Exosomes derived from bone marrow mesenchymal stem cells alleviate biliary ischemia reperfusion injury in fatty liver transplantation by inhibiting ferroptosis.

Fatty liver grafts are susceptible to ischemia reperfusion injury (IRI), increasing the risk of biliary complications after liver transplantation (LT). Ferroptosis, a newly recognized programmed cell death, is expected to be a novel therapeutic target for IRI. We investigated whether exosomes derived from heme oxygenase 1-modified bone marrow mesenchymal stem cells (HExos) relieve ferroptosis and protect biliary tracts from IRI in a rat fatty liver transplantation model. Rats were fed with a methionine choline deficient (MCD) diet for 2 weeks to induce severe hepatic steatosis. Steatotic grafts were implanted and HExos were administered after liver transplantation. A series of functional assays and pathological analysis were performed to assess ferroptosis and biliary IRI. The HExos attenuated IRI following liver transplantation, as demonstrated by less ferroptosis, improved liver function, less Kupffer and T cell activation, and less long-term biliary fibrosis. MicroRNA (miR)-204-5p delivered by HExos negatively regulated ferroptosis by targeting a key pro-ferroptosis enzyme, ACSL4. Ferroptosis contributes to biliary IRI in fatty liver transplantation. HExos protect steatotic grafts by inhibiting ferroptosis, and may become a promising strategy to prevent biliary IRI and expand the donor pool.

Heme Oxygenase-1-Modified BMMSCs Activate AMPK-Nrf2-FTH1 to Reduce Severe Steatotic Liver Ischemia-Reperfusion Injury.

BACKGROUND: Ischemia-reperfusion injury (IRI) is an important cause of graft dysfunction post-liver transplantation, where donor liver with severe steatosis is more sensitive to IRI. Liver IRI involves ferroptosis and can be alleviated by heme oxygenase-1-modified bone marrow mesenchymal stem cells (HO-1/BMMSCs). AIMS: To explore the role and mechanism of HO-1/BMMSCs in severe steatotic liver IRI. METHODS: A severe steatotic liver IRI rat model and a hypoxia/reoxygenation (H/R) of severe steatosis hepatocyte model were established. Liver and hepatocyte damage was evaluated via liver histopathology and cell activity. Ferroptosis was evaluated through ferroptosis indexes. Nuclear factor erythroid 2-related factor 2 (Nrf2) was knocked down in severe steatotic hepatocytes. The role of Nrf2 and AMPK in HO-1/BMMSC inhibition of ferroptosis was examined using the AMP-activated protein kinase (AMPK) pathway inhibitor Compound C. RESULTS: The HO-1/BMMSCs alleviated severe steatotic liver IRI and ferroptosis. HO-1/BMMSCs promoted ferritin heavy chain 1(FTH1), Nrf2, and phosphorylated (p)-AMPK expression in the H/R severe steatotic hepatocytes. Nrf2 knockdown decreased FTH1 expression levels but did not significantly affect p-AMPK expression levels. The protective effect of HO-1/BMMSCs against H/R injury in severe steatotic hepatocytes and the inhibitory effect on ferroptosis were reduced. Compound C decreased p-AMPK, Nrf2, and FTH1 expression levels, weakened the HO-1/BMMSC protective effect against severe steatotic liver IRI and H/R-injured severe steatotic hepatocytes, and reduced the inhibition of ferroptosis. CONCLUSIONS: Ferroptosis was involved in HO-1/BMMSC reduction of severe steatotic liver IRI. HO-1/BMMSCs protected against severe steatotic liver IRI by inhibiting ferroptosis through the AMPK-Nrf2-FTH1 pathway. HO-1/BMMSCs activate AMPK, which activates Nrf2, promotes its nuclear transcription, then promotes the expression of its downstream protein FTH1, thereby inhibiting ferroptosis and attenuating severe steatotic liver IRI in rats. Glu: glutamic acid; Cys: cystine; GSH: glutathione; GPX4: glutathione peroxidase 4; HO-1/BMMSCs: HO-1-modified BMMSCs; Fer-1: ferrostatin-1; DFO: deferoxamine; FTH1: ferritin heavy chain1; p-AMPK: phosphorylated AMP-activated protein kinase; Nrf2: nuclear factor erythroid 2-related factor 2; IRI: ischemia-reperfusion injury; MCD: methionine-choline deficiency.

miR-124-3p delivered by exosomes from heme oxygenase-1 modified bone marrow mesenchymal stem cells inhibits ferroptosis to attenuate ischemia-reperfusion injury in steatotic grafts.

