell death pathway activation induced by multiple pancreatic toxins. In vivo studies using different murine AP models showed that EP (usually at a dose of 40 mg/kg every 6 h) consistently reduced pain, markers of pancreatic injury, systemic inflammation and MODS. There was also a significant increase in survival rate, even when EP was administered 12 h after disease induction (compared with untreated groups or those treated with Ringer's lactate solution). Experimental studies suggest that EP and analogs are promising drug candidates for treating AP. EP or analogs can undergo medicinal chemistry modifications to improve its stability and deliverability. EP or analogs could be evaluated as a supplement to intravenous fluid therapy in AP.

[Protective Effect of a Dihydroflavonol Glycoside from Coreopsis tinctoria Nutt. in Mouse Model of Alcoholic Acute Pancreatitis].

OBJECTIVE: To investigate the protective effect of (2R, 3R)-dihydroquercetin 7-O-ß-D-glucopyranose (C1) extracted from Coreopsis tinctoria Nutt. in a mouse model of alcoholic acute pancreatitis (FAEE-AP) induced byfatty acid ethyl ester (FAEE). METHODS: The 30 healthy SPF mice were randomly divided into control group, model group, low dose group, middle dose group and high dose group, 6 in each group. Alcoholic pancreatitis was induced by ethanol and palmitoleic acid administration (1.75 g/kg ethanol, 200 mg/kg palmitoleic acid, 2 times peritoneal injections). The three treatment groups were given C1 (0 h, 4 h, 8 h) at the dose of 12.5, 25 and 50 mg/kg, respectively. After 24 h of molding, the serum amylase, lipase and IL-6 levels were detected. The trypsin level in pancreatic tissue and myeloperoxidase (MPO) level in pancreatic and lung tissue were detected. Hematoxylin-eosin (HE) staining was used to observe the pathological changes of pancreatic tissue and immunohistochemical (IHC) staining was used to detect the expression of nuclear factor-erythroid 2 related factor 2 (Nrf2) in pancreatic tissue. RESULTS: The pancreatic histopathological scores, serum amylase and lipase activity, trypsin level in pancreatic tissue, serum IL-6 level, MPO level of pancreas and lung were significantly higher in the model group than in the control group (P < 0.01). Compared with the model group, the pancreatic histopathologies of the low dose group was significantly improved (P < 0.05), as well as the serum amylase and lipase activity, trypsin level of pancreas, serum IL-6 level, the pancreas andthe lung's MPO level decreased significantly (P < 0.05), and up-regulate that expression of Nrf2 in pancreatic tissue. CONCLUSION: 12.5 mg/kg of (2R, 3R) -dihydroquercetin 7-O-ß-D-glucopyranose (C1) improved the expression of Nrf2, reduced the expression of inflammatory factor IL-6, and protected acute pancreatitis caused by FAEE.

Simulated microgravity affects some biological characteristics of Lactobacillus acidophilus.

The effects of weightlessness on enteric microorganisms have been extensively studied, but have mainly been focused on pathogens. As a major component of the microbiome of the human intestinal tract, probiotics are important to keep the host healthy. Accordingly, understanding their changes under weightlessness conditions has substantial value. This study was carried out to investigate the characteristics of Lactobacillus acidophilus, a typical probiotic for humans, under simulated microgravity (SMG) conditions. The results revealed that SMG had no significant impact on the morphology of L. acidophilus, but markedly shortened its lag phase, enhanced its growth rate, acid tolerance ability up to pH < 2.5, and the bile resistance at the bile concentration of <0.05%. SMG also decreased the sensitivity of L. acidophilus to cefalexin, sulfur gentamicin, and sodium penicillin. No obvious effect of SMG was observed on the adhesion ability of L. acidophilus to Caco-2 cells. Moreover, after SMG treatment, both the culture of L. acidophilus and its liquid phase exhibited higher antibacterial activity against S. typhimurium and S. aureus in a time-dependent manner. The SMG treatment also increased the in vitro cholesterol-lowering ability of L. acidophilus by regulating the expression of the key cholesterol metabolism genes CYP7A1, ABCB11, LDLR, and HMGCR in the HepG2 cell line. Thus, the SMG treatment did have considerable influence on some biological activities and characteristics of L. acidophilus related to human health. These findings provided valuable information for understanding the influence of probiotics on human health under simulated microgravity conditions, at least.

