EspF Interacts with nucleation-promoting factors to recruit junctional proteins into pedestals for pedestal maturation and disruption of paracellular permeability

Many pathogenic bacteria subvert normal host cell processes by delivering effector proteins which mimic eukaryotic functions directly into target cells. EspF is a multifunctional protein injected into host cells by attaching and effacing pathogens, but its mechanism of action is not understood completely. In silico analyses of EspF revealed two key motifs: proline-rich domains and PDZ domain binding motifs. Such functional domains may allow EspF to act as an actin nucleation-promoting factor by mimicking host proteins. In agreement with these predictions, we found that EspF from rabbit enteropathogenic Escherichia coli (E22) participates in the regulation of actin polymerization by binding to a complex of proteins at the tight junctions (TJ). EspF bound to actin and profilin throughout the course of infection. However, after 2 h of infection, EspF also bound to the neural Wiskott-Aldrich syndrome protein and to the Arp2/3, zonula occludens-1 (ZO-1), and ZO-2 proteins. Moreover, EspF caused occludin, claudin, ZO-1, and ZO-2 redistribution and loss of transepithelial electrical resistance, suggesting that actin sequestration by EspF may cause local actin depolymerization leading to EspF-induced TJ disruption. Furthermore, EspF caused recruitment of these TJ proteins into the pedestals. An E22 strain lacking EspF did not cause TJ disruption and pedestals were smaller than those induced by the wild-type strain. Additionally, the pedestals were located mainly in the TJ. The overexpression of EspF caused bigger pedestals located along the length of the cells. Thus, actin sequestration by EspF allows the recruitment of junctional proteins into the pedestals, leading to the maturation of actin pedestals and the disruption of paracellular permeability.     

E-selectin gene S128R polymorphism is associated with poor prognosis in patients with stage II or III colorectal cancer

Some host-related factors may predict the risk of metastasis after surgery of colorectal cancer (CRC). The endothelial adhesion molecule E-selectin is implicated in the metastatic spread of CRC. We postulated that some polymorphisms within the E-selectin gene, especially the S128R polymorphism, may increase the risk of metastases by facilitating adhesion of tumour cells to the endothelium. We collected blood samples for DNA extraction from 264 patients treated for stage II or III CRC and from 310 healthy controls in order to assess three polymorphisms within the E-selectin gene (S128R, G98T and L554F) and one within the P-selectin gene (V640L). Genotypes were analysed by the allelic discrimination TaqMan real-time PCR assay. The S128R polymorphism was detected in 59 patients (22.3%) and was strictly correlated with the G98T polymorphism. In multivariate analysis, the S128R polymorphism was associated with shorter event-free survival (EFS) and overall survival (OS) in the whole population (EFS: P = .003, HR 1.82, 95% CI 1.23–2.70; OS: P < 10^–4, HR 4.31, 95% CI 2.46–10.99), in patients with stage II CRC(EFS: P = .04, HR 1.92, 95% CI 1.02–3.60; OS: P = .02, HR 4.44, 95% CI 1.16–17.03), and in patients with stage III CRC (EFS: P = .04, HR 1.68, 95% CI 1.01–2.80; OS: P = .001, HR 4.04, 95% CI 1.73–9.46). L554F and V640L polymorphisms had no prognostic value. The S128R polymorphism is a constitutional factor associated with a higher risk of relapse and death in patients treated for CRC. This polymorphism detection may permit better selection of patients suitable for adjuvant therapy, especially among those with stage II disease.     

Hyper-alkalinization without hyper-hydration for the prevention of high-dose methotrexate acute nephrotoxicity in patients with osteosarcoma

Purpose

To evaluate the reliability and renal safety of an original schedule of high-dose methotrexate (HDMTX) administration with hyper-alkalinization, and without hyper-hydration.

Methods

Patients with osteosarcoma received HDMTX (8–12 g/m²) as a 4-h infusion. Hypertonic 8.4% sodium bicarbonate was infused prior to HDMTX, then once daily for 3 days. Methotrexate serum concentrations were measured at hour 4 (Cmax), hour 24, hour 48, and hour 72. Urinary pH was measured on each miction. Serum creatinine was assessed on days 1, 3, and 8.

Results

Twenty-six patients (median age: 18 years, range: 15–25) received a total of 344 cycles of HDMTX, including 16 patients treated in an outpatient basis. Urinary pH remained constantly higher than 7.5 in all patients. Grade 1 creatininemia toxicity was observed in 31 cycles (9%), and grade 2 creatinine toxicity was observed in one patient. No episode of acute severe nephrotoxicity was observed. No significant worsening was observed in serum creatinine and calculated creatinine clearance from baseline to the end of therapy (P = 0.74). The main extra-renal toxicity was alkalinization-related hypokalemia from H48. No re-hospitalization was required.

