Met-RANTES ameliorates fibrous airway obliteration and decreases ERK expression in a murine model of bronchiolitis obliterans.

OBJECTIVES: Bronchiolitis obliterans (BO) is the main cause of late mortality among long-term survivors of lung transplantation. Chemokine-chemokine receptor (CCR) interaction and subsequent recruitment of infiltrating cells to the graft are early events in the development of chronic rejection of transplanted lungs. The present study investigated whether blockade of chemokine receptors CCR1 and CCR5 with Met-regulated-on-activation, normal T cells expressed and secreted (RANTES), an amino-terminal modified derivative of RANTES/CCL5, affects the development of BO in murine model and we sought to determine the expression of RANTES/CCL5 and their relationship with extracellular signal-regulated kinase (ERK). Materials and Methods: BALB/c mouse tracheas were heterotopically transplanted into C57Black6 recipients and treated for 21 days with either Met-RANTES at 20 microg/day or vehicle. Animals were killed at 21 days after transplantation for histologic examination of ERK expression. RESULTS: RANTES/CCL5 was highly expressed in allografts compare to isografts. Met-RANTES treatment ameliorated fibrous airway obliteration in a mouse model of BO and decreased ERK expression. CONCLUSION: Blockade of chemokine receptors by Met-RANTES ameliorated airway obliteration and decreased ERK expression. These findings suggest that chemokine receptors CCR1 and CCR5 play significant roles in the development of chronic rejection and ERK may be a new molecular target for chronic rejection.     

ROS‐generating oxidases Nox1 and Nox4 contribute to oncogenic Ras‐induced premature senescence

Activated oncogenes induce premature cellular senescence, a permanent state of proliferative arrest in primary rodent and human fibroblasts. Recent studies suggest that generation of reactive oxygen species (ROS) is involved in oncogenic Ras‐induced premature senescence. However, the signaling mechanism controlling this oxidant‐mediated irreversible growth arrest is not fully understood. Here, we show that through the Ras/MEK pathway, Ras oncogene up‐regulated the expression of superoxide‐generating oxidases, Nox1 in rat REF52 cells and Nox4 in primary human lung TIG‐3 cells, leading to an increase in intracellular level of ROS. Ablation of Nox1 and Nox4 by small interfering RNAs (siRNAs) blocked the RasV12 senescent phenotype including β‐galactosidase activity, growth arrest and accumulation of tumor suppressors such as p53 and p16^Ink4a. This suggests that Nox‐generated ROS transduce senescence signals by activating the p53 and p16^Ink4a pathway. Furthermore, Nox1 and Nox4 siRNAs inhibited both Ras‐induced DNA damage response and p38MAPK activation, whereas overexpression of Nox1 and Nox4 alone was able to induce senescence. The involvement of Nox1 in Ras‐induced senescence was also confirmed with embryonic fibroblasts derived from Nox1 knockout mice. Together, these findings suggest that Nox1‐ and Nox4‐generated ROS play an important role in Ras‐induced premature senescence, which may involve DNA damage response and p38MAPK signaling pathways.     

Laparoscopic biliary surgery

Laparoscopic cholecystectomy has become the standard treatment for patients with symptomatic gallbladder disease. However, there is a substantial proportion of patients in whom laparoscopic cholecystectomy cannot be successfully performed, and conversion to open surgery is required because of technical difficulties or complications. The incidence of bile duct injury has increased in laparoscopic cholecystectomy. Meticulous dissection and intraoperative cholangiography could significantly reduce the rate of that injury. Laparoscopic cholecystectomy for acute cholecystitis is still controversial because of surgical difficulty. In our experience, early laparoscopic cholecystectomy is a beneficial option for patients with acute cholecystitis, and it may even be safe in the acute stage. A better alternative for high-risk early operation and septic cases is percutaneous transhepatic gallbladder drainage. The coexistence of gallbladder cancer should be ruled out and preoperative diagnosis should be done carefully. Laparoscopic management of common bile duct (CBD) stones has many advantages. However it has been reported to be demanding and time-consuming to perform, which limits its widespread adoption. In our experience with 258 patients, laparoscopic CBD exploration was feasible for almost all CBD stones. The technical difficulties associated with laparoscopic CBD exploration could be overcome with the development of suitable equipment and increased expertise.     

Synergistic induction of apoptosis by acyclic retinoid and interferon-beta in human hepatocellular carcinoma cells

Acyclic retinoid, a synthetic retinoid analog, as well as interferon alfa (IFN-alpha) and IFN-beta induce apoptosis in hepatocellular carcinoma (HCC) cells and are used clinically in the prevention of HCC. Here, we show that acyclic retinoid acts synergistically with IFNs in suppressing the growth and inducing apoptosis (as characterized by DNA fragmentation and chromatin condensation) in 5 human HCC cell lines (JHH7, HuH7, PLC/PRF/5, HLE, and HLF). This synergism was only observed when cells were pretreated with the acyclic retinoid, whereas natural retinoic acids (all-trans and 9-cis retinoic acid) were ineffective. This promotion may be due to up-regulation of type 1 IFN receptor (IFNR) expression by the retinoid. Accordingly, incubation with antitype 1 IFNR antibody abolished the synergy. Enhanced IFNR expression was accompanied by increased expression and DNA-binding activity of STAT1, an intracellular signal transducing molecule of IFNR, and increased induction of 2', 5'-oligoadenyl-5'-triphosphate synthetase, which is a target gene of STAT1. Acyclic retinoid did not have any effects on the growth of normal human hepatocytes (Hc) probably because of a lack of IFNR and STAT1 up-regulation. In conclusion, these results provide a rationale for combined biochemoprevention of HCC using acyclic retinoid and IFN-beta.     