BACKGROUND: Steatotic livers tolerate ischemia-reperfusion injury (IRI) poorly, increasing the risk of organ dysfunction. Ferroptosis is considered the initiating factor of organ IRI. Heme oxygenase oxygen-1 (HO-1)-modified bone marrow mesenchymal stem cells (BMMSCs) (HO-1/BMMSCs) can reduce hepatic IRI; however, the role of ferroptosis in IRI of steatotic grafts and the effect of HO-1/BMMSCs-derived exosomes (HM-exos) on ferroptosis remain unknown. METHODS: A model of rat liver transplantation (LT) with a severe steatotic donor liver and a model of hypoxia and reoxygenation (H/R) of steatotic hepatocytes were established. Exosomes were obtained by differential centrifugation, and the differentially expressed genes (DEGs) in liver after HM-exo treatment were detected using RNA sequencing. The expression of ferroptosis markers was analyzed. microRNA (miRNA) sequencing was used to analyze the miRNA profiles in HM-exos. RESULTS: We verified the effect of a candidate miRNA on ferroptosis of H/R treated hepatocytes, and observed the effect of exosomes knockout of the candidate miRNA on hepatocytes ferroptosis. In vitro, HM-exo treatment reduced the IRI in steatotic grafts, and enrichment analysis of DEGs suggested that HM-exos were involved in the regulation of the ferroptosis pathway. In vitro, inhibition of ferroptosis by HM-exos reduced hepatocyte injury. HM-exos contained more abundant miR-124-3p, which reduced ferroptosis of H/R-treated cells by inhibiting prostate six transmembrane epithelial antigen 3 (STEAP3), while overexpression of Steap3 reversed the effect of mir-124-3p. In addition, HM-exos from cell knocked out for miR-124-3p showed a weakened inhibitory effect on ferroptosis. Similarly, HM-exo treatment increased the content of miR-124-3p in grafts, while decreasing the level of STEAP3 and reducing the degree of hepatic ferroptosis. CONCLUSION: Ferroptosis is involved in the IRI during LT with a severe steatotic donor liver. miR-124-3p in HM-exos downregulates Steap3 expression to inhibit ferroptosis, thereby attenuating graft IRI, which might be a promising strategy to treat IRI in steatotic grafts.

Relationship between the coprecipitation of phosphorus-on-calcite by submerged macrophytes and the phosphorus cycle in water.

To assess potential phosphorus removal, we utilized Potamogeton crispus to determine the effects of calcium addition on phosphorus removal. Plastic film was used to block material exchange between the overlying water and the sediment, and we compared the experimental results with long-term monitoring results of Yimeng Lake, which contained a dense population of P. crispus. The results revealed that the first 10-40 days constituted a period of rapid P decrease, as P. crispus could effectively remove the phosphorus in the water through coprecipitation of CaCO3-P. The treatment groups indicated that P. crispus released calcium into the overlying water, and after the addition of calcium ions, P. crispus showed increased phosphorus removal efficiency in the water. Total phosphorus (TP) and P/Ca content increased with increasing pH in the treatment groups, and the TP and pH declined as the calcium content increased in the treatment groups. Long-term field observations showed that the calcium-to-phosphorus ratio in the coprecipitates was dependent on the pH during the crystallization process. Thus, water calcium driven by P. crispus plays an important role in the phosphorus cycle of water, due to P. crispus assisted precipitation. This study revealed the effect of P. crispus on the water purification, the migration and transformation of Ca and P in sediment and overlying water under the condition of sediment calcium addition, so as to provide a theoretical basis for the ecological restoration of shallow lakes eutrophication.

Long non-coding RNAs HERH-1 and HERH-4 facilitate cyclin A2 expression and accelerate cell cycle progression in advanced hepatocellular carcinoma.

