Negative regulation of the interferon response by an interferon-induced long non-coding RNA.

Long non-coding RNAs (lncRNAs) play critical roles in diverse cellular processes; however, their involvement in many critical aspects of the immune response including the interferon (IFN) response remains poorly understood. To address this gap, we compared the global gene expression pattern of primary human hepatocytes before and at three time points after treatment with IFN-α. Among ∼ 200 IFN-induced lncRNAs, one transcript showed ∼ 100-fold induction. This RNA, which we named lncRNA-CMPK2, was a spliced, polyadenylated nuclear transcript that was induced by IFN in diverse cell types from human and mouse. Similar to protein-coding IFN-stimulated genes (ISGs), its induction was dependent on JAK-STAT signaling. Intriguingly, knockdown of lncRNA-CMPK2 resulted in a marked reduction in HCV replication in IFN-stimulated hepatocytes, suggesting that it could affect the antiviral role of IFN. We could show that lncRNA-CMPK2 knockdown resulted in upregulation of several protein-coding antiviral ISGs. The observed upregulation was caused by an increase in both basal and IFN-stimulated transcription, consistent with loss of transcriptional inhibition in knockdown cells. These results indicate that the IFN response involves a lncRNA-mediated negative regulatory mechanism. lncRNA-CMPK2 was strongly upregulated in a subset of HCV-infected human livers, suggesting a role in modulation of the IFN response in vivo.

Functional liver tissue engineering by an adult mouse liver-derived neuro-glia antigen 2-expressing stem/progenitor population.

Deaths due to end-stage liver diseases are increasingly registered annually in the world. Liver transplantation is the ultimate treatment for end-stage liver diseases to date, which has been hampered by a critical shortage of organs. The potential of decellularized liver scaffolds (DLS) derived from solid organs as a three-dimensional platform has been evolved as a promising approach in liver tissue engineering for translating functional liver organ replacements, but questions still exist regarding the optimal cell population for seeding in DLS and the preparation of the DLS themselves. The aim of our study was to utilize a sodium dodecyl sulfate decellularization procedure in combination with a low concentration of trypsin (0.005%)-ethylenediaminetetraacetic acid (0.002%) process to manufacture DLS from whole mouse livers and recellularized with hepatic stem/progenitors for use in liver tissue engineering and injured liver treatment. Results showed that the DLS generated with all the necessary microstructure and the extracellular components to support seeded hepatic stem/progenitor cell attachment, functional hepatic cell differentiation. Hepatic differentiation from stem/progenitor cells loaded by DLS was more efficient than that of the stem/progenitor cells in the two-dimensional cell culture model. In summary, the method of DLS loaded by hepatic stem/progenitor cells provided by this study was effective in maintaining DLS extracellular matrix to introduce seeded stem/progenitor cell differentiation, hepatic-like tissue formation and functional hepatic protein production in vitro that promoted functional recovery and survival in a mouse model of dimethylnitrosamine-induced liver cirrhosis after auxiliary heterotopic liver transplantation. Copyright © 2016 John Wiley & Sons, Ltd.

Extended Perioperative Antibiotic Coverage in Conjunction with Intraoperative Bile Cultures Decreases Infectious Complications after Pancreaticoduodenectomy.

Background. Bile contamination from the digestive tract is a well-known risk factor for postoperative complications. Despite the literature concerning prevalence of bacterobilia and fungobilia in patients with biliary pathologies, there are no specific recommendations for perioperative antimicrobial coverage for biliary/pancreatic procedures. We evaluated the effect of at least 72 hours of perioperative broad spectrum antibiotic coverage on outcomes of pancreaticoduodenectomy (PD). Materials and Methods. A retrospective review of all patients at Case Medical Center of Case Western Reserve University undergoing PD procedure, from 2006 to 2011, was performed (n = 122). Perioperative data including demographics, comorbidities, biliary instrumentation, antibiotic coverage, culture results, and postoperative outcomes were analyzed. Propensity score matching method was used to match the patients according to duration of antibiotic coverage into two groups: 72 hours (A72) and 24 hours (A24). Results. Longer broad spectrum antibiotic coverage in group A72 resulted in significantly less surgical site infections after PD, compared to routine 24 hours of perioperative antibiotics in group A24. This study did not reveal a statistically significant decrease in postoperative fungal infections in patients receiving preoperative antifungals. Conclusion. Prolonged perioperative antibiotic therapy in conjunction with intraoperative bile cultures decreases the short-term infectious complications of PD, with no significant increase in Clostridium difficile colitis incidence.

