Opuntia ficus-indica seed attenuates hepatic steatosis and promotes M2 macrophage polarization in high-fat diet-fed mice.

Opuntia ficus-indica (L.) is a popular edible plant that possesses considerable nutritional value and exhibits diverse biological actions including anti-inflammatory and antidiabetic activities. In this study, we hypothesized that DWJ504, an extract of O ficus-indica seed, would ameliorate hepatic steatosis and inflammation by regulating hepatic de novo lipogenesis and macrophage polarization against experimental nonalcoholic steatohepatitis. Mice were fed a normal diet or a high-fat diet (HFD) for 10 weeks. DWJ504 (250, 500, and 1000 mg/kg) or vehicle (0.5% carboxymethyl cellulose) were orally administered for the last 4 weeks of the 10-week HFD feeding period. DWJ504 treatment remarkably attenuated HFD-induced increases in hepatic lipid content and hepatocellular damage. DWJ504 attenuated increases in sterol regulatory element-binding protein 1 and carbohydrate-responsive element-binding protein expression and a decrease in carnitine palmitoyltransferase 1A. Although DWJ504 augmented peroxisome proliferator-activated receptor α protein expression, it attenuated peroxisome proliferator-activated receptor γ expression. Moreover, DWJ504 promoted hepatic M2 macrophage polarization as indicated by attenuation of the M1 marker genes and enhancement of M2 marker genes. Finally, DWJ504 attenuated expression of toll-like receptor 4, nuclear factor κB, tumor necrosis factor α, interleukin 6, TIR-domain-containing adapter-inducing interferon ß, and interferon ß levels. Our results demonstrate that DWJ504 prevented intrahepatic lipid accumulation, induced M2 macrophage polarization, and suppressed the toll-like receptor 4-mediated inflammatory signaling pathway. Thus, DWJ504 has therapeutic potential in the prevention of nonalcoholic fatty liver disease.

Auxin-callose-mediated plasmodesmal gating is essential for tropic auxin gradient formation and signaling.

In plants, auxin functions as a master controller of development, pattern formation, morphogenesis, and tropic responses. A sophisticated transport system has evolved to allow the establishment of precise spatiotemporal auxin gradients that regulate specific developmental programs. A critical unresolved question relates to how these gradients can be maintained in the presence of open plasmodesmata that allow for symplasmic exchange of essential nutrients and signaling macromolecules. Here we addressed this conundrum using genetic, physiological, and cell biological approaches and identified the operation of an auxin-GSL8 feedback circuit that regulates the level of plasmodesmal-localized callose in order to locally downregulate symplasmic permeability during hypocotyl tropic response. This system likely involves a plasmodesmal switch that would prevent the dissipation of a forming gradient by auxin diffusion through the symplasm. This regulatory system may represent a mechanism by which auxin could also regulate symplasmic delivery of a wide range of signaling agents.

Losartan protects liver against ischaemia/reperfusion injury through PPAR-γ activation and receptor for advanced glycation end-products down-regulation.

BACKGROUND AND PURPOSE: PPAR-γ has been reported to be a protective regulator in ischaemia/reperfusion (I/R) injury. The receptor for advanced glycation end-products (RAGE) plays a major role in the innate immune response, and its expression is associated with PPAR-γ activation. Several angiotensin receptor blockers possess partial agonist activities towards PPAR-γ. Therefore, this study investigated the action of losartan, particularly with regard to PPAR-γ activation and RAGE signalling pathways during hepatic I/R. EXPERIMENTAL APPROACH: Mice were subjected to 60 min of ischaemia followed by 6 h of reperfusion. Losartan (0.1, 1, 3 and 10 mg · kg⁻¹) was administered 1 h prior to ischaemia and immediately before reperfusion. GW9662, a PPAR-γ antagonist, was administered 30 min prior to first pretreatment with losartan. KEY RESULTS: Losartan enhanced the DNA-binding activity of PPAR-γ in I/R. Losartan attenuated the increased serum alanine aminotransferase activity, TNF-α and IL-6 levels, and nuclear concentrations of NF-κB in I/R. GW9662 reversed these beneficial effects. Losartan caused a decrease in apoptosis as assessed by TUNEL assay, in release of cytochrome c and in cleavage of caspase-3, and these effects were abolished by GW9662 administration. Losartan attenuated not only I/R-induced RAGE overexpression, but also its downstream early growth response protein-1-dependent macrophage inflammatory protein 2 level; phosphorylation of p38, ERK and JNK; and subsequent c-Jun phosphorylation. GW9662 reversed these effects of losartan administration. CONCLUSIONS AND IMPLICATIONS: Our findings suggest that losartan ameliorates I/R-induced liver damage through PPAR-γ activation and down-regulation of the RAGE signalling pathway.