BACKGROUND: The advanced hepatocellular carcinoma (HCC), such as the recurrent tumor after liver transplantation (LT), is an obstacle of HCC treatment. The aim of this study was to discover the underlying mechanism of HCC progression caused by non-coding RNAs (ncRNAs). METHODS: To this end, we investigated the selected patient cohort of matching primary and recurrent HCC after receiving LT. The recurrent tumors after LT were regarded as clinical models of the advanced HCC. Microarrays were used to profile lncRNA and mRNA expression in HCC recurrent and primary tissue samples. The mRNA profile characteristics were analyzed by bioinformatics. Two cell lines, HepG2 and QGY-7703, were used as HCC cell models. The protein-coding potential, length, and subcellular location of the interested lncRNAs were examined by bioinformatics, Northern blot, fluorescent in situ hybridization (FISH), and quantitative RT-PCR (qRT-PCR) assays. HCC cell proliferation was detected by CCK-8, doubling time and proliferation marker gene quantitation assays. DNA replication during the cell cycle was measured by EdU/PI staining and flow cytometry analyses. Promoter activity was measured using a luciferase reporter assay. Interactions between DNA, RNA, and protein were examined by immunoprecipitation and pull-down assays. The miRNA-target regulation was validated by a fluorescent reporter assay. RESULTS: Both lncRNA and mRNA profiles exhibited characteristic alterations in the recurrent tumor cells compared with the primary HCC. The mRNA profile in the HCC recurrent tissues, which served as model of advanced HCC, showed an aberrant cell cycle regulation. Two lncRNAs, the highly expressed lncRNA in recurrent HCC (HERH)-1 and HERH-4, were upregulated in the advanced HCC cells. HERH-1/4 enhanced proliferation and promoted DNA replication and G1-S transition during the cell cycle in HCC cells. HERH-1 interacted with the transcription factor CREB1. CREB1 enhanced cyclin A2 (CCNA2) transcription, depending on HERH-1-CREB1 interaction. HERH-4 acted as an miR-29b/c sponge to facilitate CCNA2 protein translation through a competing endogenous RNA (ceRNA) pathway. CONCLUSIONS: The oncogenic lncRNA HERH-1/4 promoted CCNA2 expression at the transcriptional and post-transcriptional levels and accelerated cell cycle progression in HCC cells. The HERH-1-CREB1-CCNA2 and HERH-4-miR-29b/c-CCNA2 axes served as molecular stimuli for HCC advance.

Protective Effects of Bone Marrow Mesenchymal Stem Cells (BMMSCS) Combined with Normothermic Machine Perfusion on Liver Grafts Donated After Circulatory Death via Reducing the Ferroptosis of Hepatocytes.

BACKGROUND To improve the quality of liver grafts from extended-criteria donors donated after circulatory death (DCD), this study explored whether bone marrow mesenchymal stem cells (BMMSCs) combined with normothermic machine perfusion (NMP) have protective effects on DCD donor livers and the effects of ferroptosis in this procedure. MATERIAL AND METHODS Twenty-four male rat DCD donor livers were randomly and averagely divided into normal, static cold storage (SCS), NMP, and NMP combined with BMMSCs groups. Liver function, bile secretion, and pathological features of DCD donor livers were detected to evaluate the protective effects of NMP and BMMSCs on DCD donor livers. Hydrogen peroxide was used to induce an oxidative stress model of hepatocyte IAR-20 cells to evaluate the protective effects of BMMSCs in vitro. RESULTS Livers treated with NMP combined with BMMSCs showed better liver function, relieved histopathological damage, reduced oxidative stress injury and ferroptosis, and the mechanism of reduction was associated with downregulation of intracellular reactive oxygen species (ROS) and free Fe²âº levels. BMMSCs showed significant protective effects on the ultrastructure of DCD donor livers and ROS-induced injury to IAR-20 cells under electron microscopy. BMMSCs also significantly improved the expression level of microtubule-associated protein 1 light chain 3 (LC3)-II in both DCD donor livers and ROS-induced injured IAR-20 cells, including upregulating the expression of ferritin. CONCLUSIONS BMMSCs combined with NMP could reduce the level of ROS and free Fe²âº in oxidative stress damaged rat DCD donor livers, potentially reduce the ferroptosis in hepatocytes, and repair both morphology and function of DCD donor livers.

Bone marrow mesenchymal stem cells combine with normothermic machine perfusion to improve rat donor liver quality-the important role of hepatic microcirculation in donation after circulatory death.

