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The effect of skeletal muscle fat deposition on the prognosis of patients with chronic liver disease remains unclear. Skeletal muscle fat deposition can be estimated by attenuation of skeletal muscle in Hounsfield units (HU) on computed tomography (CT). The aim of this retrospective cohort study was to investigate the association between skeletal muscle fat deposition assessed by skeletal muscle attenuation (SMA), and hepatocellular carcinoma (HCC).We enrolled 288 patients with chronic liver disease (139 men, 149 women; mean age 67.5 ± 10.4 y; hepatitis C virus, 239; hepatitis B virus, 17; without viral infection, 32; chronic hepatitis, 227; and cirrhosis, 61) who underwent liver biopsy and CT scanning between January 2013 and February 2017. The patients were divided into two groups based on SMA levels, with the cutoff value of 31 HU. We analyzed the effect of SMA on HCC development.During the study follow-up period (median, 2.50 y; range, 0.5-4.7 y), HCC was identified in 19 patients (7%). The cumulative incidence of HCC in patients with lower SMA (31 HU) was significantly higher than in patients with SMA ≥31 HU (P = 0.007). Cox proportional hazards regression analysis confirmed cirrhosis (hazard ratio [HR], 6.626; 95% confidence interval [CI], 2.57-17.12; P 0.001) and lower SMA (HR, 3.502; 95% CI, 1.25-9.83; P = 0.017) as significant independent factors associated with HCC development in patients with chronic liver disease.Patients with cirrhosis and skeletal muscle fat deposition assessed by SMA had a higher risk for developing HCC.

Background Myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs) are attractive immune cells to induce immune tolerance. To explore a strategy for improving the efficacy of MDSC therapies, we examined the impact of adoptive transfer of several types of MDSCs on graft rejection in a murine heart transplantation model. Methods We analyzed the effects of induced syngeneic and allogeneic bone marrow-derived MDSCs (BM-MDSCs) on graft survival and suppressive capacity. We also compared the ability of syngeneic monocytic MDSCs (Mo-MDSCs) and polymorphonuclear MDSCs (PMN-MDSCs) to inhibit graft rejection and investigated the suppression mechanisms. Results Both syngeneic and allogeneic donor- or allogeneic third-party-derived BM-MDSCs prolonged graft survival, although syngeneic BM-MDSCs inhibited anti-donor immune responses most effectively in vitro. Syngeneic Mo-MDSCs, rather than PMN-MDSCs, were responsible for immune suppression through downregulating inducible nitric oxide synthase (iNOS) and expanded naturally occurring thymic originated Treg (nTreg) in vitro. Adoptive transfer of Mo-MDSCs, but not PMN-MDSCs, prolonged graft survival and increased Treg infiltration into the graft heart. Conclusion Recipient-derived Mo-MDSCs are most effective in prolonging graft survival via inhibiting T cell response and nTreg infiltration.

Background & Aims Mesalamine is a first-line drug for treatment of inflammatory bowel diseases (IBD). However, its mechanisms are not fully understood. CD4+ Foxp3+ regulatory T cells (Tregs) play a potential role in suppressing IBD. This study determined whether the anti-inflammatory activity of mesalamine is related to Treg induction in the colon. Methods We examined the frequencies of Tregs in the colons of wild-type mice, mice deficient for aryl hydrocarbon receptor (AhR-/- mice), and bone marrow–chimeric mice lacking AhR in hematopoietic cells (BM-AhR-/- mice), following oral treatment with mesalamine. We also examined the effects of mesalamine on transforming growth factor (TGF)-β expression in the colon. Results Treatment of wild-type mice with mesalamine increased the accumulation of Tregs in the colon and up-regulated the AhR target gene Cyp1A1, but this effect was not observed in AhR-/- or BM-AhR-/- mice. In addition, mesalamine promoted in vitro differentiation of naive T cells to Tregs, concomitant with AhR activation. Mice treated with mesalamine exhibited increased levels of the active form of TGF-β in the colon in an AhR-dependent manner and blockade of TGF-β signaling suppressed induction of Tregs by mesalamine in the colon. Furthermore, mice pretreated with mesalamine acquired resistance to dextran sodium sulfate–induced colitis. Conclusions We propose a novel anti-inflammatory mechanism of mesalamine for colitis: induction of Tregs in the colon via the AhR pathway, followed by TGF-β activation., Graphical abstract

