LPS-induced epithelial barrier disruption via hyperactivation of CACC and ENaC.

Gram-negative bacterial lipopolysaccharide (LPS) increases the susceptibility of cells to pathogenic diseases, including inflammatory diseases and septic syndrome. In our experiments, we examined whether LPS induces epithelial barrier disruption in secretory epithelia and further investigated its underlying mechanism. The activities of Ca2+-activated Cl- channels (CACC) and epithelial Na+ channels (ENaC) were monitored with a short-circuit current using an Ussing chamber. Epithelial membrane integrity was estimated via transepithelial electrical resistance and paracellular permeability assays. We found that the apical application of LPS evoked short-circuit current (Isc) through the activation of CACC and ENaC. Although LPS disrupted epithelial barrier integrity, this was restored with the inhibition of CACC and ENaC, indicating the role of CACC and ENaC in the regulation of paracellular pathways. We confirmed that LPS, CACC, or ENaC activation evoked apical membrane depolarization. The exposure to a high-K+ buffer increased paracellular permeability. LPS induced the rapid redistribution of zonula occludens-1 (ZO-1) and reduced the expression levels of ZO-1 in tight junctions through apical membrane depolarization and tyrosine phosphorylation. However, the LPS-induced epithelial barrier disruption and degradation of ZO-1 were largely recovered by blocking CACC and ENaC. Furthermore, although LPS-impaired epithelial barrier became vulnerable to secondary bacterial infections, this vulnerability was prevented by inhibiting CACC and ENaC. We concluded that LPS induces the disruption of epithelial barrier integrity through the activation of CACC and ENaC, resulting in apical membrane depolarization and the subsequent tyrosine phosphorylation of ZO-1.

CXCR4 Regulates Temporal Differentiation via PRC1 Complex in Organogenesis of Epithelial Glands.

CXC-chemokine receptor type 4 (CXCR4), a 7-transmembrane receptor family member, displays multifaceted roles, participating in immune cell migration, angiogenesis, and even adipocyte metabolism. However, the activity of such a ubiquitously expressed receptor in epithelial gland organogenesis has not yet been fully explored. To investigate the relationship between CXCL12/CXCR4 signaling and embryonic glandular organogenesis, we used an ex vivo culture system with live imaging and RNA sequencing to elucidate the transcriptome and protein-level signatures of AMD3100, a potent abrogating reagent of the CXCR4-CXCL12 axis, imprinted on the developing organs. Immunostaining results showed that CXCR4 was highly expressed in embryonic submandibular gland, lung, and pancreas, especially at the periphery of end buds containing numerous embryonic stem/progenitor cells. Despite no significant increase in apoptosis, AMD3100-treated epithelial organs showed a retarded growth with significantly slower branching and expansion. Further analyses with submandibular glands revealed that such responses resulted from the AMD3100-induced precocious differentiation of embryonic epithelial cells, losing mitotic activity. RNA sequencing analysis revealed that inhibition of CXCR4 significantly down-regulated polycomb repressive complex (PRC) components, known as regulators of DNA methylation. Treatment with PRC inhibitor recapitulated the AMD3100-induced precocious differentiation. Our results indicate that the epigenetic modulation by the PRC-CXCR12/CXCR4 signaling axis is crucial for the spatiotemporal regulation of proliferation and differentiation of embryonic epithelial cells during embryonic glandular organogenesis.

Epigenetic Modification of CFTR in Head and Neck Cancer.

Cystic fibrosis transmembrane conductance regulator (CFTR), a cyclic AMP (cAMP)-regulated chloride channel, is critical for secretion and absorption across diverse epithelia. Mutations or absence of CFTR result in pathogeneses, including cancer. While CFTR has been proposed as a tumor suppressing gene in tumors of the intestine, lung, and breast cancers, its effects in head and neck cancer (HNC) have yet to be investigated. This study aimed to define expression patterns and epigenetic modifications of CFTR in HNC. CFTR was expressed in normal but not in HNC cells and tissues. Treatment with 5-aza-2'-deoxycytidine (5-Aza-CdR) was associated with rescued expression of CFTR, whose function was confirmed by patch clamp technique. Further experiments demonstrated that CFTR CpG islands were hypermethylated in cancer cells and tissues and hypomethylated in normal cells and tissue. Our results suggest that CFTR epigenetic modifications are critical in both down-regulation and up-regulation of CFTR expression in HNC and normal cells respectively. We then investigated the impact of CFTR on expressions and functions of cancer-related genes. CFTR silencing was closely associated with changes to other cancer-related genes, suppressing apoptosis while enhancing proliferation, cell motility, and invasion in HNC. Our findings demonstrate that hypermethylation of CFTR CpG islands and CFTR deficiency is closely related to HNC.