Transcriptomics and Network Pharmacology Reveal the Protective Effect of Chaiqin Chengqi Decoction on Obesity-Related Alcohol-Induced Acute Pancreatitis via Oxidative Stress and PI3K/Akt Signaling Pathway.

Obesity-related acute pancreatitis (AP) is characterized by increasing prevalence worldwide and worse clinical outcomes compared to AP of other etiologies. Chaiqin chengqi decoction (CQCQD), a Chinese herbal formula, has long been used for the clinical management of AP but its therapeutic actions and the underlying mechanisms have not been fully elucidated. This study has investigated the pharmacological mechanisms of CQCQD in a novel mouse model of obesity-related alcohol-induced AP (OA-AP). The mouse OA-AP model was induced by a high-fat diet for 12 weeks and subsequently two intraperitoneal injections of ethanol, CQCQD was administered 2 h after the first injection of ethanol. The severity of OA-AP was assessed and correlated with changes in transcriptomic profiles and network pharmacology in the pancreatic and adipose tissues, and further docking analysis modeled the interactions between compounds of CQCQD and their key targets. The results showed that CQCQD significantly reduced pancreatic necrosis, alleviated systemic inflammation, and decreased the parameters associated with multi-organ dysfunction. Transcriptomics and network pharmacology analysis, as well as further experimental validation, have shown that CQCQD induced Nrf2/HO-1 antioxidant protein response and decreased Akt phosphorylation in the pancreatic and adipose tissues. In vitro, CQCQD protected freshly isolated pancreatic acinar cells from H2O2-elicited oxidative stress and necrotic cell death. The docking results of AKT1 and the active compounds related to AKT1 in CQCQD showed high binding affinity. In conclusion, CQCQD ameliorates the severity of OA-AP by activating of the antioxidant protein response and down-regulating of the PI3K/Akt signaling pathway in the pancreas and visceral adipose tissue.

Ketogenesis acts as an endogenous protective programme to restrain inflammatory macrophage activation during acute pancreatitis.

BACKGROUND: Innate immunity and metabolites link to the pathogenesis and severity of acute pancreatitis (AP). However, liver metabolism and its role in immune response and AP progression remain elusive. We investigated the function of liver metabolism in the pathogenesis of AP. METHODS: Circulating ketone body ß-hydroxybutyrate (ßOHB) levels were determined in AP clinical cohorts and caerulein-induced AP (CER-AP) mouse models receiving seven (Cer*7) or twelve (Cer*12) injection regimens at hourly intervals. Liver transcriptomics and metabolomics were compared between CER-AP (Cer*7) and CER-AP (Cer*12). Inhibition of fatty acid ß-oxidation (FAO)-ketogenesis, or supplementation of ßOHB was performed in mouse models of AP. The effect and mechanism of ßOHB were examined in vitro. FINDINGS: Elevated circulating ßOHB was observed in patients with non-severe AP (SAP) but not SAP. These findings were replicated in CER-AP (Cer*7) and CER-AP (Cer*12), which manifested as limited and hyperactive immune responses, respectively. FAO-ketogenesis was activated in CER-AP (Cer*7), while impaired long-chain FAO and mitochondrial function were observed in the liver of CER-AP (Cer*12). Blockage of FAO-ketogenesis (Cpt1a antagonism or Hmgcs2 knockdown) worsened, while supplementation of ßOHB or its precursor 1,3-butanediol alleviated the severity of CER-AP. Mechanistically, ßOHB had a discernible effect on pancreatic acinar cell damage, instead, it greatly attenuated the activation of pancreatic and systemic proinflammatory macrophages via class I histone deacetylases. INTERPRETATION: Our findings reveal that hepatic ketogenesis is activated as an endogenous protective programme to restrain AP progression, indicating its potential therapeutic value. FUNDING: This work was supported by the National Natural Science Foundation of China, Shanghai Youth Talent Support Programme, and Shanghai Municipal Education Commission-Gaofeng Clinical Medicine Grant.

