Dynamic Micropatterning Reveals Substrate-Dependent Differences in the Geometric Control of Cell Polarization and Migration

Cells are highly dynamic and adopt variable shapes and sizes. These variations are biologically important but challenging to investigate in a spatiotemporally controlled manner. Micropatterning, confining cells on microfabricated substrates with defined geometries and molecular compositions, is a powerful tool for controlling cell shape and interactions. However, conventional binary micropatterns are static and fail to address dynamic changes in cell polarity, spreading, and migration. Here, a method for dynamic micropatterning is reported, where the non-adhesive surface surrounding adhesive micropatterns is rapidly converted to support specific cell-matrix interactions while allowing simultaneous imaging of the cells. The technique is based on ultraviolet photopatterning of biotinylated polyethylene glycol-grafted poly-L-lysine, and it is simple, inexpensive, and compatible with a wide range of streptavidin-conjugated ligands. Experiments using biotinylation-based dynamic micropatterns reveal that distinct extracellular matrix ligands and bivalent integrin-clustering antibodies support different degrees of front-rear polarity in human glioblastoma cells, which correlates to altered directionality and persistence upon release and migration on fibronectin. Unexpectedly, however, neither an asymmetric cell shape nor centrosome orientation can fully predict the future direction of migration. Taken together, biotinylation-based dynamic micropatterns allow easily accessible and highly customizable control over cell morphology and motility

Prognostic and diagnostic value of REG4 serum and tissue expression in pancreatic ductal adenocarcinoma

Expression of regenerating islet-derived protein 4 (REG4), a secretory protein involved in cell differentiation and proliferation, is upregulated in inflammatory bowel diseases and in many gastrointestinal malignancies. The prognostic significance of its expression in pancreatic ductal adenocarcinoma is unknown. Our aim was to investigate tumor tissue and serum REG4 expression in pancreatic ductal adenocarcinoma patients. We also evaluated as a control the diagnostic value of serum REG4 level in patients with chronic pancreatitis. Immunohistochemical expression of REG4 was evaluated in 154 surgical specimens and serum REG4 level in 130 samples from pancreatic ductal adenocarcinoma patients treated at Helsinki University Hospital, Finland, in 2000–2011. REG4 tissue and serum expression was assessed in relation to clinicopathological parameters and patient survival. A chronic pancreatitis control group comprised 34 patients who underwent pancreatic resection because of suspicion of malignancy. Significant survival differences were detectable in subgroups: in tumor stages IA–IIA, high serum REG4 level predicted worse survival (p=0.046). In patients with grade I tumor, positive tissue REG4 expression predicted better survival (p=0.006). In multivariate analysis, neither tissue nor serum REG4 expression was independent prognostic factors. Serum REG4 levels were higher in pancreatic ductal adenocarcinoma than in chronic pancreatitis (p=0.002), with diagnostic sensitivity of 45% and specificity of 91%. In logistic regression analysis, a multivariate model with REG4, CA19-9, and age provided sensitivity of 82% and specificity of 79%. REG4 tissue expression is a prognostic marker in subgroups of pancreatic ductal adenocarcinoma patients. Serum REG4 level might be useful in differential diagnosis between pancreatic ductal adenocarcinoma and chronic pancreatitis. © 2018, © The Author(s) 2018.Peer reviewe

Suppressed bone turnover in obesity: a link to energy metabolism? A case-control study