Donation after circulatory death (DCD) can expand the donor pool effectively. A gap remains in outcome between DCD livers and living donor livers, warranting improved DCD liver quality and urgent resolution. Bone marrow mesenchymal stem cells (BMMSCs) can regulate immunity, participate in the anti-inflammatory response, and secrete cytokines. We investigated the effect of BMMSCs combined with normothermic machine perfusion (NMP) on DCD liver quality, and the role of microcirculation therein. Rat thoracic aortas were clipped to obtain DCD livers, and a rat NMP system was established. The DCD livers were grouped by preservation method: normal, static cold storage (SCS), NMP (P), and BMMSCs plus NMP (BP); storage time was up to 8 h. Liver function in outflow perfusate was detected by biochemical methods; liver tissue histopathology was observed by hematoxylin-eosin staining; hepatocyte ultrastructure was observed by transmission electron microscopy; hepatocyte apoptosis was detected by terminal deoxynucleotidyl transferase dUTP nick end labeling; liver microcirculation-related indicators were detected by immunofluorescence, immunohistochemistry, Western blotting, and enzyme-linked immunosorbent assay. Compared with SCS, P and BP significantly improved liver function and liver histological damage, reduced hepatocyte apoptosis, and repaired hepatocyte mitochondrial damage after 6 h in vitro. BP also significantly inhibited intrahepatic macrophage activation and intercellular adhesion, improved endothelial damage, and significantly improved endothelin 1-nitric oxide balance and microcirculation perfusion. In conclusion, BP can improve DCD liver microcirculation and quality. The mechanism may be the improvement of improve hepatic sinusoidal endothelial injury and microcirculation perfusion by inhibiting macrophage activation and intercellular adhesion.

Study of the protective effect on damaged intestinal epithelial cells of rat multilineage-differentiating stress-enduring (Muse) cells.

In this study, we determined whether multilineage-differentiating stress-enduring (Muse) cells exist in rat bone marrow and elucidated their effects on protection against the injury of intestinal epithelial cells associated with inflammation. Rat Muse cells were separated from bone marrow mesenchymal stem cells (BMMSCs) by trypsin-incubation stress. The group of cells maintained the characteristics of BMMSCs; however, there were high positive expression levels of stage-specific embryonic antigen-3 (SSEA-3; 75.6 ± 2.8%) and stage-specific embryonic antigen-1 (SSEA-1; 74.8 ± 3.1%), as well as specific antigens including Nanog, POU class 5 homeobox 1 (OCT 3/4), and SRY-box 2 (SOX 2). After inducing differentiation, α-fetoprotein (endodermal), α-smooth muscle actin and neurofilament medium polypeptide (ectodermal) were positive in Muse cells. Injuries of intestinal epithelial crypt cell-6 (IEC-6) and colorectal adenocarcinoma 2 (Caco-2) cells as models were induced by tumor necrosis factor-α stimulation in vitro. Muse cells exhibited significant protective effects on the proliferation and intestinal barrier structure, the underlying mechanisms of which were related to reduced levels of interleukin-6 (IL-6) and interferon-γ (IFN-γ), and the restoration of transforming growth factor-ß (TGF-ß) and IL-10 in the inflammation microenvironment. In summary, there were minimal levels of pluripotent stem cells in rat bone marrow, which exhibit similar properties to human Muse cells. Rat Muse cells could provide protection against damage to intestinal epithelial cells depending on their anti-inflammatory and immune regulatory functionality. Their functional impact was more obvious than that of BMMSCs.

Recurrent broad ligament leiomyosarcoma with pancreatic and thigh metastasis: a case report.

BACKGROUND: Leiomyosarcoma (LMS) is an uncommon mesenchymal neoplasm, which infrequently metastasizes to pancreas and thigh. Clinical presentation and imaging findings of metastatic broad ligament LMS are often nonspecific. Complete excision plays an important role in treatment of patients with localized LMS. CASE PRESENTATION: Here, we report a case of a 33-year-old woman with recurrent broad ligament LMS metastasizing to pancreas and thigh. Previously, she was diagnosed with broad ligament LMS and underwent hysterectomy, bilateral salpingo-oophorectomy. The disease-free interval was 2.5 years until metastases were found. Computerized tomography (CT) of abdomen and thighs, magnetic resonance imaging (MRI) of thighs and whole-body 18-fluorodeoxyglucose positron emission tomography - computed tomography (PET-CT) performed, revealed pancreatic and thigh metastasis. Ultrasonography-guided biopsy and histological examinations confirmed LMS at both the sites. Pancreatic metastasis was completely resected first. Then the patient underwent surgical resection of thigh metastasis when both chemotherapy and radiotherapy failed. She recovered well and remained free of disease recurrence in the 2 years follow-up. CONCLUSIONS: Though imaging lacks specificity, it is a valuable asset in assessing the burden of disease and characterizing lesions while histological examination with immunohistochemistry is helpful for the diagnosis of LMS. Complete surgical resection of all metastatic sites where-ever feasible should be strongly considered in a treated case of broad ligament LMS with a durable disease-free interval.

Immunomodulatory effects of bone marrow mesenchymal stem cells overexpressing heme oxygenase-1: Protective effects on acute rejection following reduced-size liver transplantation in a rat model.