Impact of admission and early persistent stress hyperglycaemia on clinical outcomes in acute pancreatitis.

Background: To determine the impact of glucose levels at admission and during first week (early phase) on clinical outcomes in patients with acute pancreatitis (AP) and to investigate the relationship between stress hyperglycaemia (SHG) and hypertriglyceridaemia (HTG). Methods: Two independent and prospective databases were retrospectively analysed (n = 1792). Patients admitted with pain of less than 48 hours and confirmed AP were included. SHG was defined as admission blood glucose ≥ 10.00 mmol/L (non-diabetic) or ≥ 16.67 mmol/L (diabetic). Blood glucose records for the first week were inspected to determine whether SHG lasted ≥ 48 hours (persistent) or < 48 hours (transient). Clinical outcomes were compared between designated patient groups using multivariate and trend analyses. The correlation between SHG and HTG (serum triglyceride ≥ 5.65 mmol/L) was also analysed. Results: On admission, SHG was present in 27.8% (499/1792) patients; during the first 48 hours of admission, transient and persistent SHG was found in 31% (556/1792) and 8.0% (144/1792) patients, respectively. Admission SHG was associated with higher incidence of persistent organ failure, acute necrotic collection, major infection, and mortality as well as prolonged length of hospital stay (all P < 0.05). Duration of SHG was also associated with worsened clinical outcomes (all P < 0.05). In HTG-AP patients, more severe clinical outcomes were observed in those who concomitantly had SHG (P < 0.05). Conclusions: Admission and persistent SHG during the first week of admission worsens clinical outcomes of AP patients. These effects are more pronounced when admission HTG co-existed.

Alleviation of acute pancreatitis-associated lung injury by inhibiting the p38 mitogen-activated protein kinase pathway in pulmonary microvascular endothelial cells.

BACKGROUND: Previous reports have suggested that the p38 mitogen-activated protein kinase signaling pathway is involved in the development of severe acute pancreatitis (SAP)-related acute lung injury (ALI). Inhibition of p38 by SB203580 blocked the inflammatory responses in SAP-ALI. However, the precise mechanism associated with p38 is unclear, particularly in pulmonary microvascular endothelial cell (PMVEC) injury. AIM: To determine its role in the tumor necrosis factor-alpha (TNF-α)-induced inflammation and apoptosis of PMVECs in vitro. We then conducted in vivo experiments to confirm the effect of SB203580-mediated p38 inhibition on SAP-ALI. METHODS: In vitro, PMVEC were transfected with mitogen-activated protein kinase kinase 6 (Glu), which constitutively activates p38, and then stimulated with TNF-α. Flow cytometry and western blotting were performed to detect the cell apoptosis and inflammatory cytokine levels, respectively. In vivo, SAP-ALI was induced by 5% sodium taurocholate and three different doses of SB203580 (2.5, 5.0 or 10.0 mg/kg) were intraperitoneally injected prior to SAP induction. SAP-ALI was assessed by performing pulmonary histopathology assays, measuring myeloperoxidase activity, conducting arterial blood gas analyses and measuring TNF-α, interleukin (IL)-1ß and IL-6 levels. Lung microvascular permeability was measured by determining bronchoalveolar lavage fluid protein concentration, Evans blue extravasation and ultrastructural changes in PMVECs. The apoptotic death of pulmonary cells was confirmed by performing a terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling analysis and examining the Bcl2, Bax, Bim and cle-caspase3 levels. The proteins levels of P-p38, NFκB, IκB, P-signal transducer and activator of transcription-3, nuclear factor erythroid 2-related factor 2, HO-1 and Myd88 were detected in the lungs to further evaluate the potential mechanism underlying the protective effect of SB203580. RESULTS: In vitro, mitogen-activated protein kinase (Glu) transfection resulted in higher apoptotic rates and cytokine (IL-1ß and IL-6) levels in TNF-α-treated PMVECs. In vivo, SB2035080 attenuated lung histopathological injury, decreased inflammatory activity (TNF-α, IL-1ß, IL-6 and myeloperoxidase) and preserved pulmonary function. Furthermore, SB203580 significantly reversed changes in the bronchoalveolar lavage fluid protein concentration, Evans blue accumulation, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive cell numbers, apoptosis-related proteins (cle-caspase3, Bim and Bax) and endothelial microstructure. Moreover, SB203580 significantly reduced the pulmonary P-p38, NFκB, P-signal transducer and activator of transcription-3 and Myd88 levels but increased the IκB and HO-1 levels. CONCLUSION: p38 inhibition may protect against SAP-ALI by alleviating inflammation and the apoptotic death of PMVECs.