Nuclear factor erythroid 2-related factor 2 (NRF2) is a transcription factor that plays a crucial role in protection of cells from electrophile-induced toxicity through up-regulating phase II detoxifying enzymes and phase III transporters. We previously reported that oxidative stress induces up-regulation of interleukin-11 (IL-11), a member of the IL-6 family that ameliorates acetaminophen-induced liver toxicity. However, a role for IL-11 in protection of cells from electrophile-induced toxicity remains unclear. Here we show that an environmental electrophile, 1,2-naphthoquinone (1,2-NQ), but not 15d-prostaglandin J2 (PGJ2) or tert-butylhydroxyquinone (tBHQ), induced IL-11 production. Consistent with a crucial role for prolonged ERK activation in H2O2-induced IL-11 production, 1,2-NQ, but not 15d-PGJ2 or tBHQ, elicited prolonged ERK activation. Conversely, inhibition of the ERK pathway by a MEK inhibitor completely blocked 1,2-NQ-induced IL-11 production at both protein and mRNA levels, further substantiating an intimate cross-talk between ERK activation and 1,2-NQ-induced IL-11 production. Promoter analysis of the Il11 gene revealed that two AP-1 sites were essential for 1,2-NQ-induced promoter activities. Among various members of the AP-1 family, Fra-1 was up-regulated by 1,2-NQ, and its up-regulation was blocked by a MEK inhibitor. Although NRF2 was not required for H2O2-induced IL11 up-regulation, NRF2 was essential for 1,2-NQ-induced IL11 up-regulation by increasing Fra-1 proteins possibly through promoting mRNA translation of FOSL1. Finally, intraperitoneal administration of 1,2-NQ induced body weight loss in wild-type mice, which was further exacerbated in Il11ra1−/− mice compared with Il11ra1+/− mice. Together, both Fra-1 and NRF2 play crucial roles in IL-11 production that protects cells from 1,2-NQ intestinal toxicity.

Objective Cigarette smoking is an important risk factor for periodontal disease. The aim of this study is to evaluate the effect of nicotine, a major component of cigarette smoke, on interleukin-8 (IL-8) production and cellular signalling via nicotinic acetylcholine receptors (nAChRs) in human gingival epithelial cells (HGECs). Design Messenger RNA (mRNA) expression of nAChR subunits in three different HGEC lines (epi 4, Tfx and E6E7) was assessed using reverse transcription-polymerase chain reaction (RT-PCR). HGECs were stimulated by 1 × 10 −3 M nicotine in the presence or absence of IL-1β or Porphyromonas gingivalis lipopolysaccharide (LPS). IL-8 production was then examined using real-time PCR and enzyme-linked immunosorbent assay. Nicotine-mediated signalling in the epi 4 cell line was also evaluated by Western blotting. Results HGECs expressed several nAChR subunits. Nicotine increased the secretion of IL-8 from HGECs that were cultured in the presence of IL-1β or P. gingivalis LPS and also induced the phosphorylation of extracellular signal-regulated kinase (ERK) in epi 4. Pretreatment with non-selective nAChR antagonist or intracellular calcium chelator reduced the nicotine-induced phosphorylation of ERK. Furthermore, nicotine-induced IL-8 secretion was decreased by pretreatment with non-selective nAChR antagonist, ERK1/2 inhibitor or intracellular calcium chelator. Conclusion These findings indicate that nicotine increases IL-8 production in gingival epithelial cells via ERK phosphorylation following Ca 2+ signalling after nAChR activation.