Agrimonia eupatoria protects against chronic ethanol-induced liver injury in rats.

This study examined the hepatoprotective effects of Agrimonia eupatoria water extract (AE) against chronic ethanol-induced liver injury. Rats were fed a Lieber-DeCarli liquid diet for 8 weeks. Animals were treated orally with AE at 10, 30, 100, and 300 mg/kg/day. After chronic consumption of ethanol, serum aminotransferase activities and pro-inflammatory cytokines markedly increased, and those increases were attenuated by AE. The cytochrome P450 2E1 activity and lipid peroxidation increased after chronic ethanol consumption, while reduced glutathione concentration decreased. Those changes were attenuated by AE. Chronic ethanol consumption increased the levels of Toll-like receptor 4 (TLR4) and myeloid differentiation factor 88 protein expression, inducible nitric oxide synthase and cyclooxygenase-2 protein and mRNA expression, and nuclear translocation of nuclear factor-kappa B, which was attenuated by AE. Our results suggest that AE ameliorates chronic ethanol-induced liver injury, and that protection is likely due to the suppression of oxidative stress and TLR-mediated inflammatory signaling.

Callose deposition in the phloem plasmodesmata and inhibition of phloem transport in citrus leaves infected with "Candidatus Liberibacter asiaticus".

Huanglongbing (HLB) is a destructive disease of citrus trees caused by phloem-limited bacteria, Candidatus Liberibacter spp. One of the early microscopic manifestations of HLB is excessive starch accumulation in leaf chloroplasts. We hypothesize that the causative bacteria in the phloem may intervene photoassimilate export, causing the starch to over-accumulate. We examined citrus leaf phloem cells by microscopy methods to characterize plant responses to Liberibacter infection and the contribution of these responses to the pathogenicity of HLB. Plasmodesmata pore units (PPUs) connecting companion cells and sieve elements were stained with a callose-specific dye in the Liberibacter-infected leaf phloem cells; callose accumulated around PPUs before starch began to accumulate in the chloroplasts. When examined by transmission electron microscopy, PPUs with abnormally large callose deposits were more abundant in the Liberibacter-infected samples than in the uninfected samples. We demonstrated an impairment of symplastic dye movement into the vascular tissue and delayed photoassimilate export in the Liberibacter-infected leaves. Liberibacter infection was also linked to callose deposition in the sieve plates, which effectively reduced the sizes of sieve pores. Our results indicate that Liberibacter infection is accompanied by callose deposition in PPUs and sieve pores of the sieve tubes and suggest that the phloem plugging by callose inhibits phloem transport, contributing to the development of HLB symptoms.

A combination of ischemic preconditioning and allopurinol protects against ischemic injury through a nitric oxide-dependent mechanism.

This study examined the cytoprotective mechanisms of a combination of ischemic preconditioning (IPC) and allopurinol against liver injury caused by ischemia/reperfusion (I/R). Allopurinol (50mg/kg) was intraperitoneally administered 18 and 1h before sustained ischemia. A rat liver was preconditioned by 10 min of ischemia, followed by 10 min of reperfusion, and then subjected to 90 min of ischemia, followed by 5h of reperfusion. Rats were pretreated with adenosine deaminase (ADA), 3,7-dimethyl-1-[2-propargyl]-xanthine (DMPX), and N-nitro-l-arginine methyl ester (l-NAME) before IPC. Hepatic nitrite and nitrate and eNOS protein expression levels were increased by the combination of IPC and allopurinol. This increase was attenuated by ADA, DMPX, and l-NAME. I/R induced an increase in alanine aminotransferase activity, whereas it decreased the hepatic glutathione level. A combination of IPC and allopurinol attenuated these changes, which were abolished by ADA, DMPX, and l-NAME. The increase in the liver wet weight-to-dry weight ratio after I/R was attenuated by the combination of IPC and allopurinol. In contrast, hepatic bile flow was decreased after I/R, which was attenuated by the combination of IPC and allopurinol. These changes were restored by l-NAME. I/R induced a decrease in the level of mitochondrial dehydrogenase, whereas it increased mitochondrial swelling. A combination of IPC and allopurinol attenuated these changes, which were restored by ADA, DMPX, and l-NAME. Our findings suggest that a combination of IPC and allopurinol reduces post-ischemic hepatic injury by enhancing NO generation.