Experimental Acute Pancreatitis Models: History, Current Status, and Role in Translational Research.

Acute pancreatitis is a potentially severe inflammatory disease that may be associated with a substantial morbidity and mortality. Currently there is no specific treatment for the disease, which indicates an ongoing demand for research into its pathogenesis and development of new therapeutic strategies. Due to the unpredictable course of acute pancreatitis and relatively concealed anatomical site in the retro-peritoneum, research on the human pancreas remains challenging. As a result, for over the last 100 years studies on the pathogenesis of this disease have heavily relied on animal models. This review aims to summarize different animal models of acute pancreatitis from the past to present and discuss their main characteristics and applications. It identifies key studies that have enhanced our current understanding of the pathogenesis of acute pancreatitis and highlights the instrumental role of animal models in translational research for developing novel therapies.

Chaiqin chengqi decoction alleviates severe acute pancreatitis associated acute kidney injury by inhibiting endoplasmic reticulum stress and subsequent apoptosis.

BACKGROUND: Acute kidney injury (AKI), characterized by an increase of serum creatinine and urea, is a severe complication of severe acute pancreatitis (SAP) with high mortality. Endoplasmic reticulum (ER) stress has been considered as a key pathologic process in AKI. Chaiqin chengqi decoction (CQCQD) is an effective Chinese medicine formula for SAP treatment in China and has been used for many years. Our goal is to explore the role of CQCQD on ER stress of AKI in experimental SAP. MATERIALS & METHODS: SAP was induced in rats by retrograde duct injection of 5% sodium taurocholate (NaTC, 1 ml/kg), sham operation (SO) rats simultaneously received saline infusion. Intraperitoneal injection of 4-PBA (50 mg/kg, once a day for three days before the surgery) or intragastric gavage of CQCQD (1 g/kg, 2 hourly × 3 after disease induction) was used to treat SAP rats. All animals were humanely sacrificed 12 h after disease induction. Histopathology scores of kidney and pancreas; serum biochemical indices and kidney protein levels of ER stress and apoptosis markers were assessed. Tubular epithelial cell line (HK-2) was treated either with TNF-α (10 ng/ml) or IL-6 (10 ng/ml) for 12 h plus either 4-PBA (0.1 M) or CQCQD (1 mg/ml) for in vitro study. Cell viability and markers of ER stress and apoptosis were measured. RESULTS: Ductal perfusion of NaTC caused significant increases in serum lipase, amylase and pancreatic histopathology (inflammatory cell infiltration, interstitial edema, and acinar cell necrosis). Kidney histopathology (tubular dilation, brush border loss, little tubular necrosis, and cast formation), serum creatine and urea levels were raised when compared with the SO group. Moreover, apoptotic cell death markers (caspase-9, cleaved-caspase-3, and TUNEL) and kidney ER stress proteins (BIP, IRE1-α, XBP1s, and CHOP) were elevated after NaTC administration. 4-PBA and CQCQD significantly alleviated histopathological changes of kidney and pancreas, inflammatory cytokines, biochemical markers of AKI, ER stress proteins and apoptotic cell death markers. They also protected HK-2 cells from injury of TNF-α and IL-6, and alleviated both ER stress and apoptosis proteins in vitro. CONCLUSION: CQCQD may alleviate SAP-related AKI by inhibiting ER stress-related apoptosis.

Chaiqin chengqi decoction alleviates severity of acute pancreatitis via inhibition of TLR4 and NLRP3 inflammasome: Identification of bioactive ingredients via pharmacological sub-network analysis and experimental validation.