We examined the effects of nicotine on differentiation and function of monocyte-derived human dendritic cells (DCs). NiDCs, which were the DCs differentiated in the presence of nicotine, showed lower levels of CD1a. Secretion of IL-12 and TNF-α by lipopolysaccharide (LPS)-stimulated NiDCs was significantly suppressed compared to monocyte-derived DCs grown without nicotine. NiDCs displayed a diminished capacity to induce allogeneic T cell proliferation with a reduced production of IFN-γ, and maintained/enhanced LPS-mediated expression of coinhibitory molecules. Interestingly, NiDCs enhanced the expression of nuclear receptor peroxisome proliferator-activated receptors γ (PPAR γ), which has immunomodulatory properties. Expression of PPAR γ and PPAR γ-target genes was significantly inhibited by pretreatment with d-tubocurarine, antagonist of non-selective nicotinic acetylcholine receptors (nAChR). In addition, reduction of Th1 responses was inhibited after blocking nAChR-mediated signal. These data suggest the effect of nicotine on altering DC immunogenicity by impeding Th1 immunity is partially mediated by upregulation of PPAR γ.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)/death receptor 5 (DR5)-mediated cell death plays an important role in the elimination of tumor cells and transformed cells. Recently, recombinant TRAIL and agonistic anti-DR5 monoclonal antibodies have been developed and applied to cancer therapy. However, depending on the type of cancer, the sensitivity to TRAIL has been reportedly different, and some tumor cells are resistant to TRAIL-mediated apoptosis. Using confocal microscopy, we found that large amounts of DR5 were localized in the nucleus in HeLa and HepG2 cells. Moreover, these tumor cells were resistant to TRAIL, whereas DU145 cells, which do not have nuclear DR5, were highly sensitive to TRAIL. By means of immunoprecipitation and Western blot analysis, we found that DR5 and importin β1 were physically associated, suggesting that the nuclear DR5 was transported through the nuclear import pathway mediated by importin β1. Two functional nuclear localization signals were identified in DR5, the mutation of which abrogated the nuclear localization of DR5 in HeLa cells. Moreover, the nuclear transport of DR5 was also prevented by the knockdown of importin β1 using siRNA, resulting in the up-regulation of DR5 expression on the cell surface and an increased sensitivity of HeLa and HepG2 cells to TRAIL. Taken together, our findings suggest that the importin β1-mediated nuclear localization of DR5 limits the DR5/TRAIL-induced cell death of human tumor cells and thus can be a novel target to improve cancer therapy with recombinant TRAIL and anti-DR5 antibodies.

Background Abdominal aortic aneurysm (AAA) accumulates features of a chronic inflammatory disorder and irreversible destruction of connective tissue. A recent experimental study identified c-Jun N terminal kinase (JNK) as a proximal signaling molecule in the pathogenesis of AAA and vascular dendritic cells as key players in the inflammatory reaction and degradation of the extracellular matrix. Statins can inhibit cell proliferation and vascular inflammation, which might help prevent AAA progression. However, supporting clinical data from human studies are lacking. We hypothesized that atorvastatin might inhibit JNK and dendritic cells, resulting in suppression of inflammatory cells and matrix metalloproteinases (MMPs) in human tissue of AAA. Methods Patients with AAA were randomized to atorvastatin (20 mg/day, n = 10) and non-treated ( n = 10) groups. After treatment for 4 weeks, patients underwent abdominal aorta replacement, tissue specimens were obtained, and tissue composition was assessed using immunohistochemistry with quantitative image analysis. Results Atorvastatin significantly reduced expression of JNK (1.1% vs. 8.1%, P = 0.0002) and dendritic cells (3.2 vs. 7.2, P = 0.003) compared to controls. T cells (142 vs. 315, P = 0.008), macrophages (13 vs. 24, P = 0.048) and immunoreactivity to MMP-2 (7.8% vs. 21%, P = 0.049) and MMP-9 (13% vs. 24%, P = 0.028) were also suppressed in the atorvastatin group. Serum low-density lipoprotein cholesterol level was decreased by 40% in the atorvastatin group. Conclusions Atorvastatin treatment acutely reduces JNK expression and dendritic cells, resulting in reduced inflammatory cell content and expression of MMPs in the AAA wall.

Background Full-thickness skin defects with exposed bone are often hard to heal. The lack or delayed re-vascularization is considered one of the major causes, and the periosteum is also suggested to have an important role in tissue regeneration. Materials and methods Full-thickness skin defect wounds with exposed bone were made in the parietal region of Wister rats. The periosteum of the exposed parietal bone was removed in the periosteum-lacking group, but maintained in the control group (periosteum-intact group). The wound was covered by an artificial dermis made of collagen. The wound healing process was histologically compared. Double immunostaining of α-smooth muscle actin (SMA) and von Willebrand factor (vWF) was used for re-vascularization examination, and the blood vessel density in the artificial dermis was quantified. Results The density of the blood vessels in the uninjured parietal tissue was approximately 80 vessels/mm 2 . To reach this density, 7 and 21 days were required for the control (periosteum-intact) and the periosteum-lacking groups, respectively. This coincided with complete revascularization, fibroblast migration and the reentry of blood vessels to the upper layer of the wound were observed. Conclusion The described results support the importance of the periosteum in the full-thickness skin defect healing process.