Melatonin protects liver against ischemia and reperfusion injury through inhibition of toll-like receptor signaling pathway.

This study investigated the immunomodulating effect of melatonin on toll-like receptor (TLR)-stimulated signal transduction. Rats were subjected to 60 min of ischemia followed by 1 or 5 hr of reperfusion. Melatonin (10 mg/kg) or the vehicle was administered intraperitoneally 15 min prior to ischemia and immediately before reperfusion. Melatonin treatment significantly reduced the level of serum alanine aminotransferase activity. Increased levels of TLR3 and TLR4 protein expression induced by ischemia/reperfusion (I/R) were attenuated by melatonin. Serum level of high-mobility group box 1 (HMGB1), a potent alarmin of the TLR system, increased significantly in the I/R group, and melatonin inhibited this release. Melatonin suppressed the increase in myeloid differentiation factor 88 (MyD88) protein expression, extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) phosphorylation and nuclear translocation of nuclear factor κB (NF-κB) and phosphorylated c-Jun, a component of activator protein 1. The increased level of toll-receptor-associated activator of interferon (TRIF) expression, phosphorylation of interferon (IFN) regulatory factor 3 (IRF3) and serum IFN-ß was attenuated by melatonin. Melatonin attenuated the levels of tumor necrosis factor alpha (TNF-α), interleukin (IL)-6 and inducible nitric oxide synthase (iNOS) protein and mRNA expression, while the level of heme oxygenase-1 (HO-1) was augmented. Our results suggest that melatonin ameliorates I/R-induced liver damage by modulation of TLR-mediated inflammatory responses.

Mutation of the chitinase-like protein-encoding AtCTL2 gene enhances lignin accumulation in dark-grown Arabidopsis seedlings.

Several genes that encode a chitinase-like protein (called the CTL group) have been identified in Arabidopsis, rice, pea, and cotton. Members of the CTL group have attracted much attention because of their possible role in the biosynthesis of the cell wall in plants. The hot2 mutation in the CTL1 (AtCTL1) gene of Arabidopsis thaliana causes multiple defects in growth and development. The Arabidopsis genome possesses the AtCTL2 gene, which exhibits 70% similarity to AtCTL1 at the amino acid level. We showed that the AtCTL2 gene was predominantly expressed in stems, which was in contrast to the presence of AtCTL1 transcripts in most organs of Arabidopsis. In addition, beta-glucuronidase (GUS) staining was detectable in all tissues of the stem in transgenic plants expressing the AtCTL1::GUS construct, while GUS activity under control of the AtCTL2 promoter was significantly restricted to the xylem and to interfascicular fibers in stems. The phenotypes of atctl2 single mutant and of hot2, atctl2 double mutant plants were significantly similar to those of wild-type and of hot2 single mutant plants, respectively. The expression levels of CESA1 and CESA4 transcripts were not affected in the two single mutants or corresponding double mutant plants, compared with the levels in wild-type plants. The accumulation of lignin in etiolated hypocotyls, however, was increased by mutation of AtCTL2. These findings suggest that AtCTL2 is required for proper cell wall biosynthesis in etiolated seedlings of Arabidopsis.

Altered ARA2 (RABA1a) expression in Arabidopsis reveals the involvement of a Rab/YPT family member in auxin-mediated responses.

Ras super family proteins serve as molecular switches regulating many different cellular processes. However, given the large number of family members, sequence information has provided little insight into the function of individual proteins. This study examined phenotypic alterations in an Arabidopsis ara2 mutant, in which a Ras super family member-encoding gene is disrupted by a T-DNA insertion. Although one mutant line (Salk_013811) was hypersensitive to auxin, its T-DNA insertion was in the 5'-UTR of ARA2. Thus, we examined a true ARA2 knock-out mutant (Salk_077747) which contains an insertion in the first exon of ARA2. We found that ARA2 expression is responsive to auxin and at low concentrations, ara2 mutant plants exhibit increased numbers of lateral roots. ARA2 overexpression causes plants to exhibit hypersensitivity to auxin, due to altered expression of auxin-responsive genes, and these plants exhibited reduced numbers of lateral roots. A GFP-ARA2 fusion protein localized to the endosomes, suggesting that ARA2 may play a role in vesicle trafficking of components involved in polar auxin transport. Taken together, these results show that ARA2 is an essential component of a pathway that couples auxin signaling to plant growth and development.