CONTEXT\nObservations in rodents suggest that osteocalcin (OC) participates in glucose metabolism. Based on human studies, it remains unclear whether circulating OC is simply a bone turnover marker (BTM) or also a mediator in interactions between the skeleton and glucose homeostasis.\nOBJECTIVE\nThe objective of the study was to determine the responses of BTMs, including OC, to oral glucose tolerance test (OGTT) in a case-control setting.\nDESIGN AND PATIENTS\nThirty-four normoglycemic young adults [mean age 19 y (SD 2.3)] with severe childhood-onset obesity and their gender- and age-matched nonobese controls underwent a standard 2-hour OGTT.\nMAIN OUTCOME MEASURES\nGlucose, insulin, and six BTMs including total and carboxylated OC (cOC) were determined at baseline and at 30, 60, 90, and 120 minutes during OGTT.\nRESULTS\nThe obese and control subjects were similar in height; the mean body mass indices were 40.4 and 21.9 kg/m(2), respectively. The homeostasis model assessment index was 2.7 times greater in the obese subjects. All BTMs, except bone-specific alkaline phophatase, were lower in the obese subjects compared with the controls: the differences at baseline were 40%, 35%, 17%, 31%, and 32% for N-terminal propeptides of type I collagen, cross-linked telopeptides of type I collagen, tartrate-resistant acid phosphatase, total OC, and carboxylated OC (P &lt; .05 for all) after adjusting for whole-body bone area. All BTMs decreased during OGTT. The relative values for the OGTT responses for total, but not for cOC (measured as area under the curve) differed between the two groups (P = .029 and P = .139, respectively): the decrease in total OC during the OGTT was less pronounced in the obese subjects. Responses in other BTMs were similar between the groups. No associations were observed between glucose metabolism and OCs during OGTT with linear regression.\nCONCLUSIONS\nBone turnover markers were substantially lower in obese subjects compared with controls. Total OC and cOC showed less pronounced decrease during the OGTT in obese subjects compared with controls, whereas other BTMs responded similarly in the two groups. The role of OC, if anything, in glucose homeostasis is indirect and may be mediated via other factors than glucose or insulin.</p

Fatal Outcome in Bacteremia is Characterized by High Plasma Cell Free DNA Concentration and Apoptotic DNA Fragmentation: A Prospective Cohort Study

INTRODUCTION: Recent studies have shown that apoptosis plays a critical role in the pathogenesis of sepsis. High plasma cell free DNA (cf-DNA) concentrations have been shown to be associated with sepsis outcome. The origin of cf-DNA is unclear. METHODS: Total plasma cf-DNA was quantified directly in plasma and the amplifiable cf-DNA assessed using quantitative PCR in 132 patients with bacteremia caused by Staphylococcus aureus, Streptococcus pneumoniae, ß-hemolytic streptococcae or Escherichia coli. The quality of cf-DNA was analyzed with a DNA Chip assay performed on 8 survivors and 8 nonsurvivors. Values were measured on days 1-4 after positive blood culture, on day 5-17 and on recovery. RESULTS: The maximum cf-DNA values on days 1-4 (n = 132) were markedly higher in nonsurvivors compared to survivors (2.03 vs 1.26 ug/ml, p<0.001) and the AUCROC in the prediction of case fatality was 0.81 (95% CI 0.69-0.94). cf-DNA at a cut-off level of 1.52 ug/ml showed 83% sensitivity and 79% specificity for fatal disease. High cf-DNA (>1.52 ug/ml) remained an independent risk factor for case fatality in a logistic regression model. Qualitative analysis of cf-DNA showed that cf-DNA displayed a predominating low-molecular-weight cf-DNA band (150-200 bp) in nonsurvivors, corresponding to the size of the apoptotic nucleosomal DNA. cf-DNA concentration showed a significant positive correlation with visually graded apoptotic band intensity (R = 0.822, p<0.001). CONCLUSIONS: Plasma cf-DNA concentration proved to be a specific independent prognostic biomarker in bacteremia. cf-DNA displayed a predominating low-molecular-weight cf-DNA band in nonsurvivors corresponding to the size of apoptotic nucleosomal DNA

Pilot study in human healthy volunteers on the use of magnetohydrodynamics in needle-free continuous glucose monitoring