Here we explore the T-lymphocyte suppressive and immunomodulatory effects of bone marrow mesenchymal stem cells (BMMSCs) overexpressing heme oxygenase-1 (HO-1) on acute rejection following reduced-size liver transplantation (RLT) in a rat model. The proliferation activity, cell cycle progression, secretion of proinflammatory cytokines, expression of CD25 and CD71 in lymphocytes, and activity of NK cells were found to be significantly lowered, and the proportion of regulatory T cells (Tregs) was found to be increased relative to BMMSCs when Adv-HO-1/BMMSCs were co-cultured with Con A ex vivo; secretion of anti-inflammatory cytokines was significantly higher. When treated with saline, BMMSCs or Adv-HO-1/BMMSCs, post-transplantation rats receiving Adv-HO-1/BMMSCs showed better median survival time, lower rejection activity index, higher anti-inflammatory cytokine levels, lower proinflammatory cytokine levels, more peripheral Tregs, and lower natural killer cell viability. These results suggest that HO-1 enhanced and prolonged the effects of BMMSCs on acute rejection following RLT, with immunomodulatory effects in which adaptive and innate immunity, as well as paracrine signaling, may play important roles.

Effect of heme oxygenase-1 transduced bone marrow mesenchymal stem cells on damaged intestinal epithelial cells in vitro.

In this study, we explored the effects of mesenchymal stem cells (MSCs) from bone marrow overexpressing heme oxygenase-1 (HO-1) on the damaged human intestinal epithelial barrier in vitro. Rat MSCs were isolated from bone marrow and transduced with rat HO-1 recombinant adenovirus (HO-MSCs) for stable expression of HO-1. Colorectal adenocarinoma 2 (Caco2) cells were treated with tumor necrosis factor-α (TNF-α) to establish a damaged colon epithelial model. Damaged Caco2 were cocultured with MSCs, Ad-MSCs, Ad-HO + MSCs or HO-MSCs. mRNA and protein expression of Zona occludens-1 (ZO-1) and human HO-1 and the release of cytokines were measured. ZO-1 and human HO-1 in Caco2 were significantly decreased after treatment with TNF-α; and this effect was reduced when coculture with MSCs from bone marrow. Expression of ZO-1 was not significantly affected by Caco2 treatment with TNF-α, Ad-HO, and MSCs. In contrast, ZO-1 and human HO-1 increased significantly when the damaged Caco2 was treated with HO-MSCs. HO-MSCs showed the strongest effect on the expression of ZO-1 in colon epithelial cells. Coculture with HO-MSCs showed the most significant effects on reducing the expression of IL-2, IL-6, IFN-γ and increasing the expression of IL-10. HO-MSCs protected the intestinal epithelial barrier, in which endogenous HO-1 was involved. HO-MSCs play an important role in the repair process by reducing the release of inflammatory cytokines and increasing the release of anti-inflammatory factors. These results suggested that HO-MSCs from bone marrow were more effective in repairing the damaged intestinal epithelial barrier, and the effectiveness of MSCs was improved by HO-1 gene transduction, which provides favorable support for the application of stem cell therapy in the intestinal diseases.

Mesenchymal stem cells improve the outcomes of liver recipients via regulating CD4+ T helper cytokines in rats.

BACKGROUND: Bone marrow mesenchymal stem cells (BMMSCs) exert immunosuppressive activities in transplantation. This study aimed to determine whether BMMSCs reduce acute rejection and improve outcomes of liver transplantation in rats. METHODS: Orthotopic liver transplantation from Lewis to Brown Norway rats was performed, which was followed by the infusion of BMMSCs through the penile superficial dorsal vein. Normal saline infusion was used as a control. Animals were sacrificed at 0, 24, 72, or 168 hours after BMMSCs infusion. Liver grafts, and recipient serum and spleen tissues were obtained. Histopathology, apoptosis, serum liver enzymes, serum cytokines, and circulating regulatory T (Treg), Th1, Th2 and Th17 cells were assessed at each time point. RESULTS: BMMSCs significantly attenuated acute rejection and improved the survival rate of allogeneic liver transplantation recipients. Liver enzymes and liver apoptosis were significantly alleviated. The levels of the Th1/Th2 ratio-associated cytokines such as IL-2 and IFN-gamma were significantly reduced and IL-10 was significantly increased. The levels of the Th17/Tregs axis-associated cytokines such as IL-6, IL-17, IL-23, and TNF-alpha were significantly reduced, whereas TGF-beta concentration was significantly increased. Moreover, flow cytometry analysis showed that the infusion of BMMSCs significantly increased Th2 and Treg cells and decreased Th1 and Th17 cells. CONCLUSION: BMMSCs had immunomodulatory effects, attenuated acute rejection and improved outcomes of allogeneic liver transplantation in rats by regulating the levels of cytokines associated with Th1/Th2 and Th17/Treg ratios.

