MT1-MMP collagenolytic activity is regulated through association with tetraspanin CD151 in primary endothelial cells

MT1-MMP plays a key role in endothelial function, as underscored by the angiogenic defects found in MT1-MMP deficient mice. We have studied the molecular interactions that underlie the functional regulation of MT1-MMP. At lateral endothelial cell junctions, MT1-MMP colocalizes with tetraspanin CD151 (Tspan 24) and its associated partner alpha3beta1 integrin. Biochemical and FRET analyses show that MT1-MMP, through its hemopexin domain, associates tightly with CD151, thus forming alpha3beta1 integrin/CD151/MT1-MMP ternary complexes. siRNA knockdown of HUVEC CD151 expression enhanced MT1-MMP-mediated activation of MMP2, and the same activation was seen in ex vivo lung endothelial cells isolated from CD151-deficient mice. However, analysis of collagen degradation in these experimental models revealed a diminished MT1-MMP enzymatic activity in confined areas around the cell periphery. CD151 knockdown affected both MT1-MMP subcellular localization and its inclusion into detergent-resistant membrane domains, and prevented biochemical association of the metalloproteinase with the integrin alpha3beta1. These data provide evidence for a novel regulatory role of tetraspanin microdomains on the collagenolytic activity of MT1-MMP and indicate that CD151 is a key regulator of MT1-MMP in endothelial homeostasis.     

Recruiting Rural Healthcare Providers Today: a Systematic Review of Training Program Success and Determinants of Geographic Choices

Background

Rural areas have historically struggled with shortages of healthcare providers; however, advanced communication technologies have transformed rural healthcare, and practice in underserved areas has been recognized as a policy priority. This systematic review aims to assess reasons for current providers’ geographic choices and the success of training programs aimed at increasing rural provider recruitment.

Methods

This systematic review (PROSPERO: CRD42015025403) searched seven databases for published and gray literature on the current cohort of US rural healthcare practitioners (2005 to March 2017). Two reviewers independently screened citations for inclusion; one reviewer extracted data and assessed risk of bias, with a senior systematic reviewer checking the data; quality of evidence was assessed using the GRADE approach.

Results

Of 7276 screened citations, we identified 31 studies exploring reasons for geographic choices and 24 studies documenting the impact of training programs. Growing up in a rural community is a key determinant and is consistently associated with choosing rural practice. Most existing studies assess physicians, and only a few are based on multivariate analyses that take competing and potentially correlated predictors into account. The success rate of placing providers-in-training in rural practice after graduation, on average, is 44% (range 20–84%; N = 31 programs). We did not identify program characteristics that are consistently associated with program success. Data are primarily based on rural tracks for medical residents.

Discussion

The review provides insight into the relative importance of demographic characteristics and motivational factors in determining which providers should be targeted to maximize return on recruitment efforts. Existing programs exposing students to rural practice during their training are promising but require further refining. Public policy must include a specific focus on the trajectory of the healthcare workforce and must consider alternative models of healthcare delivery that promote a more diverse, interdisciplinary combination of providers.

     

Prediction of mortality in community-living frail elderly people with long-term care needs

OBJECTIVES: To develop and validate a prognostic index for mortality in community‐living, frail elderly people. DESIGN: Cohort study of Program of All‐Inclusive Care for the Elderly (PACE) participants enrolled between 1988 and 1996. SETTING: Eleven PACE sites, a community‐based long‐term care program that cares for frail, chronically ill elderly people who meet criteria for nursing home placement. PARTICIPANTS: Three thousand eight hundred ninety‐nine PACE enrollees. The index was developed in 2,232 participants and validated in 1,667. MEASUREMENTS: Time to death was predicted using risk factors obtained from a geriatric assessment performed by the PACE interdisciplinary team at the time of enrollment. Risk factors included demographic characteristics, comorbid conditions, and functional status. RESULTS: The development cohort had a mean age of 79 (68% female, 40% white). The validation cohort had a mean age of 79 (76% female, 65% white). In the development cohort, eight independent risk factors of mortality were identified and weighted, using Cox regression, to create a risk score: male sex, 2 points; age (75–79, 2 points; 80–84, 2 points; ≥85, 3 points); dependence in toileting, 1 point; dependence in dressing (partial dependence, 1 point; full dependence, 3 points); malignant neoplasm, 2 points; congestive heart failure, 3 points; chronic obstructive pulmonary disease, 1 point; and renal insufficiency, 3 points. In the development cohort, respective 1‐ and 3‐year mortality rates were 6% and 21% in the lowest‐risk group (0–3 points), 12% and 36% in the middle‐risk group (4–5 points), and 21% and 54% in the highest‐risk group (>5 points). In the validation cohort, respective 1‐ and 3‐year mortality rates were 7% and 18% in the lowest‐risk group, 11% and 36% in the middle‐risk group, and 22% and 55% in the highest‐risk group. The area under the receiver operating characteristic curve for the point score was 0.66 and 0.69 in the development and validation cohorts, respectively. CONCLUSION: A multidimensional prognostic index was developed and validated using age, sex, functional status, and comorbidities that effectively stratifies frail, community‐living elderly people into groups at varying risk of mortality.     