Aberrant methylation profile of human malignant mesotheliomas and its relationship to SV40 infection

Malignant mesothelioma (MM) is associated with asbestos exposure and the presence of SV40 viral sequences. Recently, we reported that SV40 infection of human mesothelial cells (HM) causes aberrant methylation of the tumor suppressor gene (TSG) RASSF1A. We investigated methylation of 12 genes by methylation-specific PCR in 63 MMs, six MM cell lines, and two foci of SV40-infected HM. Methylation percentages of the tested genes ranged from 3 to 65%. The frequencies of HPP1, RASSF1A, Cyclin D2, and RRAD methylation, and the value of the methylation index, were significantly higher in SV40 sequence-positive MMs than in SV40-negative MMs. Methylation of TMS1 and HIC-1 was associated with shortened survival. SV40-infected HM showed progressive aberrant methylation of seven genes (RASSF1A, HPP1, DcR1, TMS1, CRBP1, HIC-1, and RRAD) during serial passage. Our results demonstrate a relationship between SV40 and methylation of multiple genes in MM, indicating that the virus plays a role in the pathogenesis of MM.     

DNA methylation profiles of lymphoid and hematopoietic malignancies.

PURPOSE: Aberrant methylation of the 5' gene promoter regions is an epigenetic phenomenon that is the major mechanism for silencing of tumor suppressor genes in many cancer types. The aims of our study were (a) to compare the methylation profiles of the major forms of hematological malignancies and (b) to determine the methylation profile of monoclonal gammopathy of undetermined significance (MGUS) and compare it with that of multiple myeloma (MM). EXPERIMENTAL DESIGN: We compared the aberrant promoter methylation profile of 14 known or suspected tumor suppressor genes in leukemias (n = 48), lymphomas (n = 42), and MMs (n = 40). We also examined the methylation profile of MGUS (n = 20), a premalignant plasma cell dyscrasia. The genes studied represent five of the six "hallmarks of cancer." RESULTS: Peripheral blood lymphocytes (n = 14) from healthy volunteers were negative for methylation of all genes, and methylation percentages in 41 nonmalignant tissues (peripheral blood mononuclear cells, bone marrows, and lymph nodes) from hematological patients were low (0-9%) for all 14 genes, confirming that methylation was tumor specific. Ten of the genes were methylated at frequencies of 29-68% in one or more tumor types, and the methylation indices (an indicator of overall methylation) varied from 0.25 to 0.34. With two exceptions, the methylation patterns of leukemias and lymphomas were similar. However, the pattern of MMs varied from the other tumor types for six genes. In general, the methylation pattern of MGUS was similar to that of MM, although the methylation frequencies were lower (the methylation index of MGUS was 0.15, and that of MM was 0.3). However, the methylation frequencies of six genes were significantly higher in MGUS than in control tissues. The relatively high frequencies of methylation in MGUS are consistent with it being a premalignant condition. CONCLUSIONS: The three major forms of lymphoid/hematopoietic malignancies show overlapping but individual patterns of methylation.     

Helicobacter pylori eradication therapy does not prevent gastric cancer development in all patients

We previously reported that eradication of Helicobacter pylori reduced the risk of gastric cancer developing in patients with peptic ulcer diseases. In the present study, we followed up with our patient group to investigate the occurrences and clinical features of gastric cancers that developed after cure of the infection. Prospective post-eradication evaluations were conducted on 1, 674 consecutive patients who had received successful H. pylori eradication therapy. The patients underwent endoscopic examination before eradication therapy to test for peptic ulcers, background gastric mucosal atrophy, and H. pylori infection. After confirmation of the cure of infection, the annual follow-up endoscopy was performed. The patients were followed up to more than 10 years. During the follow-up, their risk of developing gastric cancer after the cure of infection was almost the same as we reported previously. There was still a risk of developing gastric cancer of both the intestinal and diffuse types. The grade of gastric mucosal atrophy present before receiving eradication therapy was closely related to the development of gastric cancer after eradication of H. pylori. The stage of most gastric cancer was at the early TNM stage, but advanced cancer was observed in patients who skipped regular endoscopic surveillance. H. pylori eradication therapy does not prevent gastric cancer development in all infected patients. Thus, it is important to inform patients about the risk of gastric cancer after eradication therapy and to offer them surveillance endoscopy.