BACKGROUND: Chaiqin chengqi decoction (CQCQD) is a Chinese herbal formula derived from dachengqi decoction. CQCQD has been used for the management of acute pancreatitis (AP) in the West China Hospital for more than 30 years. Although CQCQD has a well-established clinical efficacy, little is known about its bioactive ingredients, how they interact with different therapeutic targets and the pathways to produce anti-inflammatory effects. PURPOSE: Toll-like receptor 4 (TLR4) and the nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3) inflammasome-mediated pro-inflammatory signaling pathways, play a central role in AP in determining the extent of pancreatic injury and systemic inflammation. In this study, we screened the bioactive ingredients using a pharmacological sub-network analysis based on the TLR4/NLRP3 signaling pathways followed by experimental validation. METHODS: The main CQCQD bioactive compounds were identified by UPLC-QTOF/MS. The TLR4/NLRP3 targets in AP for CQCQD active ingredients were confirmed through a pharmacological sub-network analysis. Mice received 7 intraperitoneal injections of cerulein (50 µg/kg; hourly) to induce AP (CER-AP), while oral gavage of CQCQD (5, 10, 15 and 20 g/kg; 3 doses, 2 hourly) was commenced at the 3rd injection of cerulein. Histopathology and biochemical indices were used for assessing AP severity, while polymerase chain reaction, Western blot and immunohistochemistry analyses were used to study the mechanisms. Identified active CQCQD compounds were further validated in freshly isolated mouse pancreatic acinar cells and cultured RAW264.7 macrophages. RESULTS: The main compounds from CQCQD belonged to flavonoids, iridoids, phenols, lignans, anthraquinones and corresponding glycosides. The sub-network analysis revealed that emodin, rhein, baicalin and chrysin were the compounds most relevant for directly regulating the TLR4/NLRP3-related proteins TLR4, RelA, NF-κB and TNF-α. In vivo, CQCQD attenuated the pancreatic injury and systemic inflammation of CER-AP and was associated with reduced expression of TLR4/NLRP3-related mRNAs and proteins. Emodin, rhein, baicalin and chrysin significantly diminished pancreatic acinar cell necrosis with varied effects on suppressing the expression of TLR4/NLRP3-related mRNAs. Emodin, rhein and chrysin also decreased nitric oxide production in macrophages and their combination had synergistic effects on alleviating cell death as well as expression of TLR4/NLRP3-related proteins. CONCLUSIONS: CQCQD attenuated the severity of AP at least in part by inhibiting the TLR4/NLRP3 pro-inflammatory pathways. Its active ingredients, emodin, baicalin, rhein and chrysin contributed to these beneficial effects.

Genome-Wide Identification and Functional Analysis of NADPH Oxidase Family Genes in Wheat During Development and Environmental Stress Responses.

As the key producers of reactive oxygen species (ROS), NADPH oxidases (NOXs), also known as respiratory burst oxidase homologs (RBOHs), play crucial roles in various biological processes in plants with considerable evolutionary selection and functional diversity in the entire terrestrial plant kingdom. However, only limited resources are available on the phylogenesis and functions of this gene family in wheat. Here, a total of 46 NOX family genes were identified in the wheat genome, and these NOXs could be classified into three subgroups: typical TaNOXs, TaNOX-likes, and ferric reduction oxidases (TaFROs). Phylogenetic analysis indicated that the typical TaNOXs might originate from TaFROs during evolution, and the TaFROs located on Chr 2 might be the most ancient forms of TaNOXs. TaNOXs are highly expressed in wheat with distinct tissue or organ-specificity and stress-inducible diversity. A large-scale expression and/or coexpression analysis demonstrated that TaNOXs can be divided into four functional groups with different expression patterns under a broad range of environmental stresses. Different TaNOXs are coexpressed with different sets of other genes, which widely participate in several important intracellular processes such as cell wall biosynthesis, defence response, and signal transduction, suggesting their vital but diversity of roles in plant growth regulation and stress responses of wheat.