Clinical analysis of hemodialysis vascular access: comparision of autogenous arterioveonus fistula & arteriovenous prosthetic graft.

BACKGROUND: Mature autogenous arteriovenous fistulas have better long term patency and require fewer secondary interventions compared to arteriovenous prosthetic graft. Our Study evaluated vascular patency rates and incidence of interventions in autogenous arteriovenous fistulas and grafts. MATERIAL AND METHODS: A total of 166 vascular access operations were performed in 153 patients between December 2002 and November 2009. Thirty seven caeses were excluded due to primary access failure and loss of follow-up. One group of 92 autogenous arterioveous fistulas and the other group of 37 arteriovenous prosthetic grafts were evaluated retrospectively. Primary and secondary patency rates were estimated using the Kaplan-Meier method. RESULTS: The primary patency rate (84%, 67%, 51% vs. 51%, 22%, 9% at 1, 3, 5 year; p=0.0000) and secondary patency rate (96%, 88%, 68% vs. 88%, 65%, 16% at 1. 3, 5 year; p=0.0009) were better in autogenous fistula group than prosthetic graft group. Interventions to maintain secondary patency were required in 23% of the autogenous fistula group (average 0.06 procedures/patient/year) and 65% of prosthetic graft group (average 0.21 procedures/patient/year). So the autogenous fistula group had fewer intervention rate than prosthetic graft group (p=0.01) The risk factor of primary patency was diabetus combined with ischemic heart disease and the secondary patency's risk factor was age. CONCLUSION: Autogenous arteriovenous fistulas showed better performance compared to prosthetic grafts in terms of primary & secondary patency and incidence of interventions.

Can magnetic resonance angiogram be a reliable alternative for donor evaluation for laparoscopic nephrectomy?

BACKGROUND: While hand-assisted laparoscopic donor nephrectomy (HLDN) is less invasive, which can encourage kidney donation, it requires more exact information about the renal vascular anatomy because of its limited visual field during nephrectomy. MRA is also an attractive choice because of its minimal invasiveness; further, it is an outpatient-based procedure, it uses non-nephrotoxic contrast material and it has no radiation. The aim of our study was to evaluate the effectiveness of gadolinium enhanced three-dimensional MRA (GdE-3D MRA) in a group of potential live donors who were candidates for HLDN. METHODS: From September 2002 to December 2004, 40 potential live renal donors were evaluated prospectively with GdE-3D MRA, and this imaging modality was performed before the gold standard, the intra-arterial digital subtraction angiogram (IA-DSA), was carried out. All the images were reviewed in a blinded manner by the attending vascular radiologist. The MRA findings were compared with the DSA findings and the surgical findings as the reference methods. We evaluated the accuracy of MRA for imaging the renal architectures, and especially for imaging the renal accessory arteries and the early branching arteries that are important determinants for selection of the donor kidney. RESULTS: Both the MRA and DSA images showed consistent findings with the surgical findings in 92.5% of the 40 donors. There were no discrepant cases in depicting the main renal artery. MRA showed 100% specificity for imaging both the renal accessory arteries and the early branching arteries, when compared with the surgical findings. The kappa values for the MRA and DSA for the accessory arteries were all 0.66 compared with the intraoperative findings. MRA also depicted one huge renal cyst in one donor and many small renal cysts in the other donors that could not be imaged by DSA. There were no adverse events during the MRA procedure. None of the findings missed by MRA resulted in deleterious consequences at laparoscopic nephrectomy for the donor and graft. CONCLUSIONS: Our limited experience with GdE-3D MRA for imaging the renal structures in kidney donor evaluation for HLDN has been quite satisfactory.

Coronary-artery calcium scores using electron beam CT in patients with chronic renal failure.

We evaluated the risk of coronary-artery disease in patients with chronic renal failure (CRF) by measuring the coronary-artery calcium scores with electron beam CT (EBCT). A total of 81 CRF patients were divided into three groups; pre-dialysis (group I, n = 35), hemodialysis (group II, n = 31) and peritoneal dialysis (group III, n = 15). The several serum biochemical markers and calcium score levels by EBCT were determined. The Ca x P products were significantly higher in groups II (p < 0.05) and III (p < 0.01) than in group I. The serum calcium levels were significantly higher in group III than in both group I (p < 0.01) and II (p < 0.05). The serum calcium level in 15 patients with a calcium score > 400 was significantly higher than the 66 patients with a score < or =400 (p < 0.01). The calcium score was significantly higher in the 15 patients with cardiovascular complications than in the 66 patients without cardiovascular complications (628.9+/-904.8 vs. 150.4+/-350.9, p < 0.01). EBCT seemed to be a good diagnostic tool for evaluating the risk of coronary-artery disease ''noninvasively'' in CRF patients who are at increased risk of cardiovascular morbidity and mortality.