Conclusion

Hyper-alkalinization appears an efficient and reliable method to prevent the acute renal toxicity of HDMTX and allows its safe administration in the outpatient setting.     

Gluten-free diet induces regression of T-Cell activation in the rectal mucosa of patients with celiac disease

Objective: An increase in the number of intraepithelial lymphocytes (IEL) in the rectal epithelium of patients with active celiac disease has been described. No data are available about how they vary during a gluten-free diet. The aim of the study was to assess the effect of a gluten-free diet on T-cell activation in the rectal mucosa of adult patients with celiac disease.
Methods: Frozen duodenal and rectal biopsies were available in four celiac patients (one male, three female, mean age 39 yr) both before and after 7 to 24 months on a gluten-free diet. Biopsy samples were stained using monoclonal antibodies directed against CD3, βF1, TcRδ1, CD25, and HLADR. Numbers of IEL were estimated by counting the peroxidase-stained cells per 100 epithelial cells. Four patients without histological abnormalities were used as control subjects.
Results: In the four patients with active celiac disease but in none of the controls, CD25 was expressed by both duodenal and rectal lamina propria cells and HLADR was expressed by duodenal (4/4) and rectal (2/4) epithelial cells. In addition, two patients with active celiac disease had features of lymphocytic colitis, i.e. , >20 IEL per 100 epithelial cells. After a gluten-free diet, the mean number of rectal CD3⁺βF1⁺ IEL decreased (9% vs 21%) and the expression of CD25 and HLADR was no longer present. These changes mirrored those found in the small intestinal biopsies.
Conclusion: These results suggest that in celiac disease, gluten-driven T-cell activation is not restricted to the proximal part of the intestine but is present on the whole intestinal length. Assessment of the effectiveness of a gluten-free diet through rectal biopsies warrants investigation, as it could lessen discomfort for patients and prove more cost-effective.     

Computed tomographic evidence of hepatic portal venous gas after blunt abdominal trauma does not necessitate surgery

Historically, hepatic portal venous gas (HPVG) seen on abdominal radiographic examination indicated serious intra-abdominal pathology requiring urgent operative intervention. The mortality attributed to HPVG is associated closely with its causative source rather than a direct effect of the presence of venous air and, therefore, the finding should be correlated with a careful clinical examination before any therapeutic endeavor. Fourteen cases of HPVG associated with blunt trauma have been reported over the past 20 years, and only half of these have resulted in surgery. We report the case of a 24-year-old woman who presented with no abdominal pathology other than HPVG after a severe motor vehicle crash. She was managed nonoperatively and made a successful recovery.     

Depressive symptoms, antidepressants and disability and future coronary heart disease and stroke events in older adults: the Three City Study

To investigate the association between baseline depressive symptoms and first fatal and non fatal coronary heart disease (CHD) and stroke in older adults, taking antidepressants and disability into account. In the Three City Study, a community-based prospective multicentric observational study cohort, 7,308 non-institutionalized men and women aged ≥65 years with no reported history of CHD, stroke or dementia, completed the 20-item Center for Epidemiologic Studies Depression Scale (CESD) questionnaire. First CHD and stroke events during follow-up were adjudicated by an independent expert committee. Hazard ratios (HRs) were estimated by Cox proportional hazard model. After a median follow-up of 5.3 years, 338 subjects had suffered a first non-fatal CHD or stroke event, and 82 had died from a CHD or stroke. After adjustment for study center, baseline socio-demographic characteristics, and conventional risk factors, depressive symptoms (CESD ≥ 16) were associated with fatal events only: fatal CHD plus stroke (HR = 2.50; 95 % CI 1.57–3.97), fatal CHD alone (n = 57; HR = 2.21 ; 95 %CI 1.27–3.87), and fatal stroke alone (n = 25; HR = 3.27; 95 % CI 1.42–7.52). These associations were even stronger in depressed subjects receiving antidepressants (HR = 4.17; 95 % CI 1.84–9.46) and in depressed subjects with impaired Instrumental Activities of Daily Living (HR = 8.93; 95 % CI 4.60–17.34). By contrast, there was no significant association with non fatal events (HR for non-fatal CHD or stroke = 0.94; 95 % CI 0.66–1.33). In non-institutionalized elderly subjects without overt CHD, stroke or dementia, depressive symptoms were selectively and robustly associated with first fatal CHD or stroke events.