Suppression of MicroRNA-203 improves survival of rat bone marrow mesenchymal stem cells through enhancing PI3K-induced cellular activation.

As a group of heterogeneous multipotent cells, mesenchymal stem cells (MSCs) have potential in treatment of a variety of clinical diseases. However, the low survival of the transplanted MSCs reduced their therapeutic effects. In this study, we revealed that rno-miR-203 suppressed activity and colony formation and enhanced apoptosis of the rat bone marrow-derived MSCs (BM-MSCs). Using bioinformatics analysis, we found a potential miR-203 binding site within rat phosphatidylinositol 3-kinase (PI3K) 3'UTR, and fluorescent reporter experiments validated the direct and negative regulation of PI3K expression by miR-203 through this site. Ectopic expression of PI3K rescued BM-MSCs from depressed activity induced by miR-203, and suppression of PI3K attenuated the increased BM-MSCs activity by miR-203 inhibitor treatment. Moreover, miR-203 blocking partly protected BM-MSCs from impairment caused by low nutrition. We conclude that inhibition of endogenous miR-203 elevated PI3K expression, which may strengthen PI3K/Akt pathway and promote BM-MSCs activity and survival. © 2014 IUBMB Life, 66(3):220-227, 2014.

Attenuation of early liver fibrosis by herbal compound "Diwu Yanggan" through modulating the balance between epithelial-to-mesenchymal transition and mesenchymal-to-epithelial transition.

BACKGROUND: Diwu Yanggan (DWYG) is a Chinese compound herbal preparation which consists of five Chinese herbs. This study investigates the preventative effects of DWYG on liver fibrosis induced by carbon tetrachloride (CCl4) and explores its possible mechanisms of action. METHODS: Liver fibrosis was induced in male Wistar rats by injecting a 50% CCl4/soybean oil solution subcutaneously twice a week for six weeks. After six weeks of treatment, serum aspartate transaminase (AST) and alanine transaminase (ALT) assay, liver tissue histological assessment and hepatic hydroxyproline assay were respectively carried out to examine the effects of DWYG on liver function and fibrosis degree. The impacts of DWYG on the expression levels of epithelial marker E-cadherin, mesenchymal marker Vimentin, transforming growth factor ß1 (TGF-ß1) and bone morphogenetic protein-7 (BMP-7) were further examined by quantitative real-time RT-PCR and Western blot analysis. In addition, the differences of Hedgehog (Hh) signaling pathway activity between DWYG-treated and DWYG-untreated fibrotic liver tissues were also evaluated by quantitative real-time RT-PCR and Western blot analysis. RESULTS: Upon DWYG treatment, the serum levels of ALT and AST, hepatic hydroxyproline content and the degree of fibrosis in CCl4-induced fibrotic model rats were dramatically declined. In accompany with the alleviation of the degree of fibrosis, DWYG treatment provoked the reversal of epithelial-to-mesenchymal transition (EMT) to mesenchymal-to-epithelial transition (MET) in the fibrotic liver tissues, which was characterized with the up-regulation expression of E-cadherin and down-regulation expression of Vimentin. Furthermore, we observed that the expression level of TGF-ß1 was reduced whereas the expression level of BMP-7 was enhanced in liver tissues of DWYG-treated rats, therefore the expression ratio of TGF-ß1/BMP-7 was dramatically decreased compared to CCl4-induced fibrosis model rats. In addition, quantitative real-time RT-PCR and Western blot analysis demonstrated that after DWYG treatment the expressions of Hh ligand Shh, receptor Smo and Ptc, and transcription factor Gli1 in CCl4-induced fibrotic liver tissues were dramatically repressed. CONCLUSIONS: DWYG demonstrates therapeutic potential to prevent liver fibrosis by modulating the balance between EMT and MET through reducing the expression ratio of TGF-ß1/BMP-7 and inhibiting the excessive activation of Hh signaling pathway.

Effects of continual burial by sediment on seedling emergence and morphology of Suaeda salsa in the coastal marsh of the Yellow River estuary, China.