Glucosamine causes overproduction of reactive oxygen species, leading to repression of hypocotyl elongation through a hexokinase-mediated mechanism in Arabidopsis.

Glucosamine (GlcN) is a naturally occurring amino-sugar that is synthesized by amidation of fructose-6-phosphate. Although a number of reports have examined the biological effects of GlcN on insulin resistance in mammalian systems, little is known about its effects on plant growth. In this study, we have shown that exogenous GlcN inhibits hypocotyl elongation in Arabidopsis, whereas glucose and its analogs alleviate this inhibitory effect. The hexokinase (HXK)-specific inhibitor mannoheptulose also restored hypocotyl elongation. The gin2-1 mutants with an alteration in AtHXK1 exhibited higher tolerance to GlcN. We also found that GlcN induces a significant increase in the production of reactive oxygen species (ROS). In addition, the GlcN-mediated inhibition of hypocotyl elongation was relieved by reducing agents such as ascorbic acid and glutathione. GlcN treatment resulted in significant induction of expression of GST1, GST2 and GST6, which are marker genes for ROS production. The gin2 mutation also represses the ROS production and the GST2 induction by GlcN treatment. Taken together, these results provide evidence that GlcN induces HXK-mediated induction of oxidative stress, leading to growth repression in Arabidopsis thaliana.

The ABA effect on the accumulation of an invertase inhibitor transcript that is driven by the CAMV35S promoter in ARABIDOPSIS.

Invertase (beta-D-fructofuranosidase; EC 3.2.1.26) catalyzes the conversion of sucrose into glucose and fructose and is involved in an array of important processes, including phloem unloading, carbon partitioning, the response to pathogens, and the control of cell differentiation and development. Its importance may have caused the invertases to evolve into a multigene family whose members are regulated by a variety of different mechanisms, such as pH, sucrose levels, and inhibitor proteins. Although putative invertase inhibitors in the Arabidopsis genome are easy to locate, few studies have been conducted to elucidate their individual functions in vivo in plant growth and development because of their high redundancy. In this study we assessed the functional role of the putative invertase inhibitors in Arabidopsis by generating transgenic plants harboring a putative invertase inhibitor gene under the control of the CaMV35S promoter. A transgenic plant that expressed high levels of the putative invertase inhibitor transcript when grown under normal conditions was chosen for the current study. To our surprise, the stability of the invertase inhibitor transcripts was shown to be down-regulated by the phytohormone ABA (abscisic acid). It is well established that ABA enhances invertase activity in vivo but the underlying mechanisms are still poorly understood. Our results thus suggest that one way ABA regulates invertase activity is by down-regulating its inhibitor.

A possible role for NDPK2 in the regulation of auxin-mediated responses for plant growth and development.

Auxin plays many crucial roles in the course of plant growth and development, such as hook opening, leaf expansion and inhibition of mesocotyl elongation. Although its mechanism of action has not been clarified at the molecular level, recent studies have indicated that auxin triggers the induction of a number of genes known as primary auxin-responsive genes. Hence, the identification of the regulatory components in auxin-mediated cellular responses would help to elucidate the mechanism of the action of this hormone in plant growth and development. NDPK2 encodes a nucleoside diphosphate kinase 2 (NDPK2) in Arabidopsis. We aim to elucidate the possible role of NDPK2 in auxin-related cellular processes, in view of the finding that a ndpk2 mutant displays developmental defects associated with auxin. Interestingly, the ndpk2 mutant exhibits defects in cotyledon development and increased sensitivity to an inhibitor of polar auxin transport (naphthylphthalamic acid; NPA). Consistent with this phenotype, the transcript levels of specific auxin-responsive genes were reduced in the ndpk2 mutant plants treated with auxin. The amount of auxin transported from the shoot apex to the shoot/root transition zone of ndpk2 mutant plants was increased, compared with that in the wild-type plants. These results collectively suggest that NDPK2 appears to participate in auxin-regulated processes, partly through the modulation of auxin transport.