Protective effects of flavonoids from Coreopsis tinctoria Nutt. on experimental acute pancreatitis via Nrf-2/ARE-mediated antioxidant pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Oxidative stress is a prominent feature of clinical acute pancreatitis (AP). Coreopsis tinctoria has been used traditionally to treat pancreas disorders like diabetes mellitus in China and Portugal and its flavonoid-rich fraction contain the main phytochemicals that have antioxidant and anti-inflammatory activities. AIM OF THE STUDY: To investigate the effects of flavonoids isolated from C. tinctoria on experimental AP and explore the potential mechanism. MATERIALS AND METHODS: LC-MS based online technique was used to analyse and isolate targeted flavonoids from C. tinctoria. Freshly isolated mouse pancreatic acinar cells were treated with taurocholic acid sodium salt hydrate (NaT, 5 mM) with or without flavonoids. Fluorescence microscopy and a plate reader were used to determine necrotic cell death pathway activation (propidium iodide), reactive oxygen species (ROS) production (H2-DCFDA) and ATP depletion (luminescence) where appropriate. AP was induced by 7 repeated intraperitoneal caerulein injections (50 µg/kg) at hourly interval in mice or retrograde infusion of taurolithocholic acid 3-sulfate disodium salt (TLCS; 5 mM, 50 µL) into the pancreatic duct in mice or infusion of NaT (3.5%, 1 mL/kg) in rats. A flavonoid was intraperitoneally administered at 0, 4, and 8 h after the first caerulein injection or post-operation. Disease severity, oxidative stress and antioxidant markers were determined. RESULTS: Total flavonoids extract and flavonoids 1-6 (C1-C6) exhibited different capacities in reducing necrotic cell death pathway activation with 0.5 mM C1, (2 R,3 R)-taxifolin 7-O-ß-D-glucopyranoside, having the best effect. C1 also significantly reduced NaT-induced ROS production and ATP depletion. C1 at 12.5 mg/kg and 8.7 mg/kg (equivalent to 12.5 mg/kg for mice) significantly reduced histopathological, biochemical and immunological parameters in the caerulein-, TLCS- and NaT-induced AP models, respectively. C1 administration increased pancreatic nuclear factor erythroid 2-related factor 2 (Nrf2) and Nrf2-medicated haeme oxygenase-1 expression and elevated pancreatic antioxidant enzymes superoxide dismutase and glutathione peroxidase levels. CONCLUSIONS: Flavonoid C1 from C. tinctoria was protective in experimental AP and this effect may at least in part be attributed to its antioxidant effects by activation of Nrf2-mediated pathways. These results suggest the potential utilisation of C. tinctoria to treat AP.

Mechanisms of Pancreatic Injury Induced by Basic Amino Acids Differ Between L-Arginine, L-Ornithine, and L-Histidine.

Pancreatic acinar cells require high rates of amino acid uptake for digestive enzyme synthesis, but excessive concentrations can trigger acute pancreatitis (AP) by mechanisms that are not well understood. We have used three basic natural amino acids L-arginine, L-ornithine, and L-histidine to determine mechanisms of amino acid-induced pancreatic injury and whether these are common to all three amino acids. Caffeine markedly inhibited necrotic cell death pathway activation in isolated pancreatic acinar cells induced by L-arginine, but not L-ornithine, whereas caffeine accelerated L-histidine-induced cell death. Both necroptosis inhibitors of RIPK1 and RIPK3 and a necroptosis activator/apoptosis inhibitor z-VAD increased cell death caused by L-histidine, but not L-arginine or L-ornithine. Cyclophilin D knock-out (Ppif-/-) significantly attenuated cell death induced by L-histidine, but not L-arginine, or L-ornithine. Allosteric modulators of calcium-sensing receptor (CaSR) and G-protein coupled receptor class C group 6 member A (GPRC6A) had inhibitory effects on cell death induced by L-arginine but not L-ornithine or L-histidine. We developed a novel amino acid-induced AP murine model with high doses of L-histidine and confirmed AP severity was significantly reduced in Ppif-/- vs. wild type mice. In L-arginine-induced AP neither Ppif-/-, caffeine, or allosteric modulators of CaSR or GPRC6A reduced pancreatic damage, even though CaSR inhibition with NPS-2143 significantly reduced pancreatic and systemic injury in caerulein-induced AP. These findings demonstrate marked differences in the mechanisms of pancreatic injury induced by different basic amino acids and suggest the lack of effect of treatments on L-arginine-induced AP may be due to conversion to L-ornithine in the urea cycle.