We showed previously that adult small bowel could be transplanted successfully in rats without vascular reconstruction by removing the graft serosa. In this study, we assessed if granulocyte colony-stimulating factor (G-CSF) or basic fibroblast growth factor (bFGF) could improve graft survival in the same rat model.A 10-mm-long adult small bowel graft from an adult 12-week-old Lewis rat was transplanted into a pouch created in the omentum of a 5-week-old Lewis rat (syngeneic bowel transplantation [SBTx], n = 49). Graft serosa was removed just before SBTx in the serosectomy group (n = 29) and left intact in the nonserosectomy group (n = 20). Each group was divided into 3 subgroups (sG): sG-1 had no G-CSF or bFGF; sG-2 had daily subcutaneous injections of G-CSF; and sG-3 had continuous infusion of bFGF around the graft in the omentum. All grafts were harvested 14 days after SBTx and studied histologically. A mucosal surface expansion score (MSES) was used where 0 = no mucosa on the graft, 1 = mucosa on one fourth of the graft, 2 = mucosa on one half of the graft, 3 = mucosa on three fourths of the graft, and 4 = mucosa on the whole graft. The density of CD34-positive capillaries per 1000 nuclei was also measured.Serosectomy group MSES were significantly higher than nonserosectomy group MSES indicating that grafts survived (P.0001). CD34-positive capillaries in serosectomy group subgroups for mucosa were 103.9 +/- 34.2, 130.2 +/- 52.0, and 132.3 +/- 37.7, respectively; for muscle, 74.4 +/- 38.0, 86.2 +/- 32.9, and 82.4 +/- 30.3, respectively; and for omentum, 73.8 +/- 30.1, 151.3 +/- 60.3, and 140.0 +/- 49.0, respectively. Mucosal surface expansion score and overall CD34-positive capillaries for sG-2 and sG-3 were significantly higher than for sG-1 (both, P.05).Our results suggest that G-CSF and bFGF enhance angiogenesis and mucosal surface expansion.

The aim of this study was to assess whether adult small bowel grafts (ASBGs) can survive transplantation without vascular reconstruction if graft serosectomy (SS) is performed.Syngeneic ASBG transplants were performed in 85 Lewis rats. The entire serosa was removed just before transplantation in the SS group (n = 50) and left intact in the nonserosectomy group (n = 35). Transplanted ASBG was harvested 1, 3, 5, 7, 14, 21, or 28 days after transplantation and studied using staining with hematoxylin-eosin, immunohistochemistry for protein gene product 9.5, S-100, CD34 and vascular endothelial growth factor (VEGF), and quantification of VEGF messenger RNA (mRNA). Adult small bowel graft viability was assessed blindly using a mucosal surface expansion score (0, no mucosa; 1, mucosa on one fourth of graft; 2, mucosa on one half of graft; 3, mucosa on three fourths of graft; and 4, circumferential mucosa on graft).No rejection was identified in any ASBG. Average mucosal surface expansion score and VEGF mRNA expression were significantly higher in the SS group (both P.01). Vascular endothelial growth factor protein was detected in enterocytes from day 3 posttransplant in the SS group. Distribution of protein gene product 9.5 and S-100 was normal in SS-group ASBG.Our results suggest that SS allows VEGF mRNA and, subsequently, VEGF protein in ASBG to be induced very soon after transplantation, which may contribute to the survival of ASBG transplanted without vascular reconstruction.