The benefits of continuous glucose monitoring (CGM) in diabetes management are extensively documented. Yet, the broader adoption of CGM systems is limited by their cost and invasiveness. Current CGM devices, requiring implantation or the use of hypodermic needles, fail to offer a convenient solution. We have demonstrated that magnetohydrodynamics (MHD) is effective at extracting dermal interstitial fluid (ISF) containing glucose, without the use of needles. Here we present the first study of ISF sampling with MHD for glucose monitoring in humans. We conducted 10 glucose tolerance tests on 5 healthy volunteers and obtained a significant correlation between the concentration of glucose in ISF samples extracted with MHD and capillary blood glucose samples. Upon calibration and time lag removal, the data indicate a Mean Absolute Relative Difference (MARD) of 12.9% and Precision Absolute Relative Difference of 13.1%. In view of these results, we discuss the potential value and limitations of MHD in needle-free glucose monitoring.Peer reviewe

Liver Enzyme Abnormalities and Associated Risk Factors in HIV Patients on Efavirenz-Based HAART with or without Tuberculosis Co-Infection in Tanzania.

To investigate the timing, incidence, clinical presentation, pharmacokinetics and pharmacogenetic predictors for antiretroviral and anti-tuberculosis drug induced liver injury (DILI) in HIV patients with or without TB co-infection. A total of 473 treatment naïve HIV patients (253 HIV only and 220 with HIV-TB co-infection) were enrolled prospectively. Plasma efavirenz concentration and CYP2B6*6, CYP3A5*3, *6 and *7, ABCB1 3435C/T and SLCO1B1 genotypes were determined. Demographic, clinical and laboratory data were collected at baseline and up to 48 weeks of antiretroviral therapy. DILI case definition was according to Council for International Organizations of Medical Sciences (CIOMS). Incidence of DILI and identification of predictors was evaluated using Cox Proportional Hazards Model. The overall incidence of DILI was 7.8% (8.3 per 1000 person-week), being non-significantly higher among patients receiving concomitant anti-TB and HAART (10.0%, 10.7 per 1000 person-week) than those receiving HAART alone (5.9%, 6.3 per 1000 person-week). Frequency of CYP2B6*6 allele (p = 0.03) and CYP2B6*6/*6 genotype (p = 0.06) was significantly higher in patients with DILI than those without. Multivariate cox regression model indicated that CYP2B6*6/*6 genotype and anti-HCV IgG antibody positive as significant predictors of DILI. Median time to DILI was 2 weeks after HAART initiation and no DILI onset was observed after 12 weeks. No severe DILI was seen and the gain in CD4 was similar in patients with or without DILI. Antiretroviral and anti-tuberculosis DILI does occur in our setting, presenting early following HAART initiation. DILI seen is mild, transient and may not require treatment interruption. There is good tolerance to HAART and anti-TB with similar immunological outcomes. Genetic make-up mainly CYP2B6 genotype influences the development of efavirenz based HAART liver injury in Tanzanians

Plasma Heme Oxygenase-1 Concentration in Relation to Impaired Glucose Regulation in a Non-Diabetic Chinese Population

Background: Our previous study has recently shown that plasma heme oxygenase-1 (HO-1), a stress-responsive protein, is elevated in individuals with type 2 diabetes. The current study aimed to examine the association between plasma HO-1 concentration and impaired glucose regulation (IGR) in non-diabetic individuals. Methods: We conducted a case-control study including a total of 865 subjects (262 IGR individuals and 603 healthy controls) in a Chinese population. Basic characteristics were collected by questionnaire and standardized anthropometric measurements. Plasma HO-1 concentration was determined by ELISA. Results: Plasma HO-1 concentration was significantly increased in IGR individuals compared with healthy controls (1.34 (0.81–2.29) ng/ml vs 0.98 (0.56–1.55) ng/ml, P,0.001). After adjustment for age, sex, and BMI, the ORs for IGR in the highest quartile of plasma HO-1 concentrations, compared with the lowest, was 3.42 (95 % CI 2.11–5.54; P for trend,0.001). The trend remained significant even after additional adjustment for smoking, alcohol drinking, hypertension, family history of diabetes, lipid profiles and C-reactive protein. In the receiver-operating characteristic curve analysis, addition of plasma HO-1 concentration to a model with known risk factors yielded significantly improved discriminative value for IGR (area under the curves 0.75 (95 % CI 0.71–0.78) vs. 0.72 (95 % CI 0.69–0.76); P for difference = 0.026)