A greenhouse study was conducted to determine the impacts of continual burial on seedling emergence and morphology of Suaeda salsa, a pioneer species in the coastal marsh of the Yellow River estuary. From May to June 2012, seeds of S. salsa were artificially buried to depths of 0 cm (no burial), 2 cm (burial of 1 mm d(-1)), 4 cm (burial of 2 mm d(-1)), 6 cm (burial of 3 mm d(-1)), 8 cm (burial of 4 mm d(-1)) and 10 cm (burial of 5 mm d(-1)) in plastic pots filled with unsterilized sediment. Results showed that the percent emergence of seedlings had a significantly negative correlation with continual burial depth (p < 0.001). A large percentage of seedlings emerged from 2, 4 and 6 cm burial depths, with the highest emergence (56.00 ± 6.60%) occurring from 2 cm depth. The shortest emergence time occurred at 4 cm burial depth and seeds buried at 10 cm depth took longer to emerge than those at other depths. At shallow or moderate burials, a stimulatory effect on seedling height, stem diameter, number and length of branch, taproot length and dry mass were observed. With increasing burial depth, root-mass and leaf-mass ratios generally increased while stem-mass ratio decreased. Sediment burial also stimulated part of the hypocotyl below the sediment to form adventitious roots, implying that S. salsa seedlings had a special adaptive strategy in response to the rapid and dynamic burial environment in the coastal marsh of the Yellow River estuary. The use of thin-layer continual burial (1-2 mm d(-1)) to promote the emergence of S. salsa seedlings in degraded marsh was feasible, and our study provided another way for the restoration of S. salsa marsh during the initial stage of seedling establishment and laid a good foundation for the scientific decision-making and management of restoration project at a large scale.

Ferrostatin-1 improves prognosis and regulates gut microbiota of steatotic liver transplantation recipients in rats.

Aims: To investigate the effects of Ferrostatin-1 (Fer-1) on improving the prognosis of liver transplant recipients with steatotic liver grafts and regulating gut microbiota in rats. Methods: We obtained steatotic liver grafts and established a liver transplantation model. Recipients were divided into sham, liver transplantation and Fer-1 treatment groups, which were assessed 1 and 7 days after surgery (n = 6). Results & conclusion: Fer-1 promotes recovery of the histological structure and function of steatotic liver grafts and the intestinal tract, and improves inflammatory responses of recipients following liver transplantation. Fer-1 reduces gut microbiota pathogenicity, and lowers iron absorption and improves fat metabolism of recipients, thereby protecting steatotic liver grafts.

Oleanolic acid alleviating ischemia-reperfusion injury in rat severe steatotic liver via KEAP1/NRF2/ARE.

Severe steatosis in donor livers is contraindicated for transplantation due to the high risk of ischemia-reperfusion injury (IRI). Although Ho-1 gene-modified bone marrow mesenchymal stem cells (HO-1/BMMSCs) can mitigate IRI, the role of gut microbiota and metabolites in this protection remains unclear. This study aimed to explore how gut microbiota and metabolites contribute to HO-1/BMMSCs-mediated protection against IRI in severe steatotic livers. Using rat models and cellular models (IAR20 and THLE-2 cells) of steatotic liver IRI, this study revealed that ischemia-reperfusion led to significant liver and intestinal damage, heightened immune responses, impaired liver function, and altered gut microbiota and metabolite profiles in rats with severe steatosis, which were partially reversed by HO-1/BMMSCs transplantation. Integrated microbiome and metabolome analyses identified gut microbial metabolite oleanolic acid as a potential protective agent against IRI. Experimental validation showed that oleanolic acid administration alone alleviated IRI and inhibited ferroptosis in both rat and cellular models. Network pharmacology and molecular docking implicated KEAP1/NRF2 pathway as a potential target of oleanolic acid. Indeed, OA experimentally upregulated NRF2 activity, which underlies its inhibition of ferroptosis and protection against IRI. The gut microbial metabolite OA protects against IRI in severe steatotic liver by promoting NRF2 expression and activity, thereby inhibiting ferroptosis.

Response of nutrient loss to natural erosive rainfall events under typical tillage practices of contour ridge system in the rocky mountain areas of Northern China.