Effects of chronic iodine excess in a cohort of long-term American workers in West Africa

A cross-sectional survey of 102 Peace Corps volunteers in Niger, West Africa, in 1998 had previously demonstrated a high rate of thyroid dysfunction and goiter attributable to excess iodine from their water filters. The Peace Corps volunteers were followed-up a mean of 30 wk after they ceased using iodine-based water filtration systems. Goiter was present in 44% of subjects during excess iodine ingestion and in 30% after removal of excess iodine. Mean serum iodine decreased from 293 micro g/liter during excess iodine ingestion to 84 micro g/liter after cessation of excess iodine. Mean total serum T(4) values increased from 100.4 to 113.3 nmol/liter (7.8 to 8.8 micro g/dl). Mean serum free T(4) increased from 32.2 to 34.7 pmol/liter (2.5 to 2.7 ng/dl). Mean serum TSH decreased from 4.9 to 1.8 mU/liter. Mean serum thyroid peroxidase antibody levels decreased from 33,000 to 22,000 IU/liter (33 to 22 IU/ml). We found that during prolonged excess iodine exposure there were marked increases in serum total iodine concentrations, and the prevalence of goiter, elevated serum TSH values, and elevated serum thyroid peroxidase antibody values increased. The prevalence of all abnormalities decreased after removal of excess iodine from the drinking water system.     

Flow cytometric analysis of intestinal intraepithelial lymphocytes in the diagnosis of refractory celiac sprue

The etiology of refractory celiac sprue (RCS) is unclear. In a high proportion of cases, the clonal nature of intestinal intraepithelial lymphocytes (IEL) can be demonstrated and a pathogenetic implication of intestinal IEL has been postulated. The prognosis of this subgroup of RCS is poor, with a high risk to develop an overt lymphoma and uncontrolled malabsorption despite steroid/immunosuppressive therapy. Cases with a relatively indolent clinical course, however, exist and their early diagnosis may be difficult. To gain insight into the pathogenic implication of intestinal IEL in refractory celiac sprue, we have performed an extensive phenotypic and functional characterization of clonal intestinal IEL in a patient with an indolent form of refractory celiac sprue, using multiparametric flow cytometry. The abnormal lymphocyte infiltrate lacked surface membrane expression of CD3/T-cell receptor (TCR) complexes (TCR(-), CD4(-), CD8(-), sCD3(-)), but contained intracellular CD3(epsilon) (CyCD3(+)) and surface CD103(+) and CD7(+). In particular, these cells showed a unique spontaneous ex-vivo cytokine secretion profile with an increased percentage of CD3(-) IEL containing TNF-alpha and IL-10, in the absence of IL-2, IL-4 and IFN-gamma. Altogether our results suggest that flow cytometry immunophenotyping of intestinal IEL, in cases suspected of celiac disease and their complicated forms, could be of great help in the correct diagnosis of RCS and the understanding of the immunopathogenic mechanisms of the disease and their clinical and/or therapeutical implications.     

Echocardiographic assessment of left ventricular midwall mechanics in spontaneously hypertensive rats.

AIMS: The present study was attempted to determine whether LV midwall mechanics yielded different conclusions about LV systolic function than the assessment of endocardial LV mechanics by echocardiography in spontaneously hypertensive rats (SHR). METHODS AND RESULTS: Thirty-six (18 Wistar normotensive (W), 18 [SHR]) anesthetized rats were studied with two-dimensional directed M-mode echocardiogram to analyze LV structure (LV diameter, left ventricular wall thickness and LV mass [LVM]) and LV function (endocardial shortening [ES] and midwall shortening [MS]). Measurements were made from three consecutive cardiac cycles on the M-mode tracings. There was no significant difference in LV dimension. LVM was higher in SHR (SHR: 595 +/- 111 mg, W: 413 +/- 83 mg--p < 0.01). ES was higher in SHR (SHR: 64.1 +/- 6%, w: 58.2 +/- 4%--p < 0.01), whereas no significant difference was found in MS (SHR: 24 +/- 4%, W: 27.6 +/- 4%--ns). Twelve of 18 (66%) SHR showed endocardial shortening higher than normally predicted, compared with 3/18 (16%) with observed enhanced MS (p < 0.01). CONCLUSION: These results suggest that the analysis of midwall mechanics by echo allows us to better understand the LV performance in SHR and that the exaggerated endocardial motion could not represent a really supernormal systolic performance.     

