Tet2 loss leads to increased hematopoietic stem cell self-renewal and myeloid transformation

Somatic loss-of-function mutations in the ten-eleven translocation 2 (TET2) gene occur in a significant proportion of patients with myeloid malignancies. Although there are extensive genetic data implicating TET2 mutations in myeloid transformation, the consequences of Tet2 loss in hematopoietic development have not been delineated. We report here an animal model of conditional Tet2 loss in the hematopoietic compartment that leads to increased stem cell self-renewal in vivo as assessed by competitive transplant assays. Tet2 loss leads to a progressive enlargement of the hematopoietic stem cell compartment and eventual myeloproliferation in vivo, including splenomegaly, monocytosis, and extramedullary hematopoiesis. In addition, Tet2(+/-) mice also displayed increased stem cell self-renewal and extramedullary hematopoiesis, suggesting that Tet2 haploinsufficiency contributes to hematopoietic transformation in vivo.     

Claudin-4 forms paracellular chloride channel in the kidney and requires claudin-8 for tight junction localization

Tight junctions (TJs) play a key role in mediating paracellular ion reabsorption in the kidney. The paracellular pathway in the collecting duct of the kidney is a predominant route for transepithelial chloride reabsorption that determines the extracellular NaCl content and the blood pressure. However, the molecular mechanisms underlying the paracellular chloride reabsorption in the collecting duct are not understood. Here we showed that in mouse kidney collecting duct cells, claudin-4 functioned as a Cl(-) channel. A positively charged lysine residue at position 65 of claudin-4 was critical for its anion selectivity. Claudin-4 was observed to interact with claudin-8 using several criteria. In the collecting duct cells, the assembly of claudin-4 into TJ strands required its interaction with claudin-8. Depletion of claudin-8 resulted in the loss of paracellular chloride conductance, through a mechanism involving its recruitment of claudin-4 during TJ assembly. Together, our data show that claudin-4 interacts with claudin-8 and that their association is required for the anion-selective paracellular pathway in the collecting duct, suggesting a mechanism for coupling chloride reabsorption with sodium reabsorption in the collecting duct.     

Epithelial-to-mesenchymal transition and chronic allograft tubulointerstitial fibrosis

Chronic allograft tubular atrophy/interstitial fibrosis (TA/IF) is a major cause of late allograft loss. A major challenge to the future of kidney transplantation is to dissect the identifiable causes of chronic allograft TA/IF and to develop cause-specific treatment strategies. Emerging evidence suggests that epithelial-to-mesenchymal transition (EMT) is an important event in native and transplant kidney injury, including chronic allograft TA/IF. During EMT, tubular epithelial cells are transformed into myofibroblasts through a stepwise process including loss of cell-cell adhesion and E-cadherin expression, de novo alpha-smooth muscle actin expression, actin reorganization, tubular basement membrane disruption, cell migration, and fibroblast invasion with production of profibrotic molecules such as collagen types I and III and fibronectin. We examined in this review the molecular and cellular pathways of EMT and their involvement in chronic allograft tubulointerstitial fibrosis. We examined the role of alloimmune T cells and oxidative stress in this context and evaluated EMT as a marker of disease progression. Potential therapeutic options are discussed. In conclusion, there is enough evidence demonstrating that EMT is involved in the pathogenesis of chronic allograft tubulointerstitial fibrosis. However, the extent of its contribution to allograft fibrogenesis remains unknown, and only interventional trials will enable us to clarify this question. Furthermore, additional data are required to determine whether EMT may be used as a surrogate marker of disease progression in kidney transplant recipients.     

Molecular profiling of chronic lymphocytic leukaemia: genetics meets epigenetics to identify predisposing genes

Molecular profiling may lead to a better understanding of a disease. This knowledge is especially important in malignancies, where multiple alterations are required during the progression from premalignant to malignant stages. Such information can be useful for the development of novel biomarkers that allow the prediction of a clinical course, response to treatment or early detection. Molecular data is also utilized to develop targeted therapies. Moreover, gene defects identified in profiling studies will help to understand the molecular pathways disrupted in the disease. This review provides an overview of molecular profiling approaches in chronic lymphocytic leukaemia (CLL). We will describe our current understanding of genetic alterations in CLL, the use of familial CLL for the identification of predisposing mutations, and the search for epigenetic alterations in CLL.     