Repair of liver mediated by adult mouse liver neuro-glia antigen 2-positive progenitor cell transplantation in a mouse model of cirrhosis.

NG2-expressing cells are a population of periportal vascular stem/progenitors (MLpvNG2(+) cells) that were isolated from healthy adult mouse liver by using a "Percoll-Plate-Wait" procedure. We demonstrated that isolated cells are able to restore liver function after transplantation into a cirrhotic liver, and co-localized with the pericyte marker (immunohistochemistry: PDGFR-ß) and CK19. Cells were positive for: stem cell (Sca-1, CD133, Dlk) and liver stem cell markers (EpCAM, CD14, CD24, CD49f); and negative for: hematopoietic (CD34, CD45) and endothelial markers (CD31, vWf, von Willebrand factor). Cells were transplanted (1 × 10(6) cells) in mice with diethylnitrosamine-induced cirrhosis at week 6. Cells showed increased hepatic associated gene expression of alpha-fetoprotein (AFP), Albumin (Alb), Glucose-6-phosphatase (G6Pc), SRY (sex determining region Y)-box 9 (Sox9), hepatic nuclear factors (HNF1a, HNF1ß, HNF3ß, HNF4α, HNF6, Epithelial cell adhesion molecule (EpCAM), Leucine-rich repeated-containing G-protein coupled receptor 5-positive (Lgr5) and Tyrosine aminotransferase (TAT). Cells showed decreased fibrogenesis, hepatic stellate cell infiltration, Kupffer cells and inflammatory cytokines. Liver function markers improved. In a cirrhotic liver environment, cells could differentiate into hepatic lineages. In addition, grafted MLpvNG2(+) cells could mobilize endogenous stem/progenitors to participate in liver repair. These results suggest that MLpvNG2(+) cells may be novel adult liver progenitors that participate in liver regeneration.

Thrombotic events and markers of oxidation and inflammation in hemodialysis.

The goal of this study was to determine whether antioxidant therapy with vitamin E would alter the rate of vascular access complications or other macrovascular complications in hemodialysis (HD) patients. A secondary goal of the study was to explore the relationship between baseline pretreatment markers of oxidative stress (the advanced glycation end product pentosidine and basal levels of vitamin Ealpha and gamma) and the subsequent development of access failure. Thirty-five stable patients treated by HD were recruited for the study. Patients were provided with vitamin E (800 IU) or placebo capsules to be taken daily. Clinical variables, vascular access function (flow meter access flow measurements), and circulating blood markers were obtained initially and every 3 months throughout the study. Vitamin Ealpha levels rose in treated patients from 12.7 +/- 4.4 to 25.1 +/- 15.1 microg/mL at 3 months and 28.6 +/- 14.8 microg/mL at 6 months. Vitamin Egamma levels fell in treated patients from 3.9 +/- 1.7 to 2.3 +/- 1.5 microg/mL at 3 months and 1.7 microg/mL at 6 months. Patients who subsequently developed repeated thrombotic vascular access events were characterized by higher baseline pentosidine content of circulating proteins. Patients who developed a myocardial infarction had higher pentosidine, lower vitamin Ealpha, and much lower vitamin Egamma than patients who did not develop thrombotic events. These findings lead to the speculation that the anti-inflammatory effects of vitamin Egamma may play a more important role in thrombotic vascular events than the antioxidant effects of vitamin Ealpha. Additional studies of these interactions are in progress.

Rescue ALPPS: Intraoperative Conversion to ALPPS during Synchronous Resection of Rectal Cancer and Liver Metastasis.

Future liver remnant (FLR) is the most important deciding factor in planning for liver resection. Portal vein embolization (PVE) was first introduced in the 1980s to induce liver hypertrophy, enabling removal of multiple/bilobar tumors. PVE was later combined with sequential hepatectomies with the aim of allowing the liver remnant to hypertrophy (15-20%) between procedures. However, the interval between the two procedures (3-8 weeks) put patients at risk for disease progression. With portal vein ligation alone or when combined with sequential hepatectomy, there is also a risk for inadequate liver hypertrophy because of intrahepatic portal collaterals leading to a high (19-30%) dropout rate. The ALPPS procedure (associating liver partition and portal vein ligation for staged hepatectomy) was recently developed as a feasible means to perform extensive/bilobar liver resections. It produces rapid, enormous hypertrophy of the remnant, making previously unresectable lesions resectable. Indications for ALPPS include any extensive liver resection with inadequate FLR. Here we present a novel indication for ALPPS as a rescue when inadequate FLR was faced intraoperatively, during a simultaneous resection of rectal primary and liver metastasis.