Smad3 deficiency attenuates renal fibrosis, inflammation, and apoptosis after unilateral ureteral obstruction. Background Transforming growth factor-β (TGF-β) has been implicated in the development of renal fibrosis induced by unilateral ureteral obstruction (UUO). However, there is little information on signaling pathways mediating TGF-β activity involved in molecular and cellular events leading to renal fibrosis induced by UUO. In this study, we sought to determine whether Smad3, a major signaling component of TGF-β, mediated renal fibrosis induced by UUO. Methods Renal fibrosis, inflammation, and apoptosis induced by UUO were macroscopically and histologically compared between wild-type mice and Smad3 null mice. Results Gross appearance of the kidney after UUO showed relatively intact kidney in Smad3 null mice [Smad3(-/-) mice] when compared with that of wild-type mice [Smad3(+/+) mice]. Renal interstitial fibrosis based on the interstitial area stained with Aniline-blue or Sirius red solution was significantly attenuated in the obstructed kidney of Smad3(-/-) mice when compared with that of Smad3(+/+) mice. Deposition of type I and type III collagens were also significantly reduced in the obstructed kidney of Smad3(-/-) mice. In addition, the numbers of myofibroblasts, macrophages, and CD4/CD8 T cells infiltrated into the kidney after UUO were significantly attenuated in the obstructed kidney of Smad3(-/-) mice when compared with that of Smad3(+/+) mice. Furthermore, terminal deoxynucleotidyltransferase-mediated deoxyuridine triphosphate (dUTP) nick-end labeling (TUNEL) staining after UUO showed significantly reduced number of tubular apoptotic cells in the obstructed kidney of Smad3(-/-) mice when compared with that of Smad3(+/+) mice. Endogenous Smadpathway was activated in the obstructed kidney after UUO in wild-type mice as judged by the increase of phosphorylated Smad2 or phosphorylated Smad2/3-positive cells in renal interstitial area. Conclusion Smad3 deficiency attenuated renal fibrosis, inflammation, and apoptosis after UUO, suggesting that Smad3 was a key molecule mediating TGF-β activity leading to real fibrosis after UUO.

Purpose: The study goal was to evaluate the change in hyoid bone position after neck dissection and tumor resection for oral cancer. Patients and Methods: Twenty-seven patients who underwent surgery were retrospectively evaluated. The hyoid bone position was measured before and after the surgery using a topogram of computed tomography scans. Results: There was a statistically significant difference in the hyoid bone position between preoperative and postoperative images in anteroposterior direction (Wilcoxon signed rank test, P

We extracted Triton-treated erythrocyte ghosts with 2 M KCl (Triton/KCl/ghosts), and then with 1.2 M KBr at pH 5.5 (Triton/KCl/KBr ghosts). Triton/KCl/KBr ghosts were very similar in shape to untreated ghosts, Triton ghosts and Triton/KCl ghosts under a phase-contrast microscope at various pH values and salt concentrations, despite having lost most of their phospholipids and proteins, except for spectrin. Negatively stained Triton ghosts, Triton/KCl ghosts and Triton/KCl/KBr ghosts appeared similar to each other, but the regularity of the spectrin network structure decreased somewhat in that order. Triton/KCl/KBr ghosts were stabilized by adding both actin and band 4.1, but not by adding either alone. These and previous findings strongly suggest that the spectrin network is visible and the simplest inframembrane structure.

culture markedly decreased nuclear LXR protein levels, suggesting that 25HC3S may complex with LXR to prevent its activation. 25HC3S administration was followed by doseand timedependent decreases in SREBP-1 protein and mRNA, consistent with down regulation of SREBP-1 transcription. 25HC3S administration also led to a decrease in mature SREBP-1 accompanied by an increase in SREBP-1 precursor, consistent with post-translational downregulation of SREBP-1 activity. 25HC3S decreased the expression of a number of SREBP1-responsive genes including acetyl CoA carboxylase-1 (ACC-1) and fatty acid synthase (FAS), key enzymes involved in fatty acid biosynthesis, as well as Cyp7A1, HMGR, and LDLR, which are key proteins involved in cholesterol metabolism. 25HC, the precursor of 25HC3S, had opposite effects, increasing SREBP-1 and FAS mRNA levels in the hepatocytes. Conclusion: 25HC3S, initially produced in mitochondria in response to excess cholesterol, is a potent down-regulator of the cholesterol and fatty acid biosynthetic pathways. The effects of 25HC3S appear to be mediated via inhibition of LXR/SREBP-1 signaling.