An altered composition of the microbiome in microscopic colitis is driven towards the composition in healthy controls by treatment with budesonide

Background and aim: Microscopic colitis (MC) is an inflammatory disease of the bowel, hypothetically induced by an immunologic response to a luminal microbial agent. We aimed to characterize the microbiome composition in MC and subtypes collagenous colitis (CC) and lymphocytic colitis (LC) and to identify a possible microbial effect of treatment.

Method: Stool samples were collected from MC patients prior to treatment, at 8 weeks (during treatment) and at 16 weeks (after treatment), and from healthy controls, not receiving treatment, at matched time-points. Microbiome composition was analyzed by sequencing of the 16S and 18S genes. Differences between patients and controls were analyzed by Shannon’s diversity index (mean, standard deviation (SD)) and principal coordinate analysis (PCoA) complemented with a permanova test of UniFrac distances.

Results: Ten LC patients, 10?CC patients and 10 controls were included. By PCoA, the bacterial composition in MC patients differed from controls at baseline (p?=?.02), but not during and after treatment (p?=?.09 and p?=?.33, respectively). At baseline, bacterial diversity was lower in MC patients compared to controls (2.5, SD: 0.5 vs 3.5, SD: 0.3, p?<?.05). Diversity in MC patients increased during (3.0, SD: 0.6) and after treatment and (2.9, SD: 0.5) compared with baseline (p?<?.01). Eukaryotes were detected in fewer samples from MC patients compared with controls (11/20 (55%) vs. 9/10 (90%), p?=?.06) with no effect of treatment.

Conclusion: Microbiome composition is altered in MC patients. During and after treatment with budesonide the microbiome composition in MC patients was driven towards the composition in healthy controls.     

Isotope Ratio Mass Spectrometry (IRMS) Versus Laser-Assisted Ratio Analyzer (LARA)

This study was carried out to compare the measurements and the diagnostic accuracy of the traditional expensive IRMS and the new economical LARA system using two doses of [¹³C]urea + two different test meals in patients undergoing upper gastrointestinal endoscopy, both before and after anti-Helicobacter treatment. A total of 354 dyspeptic patients underwent endoscopy with gastric biopsies to diagnose H. pylori infection by CLO-test and histology. No patients had taken antibiotics, bismuth, or antisecretory drugs in the 4 weeks before testing. After overnight fasting, breath samples were collected simultaneously in both plastic and glass tubes at baseline and at 30 and 60 min after urea ingestion. In 237 patients 100 mg [¹³C]urea + Ensure and in 117 patients 75 mg [¹³C]urea + citric acid were given. The test was also performed with the two urea dosages and meals in 67 and 64 infected patients, respectively, four weeks after anti-Helicobacter therapy. H. pylori was considered eradicated when both biopsy-based tests were negative. A value >5 was considered positive. Breath samples with insufficient CO₂ levels at both 30 and 60 min were excluded from final analysis (N = 37 in pre- and N = 8 in posttreatment). There was excellent agreement between overall values of the two machines with both [¹³C]urea 100 mg + Ensure and [¹³C]urea 75 mg + citric acid. The 95% CI of the difference against the mean was wider with the former (mean –1.3, +6.3, and –9.4) than with the latter urea dosage and test meal (mean –1.2, +5.2 and –8.1). LARA and IRMS were equally effective (P = NS) in distinguishing infected from uninfected patients before therapy using both doses of [¹³C]urea and test meals (sensitivity ranged from 95% to 99% and specificity from 95% to 97%). This good performance was maintained in the posttreatment phase (sensitivity ranged from 90% to 100% and specificity from 90% to 97%), without any statistical difference among the various combinations (P = NS). The LARA system is a valid alternative to IRMS in the diagnosis of H. pylori infection. Both machines provide highly reliable results after 30 min, so that the 60 min sample can be avoided. The dose of 75 mg + citric acid suffices to ensure an accurate UBT. The test performed with both devices and [¹³C]urea dosages is very effective also for posttherapy evaluation of H. pylori status.