Changes in natural rainfall characterized by heavy precipitation and high rainfall intensity would increase the risks and uncertainty of nutrients losses. Losses of nitrogen (N) and phosphorus (P) with water erosion from agriculture-related activities has become the principal nutrients resulting the eutrophication of water bodies. However, a little attention has been paid to the loss characteristic of N and P responding to natural rainfall in widely used contour ridge systems. To explore the loss mechanism of N and P in contour ridge system, nutrient loss associated with runoff and sediment yield was observed in in situ runoff plots of sweet potato (SP) and peanut (PT) contour ridges under natural rainfall. Rainfall events were divided into light rain, moderate rain, heavy rain, rainstorm, large rainstorm, and extreme rainstorm level, and rainfall characteristics for each rainfall level were recorded. Results showed that rainstorm, accounting for 46.27% of the total precipitation, played a destructive role in inducing runoff, sediment yield, and nutrient loss. The average contribution of rainstorm to sediment yield (52.30%) was higher than that to runoff production (38.06%). Rainstorm respectively generated 43.65-44.05% of N loss and 40.71-52.42% of P loss, although light rain induced the greatest enrichment value for total nitrogen (TN, 2.44-4.08) and PO4-P (5.40). N and P losses were dominated by sediment, and up to 95.70% of the total phosphorus and 66.08% of TN occurred in sediment. Nutrient loss exhibited the highest sensitivity to sediment yield compared to runoff and rainfall variables, and a significant positive linear relationship was observed between nutrient loss and sediment yield. SP contour ridge presented higher nutrient loss than that in PT contour ridge, especially for P loss. Findings gained in this study provide references for the response strategies of nutrient loss control to natural rainfall change in contour ridge system.

Small extracellular vesicles from HO-1-modified bone marrow-derived mesenchymal stem cells attenuate ischemia-reperfusion injury after steatotic liver transplantation by suppressing ferroptosis via miR-214-3p.

Donor shortage is a major problem that limits liver transplantation availability. Steatotic donor liver presents a feasible strategy to solve this problem. However, severe ischemia-reperfusion injury (IRI) is an obstacle to the adoption of steatotic transplanted livers. Evidence from our prior studies indicated that bone marrow mesenchymal stem cells modified with heme oxygenase-1 (HMSCs) can attenuate non-steatotic liver IRI. However, the contribution of HMSCs in transplanted steatotic liver IRI is unclear. Here, HMSCs and their derived small extracellular vesicles (HM-sEVs) alleviated IRI in transplanted steatotic livers. After liver transplantation, there was significant enrichment of the differentially expressed genes in the glutathione metabolism and ferroptosis pathways, accompanied by ferroptosis marker upregulation. The HMSCs and HM-sEVs suppressed ferroptosis and attenuated IRI in the transplanted steatotic livers. MicroRNA (miRNA) microarray and validation experiments indicated that miR-214-3p, which was abundant in the HM-sEVs, suppressed ferroptosis by targeting cyclooxygenase 2 (COX2). In contrast, COX2 overexpression reversed this effect. Knockdown of miR-214-3p in the HM-sEVs diminished its ability to suppress ferroptosis and protect liver tissues/cells. The findings suggested that HM-sEVs suppressed ferroptosis to attenuate transplanted steatotic liver IRI via the miR-214-3p-COX2 axis.

Potential ecological risks of heavy metals and Cd accumulation characteristics of Suaeda salsa under different Cd input and water logging conditions in the Yellow River estuary, China.

To improve the remediation of heavy metal pollution by typical wetland vegetation and maintain the health of wetland ecosystems under the water-sediment regulation scheme (WSRS) application, we evaluated the potential ecological risk of heavy metals in surface sediment in the Yellow River estuary affected by the WSRS. The ranges of Cr, Cu, Zn, Cd, and Pb content in surface sediment were 52.44-100.80 mg·kg-1 dry weight (DW), 16.38-21.19 mg·kg-1 DW, 64.77-255.50 mg·kg-1 DW, 0.12-0.24 mg·kg-1 DW, and 5.40-8.63 mg·kg-1 DW, respectively, and potential ecological risk coefficients showed that Cd was associated with moderate potential risk. We further examined effects of Cd in a greenhouse experiment to explore the influence of short-term Cd input and water logging condition changes induced by WSRS on the Cd absorption characteristics of Suaeda salsa (L.) Pall in the Yellow River estuary. The results showed that total biomass decreased but Cd content in tissue of S. salsa increased with increasing Cd input and the accumulation factor reached maximum values at 100 µg·L-1 of Cd, indicating that S. salsa efficiently accumulated Cd. Water logging depth significantly affected S. salsa growth and Cd absorption with deeper water logging being detrimental to growth. The interaction effect of Cd input and water logging depth on Cd content and accumulation factor was significant. These results suggest that WSRS caused short-term heavy metal input and changes in water conditions affect wetland vegetation growth and heavy metal absorption in the downstream estuary.