Early experience with intraoperative radiotherapy in patients with resected pancreatic adenocarcinoma.

BACKGROUND: The use of intraoperative radiotherapy (IORT) in patients with resected pancreatic adenocarcinoma has not been clearly defined. METHODS: The medical records of our first 22 patients receiving IORT for resected pancreatic adenocarcinoma (2001 to 2006) were reviewed and compared with the records of 27 consecutive patients not receiving IORT for resected pancreatic adenocarcinoma (2004 to 2006). RESULTS: There were no 30-day mortalities in either group, and complication rates were similar. Local recurrence occurred in 18% in the IORT group (median 14 months) and 12% in the no-IORT group (median 7 months). Distant recurrence occurred in 47% in the IORT group (median 11 months) and 32% in the no-IORT group (median 6.5 months). Median overall, stage-specific, and location-specific survival did not differ between the groups. CONCLUSIONS: Although limited in size and follow-up, our experience showed that complications, recurrence, and survival were not affected by IORT, but time to recurrence may be longer with IORT.

The impact of resection margin status and postoperative CA19-9 levels on survival and patterns of recurrence after postoperative high-dose radiotherapy with 5-FU-based concurrent chemotherapy for resectable pancreatic cancer.

OBJECTIVES: To analyze the impact of surgical margins and other clinicopathological data on treatment outcomes on 75 patients treated from 1999 to 2006 by initial potentially curative surgery (+/- intraoperative radiotherapy), followed by high-dose 3-dimensional conformal radiation therapy and concomitant fluoropyrimidine-based chemoradiotherapy. MATERIALS AND METHODS: All clinical and pathologic data on this patient cohort were analyzed by actuarial Kaplan-Meier survival methodology and by univariate and multivariate Cox proportional hazards methods to measure effects on survival and patterns of failure. RESULTS: With a median follow-up of 28 months, the median, 2-year and 5-year overall survival (OS) rates were 18.1 month, 41% and 23.6%, respectively. Disease-free survival (DFS) rates were of 11.4 months, 35% and 20%, respectively. Only 2 clinicopathological features, positive (< or =1 mm) surgical margins (P < 0.05) and a 2-fold (>70 U/mL) elevation of the postoperative serum CA19-9 (P < 0.001) impacted OS and disease-free survival. In patients with negative (>1 mm) surgical margins and a low (< or =70 U/mL) postoperative CA19-9, the projected 2- and 5-year OS were 80% and 65%, respectively, compared with 40% and 10% with positive surgical margins and a low CA19-9 and to 10% and 0% with positive or negative surgical margins and a high (>70 U/mL) CA19-9. Positive surgical margins (P < 0.001) and an elevated postoperative CA19-9 (P < 0.001) also predicted early development of distant metastases, whereas isolated loco-regional failure was less common and not affected by these or other clinicopathological features. CONCLUSIONS: Using this fluoropyrimidine-based chemoradiotherapy regimen after surgical resection (+/- intraoperative radiotherapy), positive surgical margins and an elevated (2-fold) postoperative serum CA19-9 level predicted for reduced survival and early development of distant metastatic disease.

Comparison of sirolimus vs. mycophenolate mofetil on surgical complications and wound healing in adult kidney transplantation.

Mycophenolate mofetil (MMF) and sirolimus impair wound healing. We compared sirolimus vs. MMF to determine the relative impact on surgical complications and wound healing in adult kidney transplant recipients. This retrospective, single center study of 235 adult kidney transplants performed between 1 January 2000 and 31 January 2002 identified 158 adult, kidney-only recipients treated with tacrolimus and prednisone, from which two groups were defined: group 1 (n = 84) received MMF, group 2 (n = 74) received sirolimus. The incidence of fluid collections, wound problems, dehiscence, and urine leak were compared. A multivariate stepwise logistical regression analysis was performed to identify risk factors. The overall incidence of complications was 21.5%, with rates significantly lower in group 1 (2.4%) vs. group 2 (43.2%, p < 0.0001). Regression analysis showed only sirolimus (p < 0.001) and hypo-albuminemia (p = 0.006) to independently correlate with complication occurrence. In subanalyses, lymphoceles correlated only with sirolimus (p = 0.003), while other wound problems also correlated with higher body mass index (p = 0.067). The use of sirolimus, tacrolimus and prednisone was associated with a greater incidence of lymphoceles, non-lymphocele perinephric fluid collections and other consequences of poor wound healing, as compared to contemporary patients treated with MMF, tacrolimus and prednisone.