ProSeal as an alternative to endotracheal intubation in pediatric laparoscopy

BACKGROUND: The increasing use of laparoscopic surgery in children mandates a leak-free airway device during carboperitoneum for which tracheal tubes (TT) have been traditionally used. The pediatric ProSeal is a recent introduction to the laryngeal mask airway (LMA) family. The ProSeal LMA (PLMA) has been successfully used in adult laparoscopic procedures. We hypothesized that the ProSeal can be equally effective in achieving adequate oropharyngeal seal and pulmonary ventilation during elective laparoscopic procedures in children. METHODS: Sixty ASA I and II children, 6 months to 8 years, scheduled for elective laparoscopic surgeries (duration of carboperitoneum <60 min) were randomly assigned to two groups of 30 each (TT and ProSeal(TM)). Patients at risk of aspiration, difficult airway and upper respiratory tract infection were excluded. Anesthesia technique included 8% of sevoflurane/O(2)/N(2)O and neuromuscular blockade. Ventilation was set to 10 ml.kg(-1) with positive endexpiratory pressure of 5 cmH(2)O. Respiratory rate was adjusted to maintain endtidal CO(2) (P(E)CO(2)) between 4.6 and 5.8 kPa (35-45 mmHg). Carboperitoneum was achieved at 10 mmHg. Hemodynamic parameters, peak inspiratory pressure (PIP) and P(E)CO(2) were noted at different time points. Chi-squared test, Mann-Whitney U-test and Wilcoxan W-test were applied; P < 0.05 was considered significant. RESULTS: There were no statistically significant differences in SpO(2) and P(E)CO(2) between PLMA and TT groups before and after peritoneal insufflation. Significant change was seen in PIP (cmH(2)O) from induction to carboperitoneum in both groups (14.3 +/- 1.48 in PLMA, 14.2 +/- 1.46 in TT to 23.36 +/- 1.7 in PLMA, and 23.73 +/- 1.9 in TT); and in P(E)CO(2) (mmHg), from 32 +/- 2.2 to 48 +/- 3.6 in PLMA and 30 +/- 3.1 to 47 +/- 2.8 in the TT group. The changes were not significant; P > 0.05 between the groups. The first-time success rate for PLMA was 88%, mean oropharyngeal leak pressure was 29 +/- 3 cmH(2)O. CONCLUSIONS: The pediatric PLMA and TT have comparable ventilatory efficacy for elective short laparoscopic procedures.     

Identification of Ras-related nuclear protein, targeting protein for xenopus kinesin-like protein 2, and stearoyl-CoA desaturase 1 as promising cancer targets from an RNAi-based screen

To identify new candidate cancer drug targets, we used RNAi as a tool to functionally evaluate genes that play a role in maintaining human tumor cell survival. We screened a small interfering RNA (siRNA) library directed against approximately 3,700 individual genes to assess the ability of siRNAs to induce cell death in an in vitro cell cytotoxicity assay. We found that siRNAs specifically targeting ras-related nuclear protein (Ran), targeting protein for Xenopus kinesin-like protein 2 (TPX2), and stearoyl-CoA desaturase 1 (SCD1), significantly reduced the survival of multiple human tumor cell lines. Further target validation studies revealed that treatment with Ran and TPX2 siRNAs differentially reduced the survival of activated K-Ras-transformed cells compared with their normal isogenic counterparts in which the mutant K-Ras gene had been disrupted (DKS-8). Knockdown of Ran and TPX2 in activated mutant K-Ras cells selectively induced S-phase arrest or transient G(2)-M arrest phenotypes, respectively, that preceded apoptotic cell death. Given our observations that Ran and TPX2 depletion preferentially reduces the survival of activated K-Ras-transformed cells, these two proteins may serve as useful anticancer targets in tumors expressing the activated K-Ras oncogene.