Hydrogels for delivery of therapeutic compounds

In some aspects, methacrylate co-polymers crosslinked with an enzymatically cleavable peptide linker are provided and may be used for the oral delivery of a therapeutic. The peptide linker may be cleavable by an enzyme in the small intestine and may allow for the delivery of a therapeutic protein or nucleic acid to the small intestine. Also provided are methods of using the polymers for the treatment of a disease.Board of Regents, University of Texas Syste

Temperature-Dependence of Weibel-Palade Body Exocytosis and Cell Surface Dispersal of von Willebrand Factor and Its Propolypeptide

Background: Weibel-Palade bodies (WPB) are endothelial cell (EC) specific secretory organelles containing Von Willebrand factor (VWF). The temperature-dependence of Ca2+-driven WPB exocytosis is not known, although indirect evidence suggests that WPB exocytosis may occur at very low temperatures. Here we quantitatively analyse the temperature-dependence of Ca2+-driven WPB exocytosis and release of secreted VWF from the cell surface of ECs using fluorescence microscopy of cultured human ECs containing fluorescent WPBs. Principal Findings: Ca2+-driven WPB exocytosis occurred at all temperatures studied (7–37°C). The kinetics and extent of WPB exocytosis were strongly temperature-dependent: Delays in exocytosis increased from 0.92 s at 37°C to 134.2 s at 7°C, the maximum rate of WPB fusion decreased from 10.0±2.2 s−1 (37°C) to 0.80±0.14 s−1 (7°C) and the fractional extent of degranulation of WPBs in each cell from 67±3% (37°C) to 3.6±1.3% (7°C). A discrepancy was found between the reduction in Ca2+-driven VWF secretion and WPB exocytosis at reduced temperature; at 17°C VWF secretion was reduced by 95% but WPB exocytosis by 75–80%. This discrepancy arises because VWF dispersal from sites of WPB exocytosis is largely prevented at low temperature. In contrast VWF-propolypeptide (proregion) dispersal from WPBs, although slowed, was complete within 60–120 s. Novel antibodies to the cleaved and processed proregion were characterised and used to show that secreted proregion more accurately reports the secretion of WPBs at sub-physiological temperatures than assay of VWF itself. Conclusions : We report the first quantitative analysis of the temperature-dependence of WPB exocytosis. We provide evidence; by comparison of biochemical data for VWF or proregion secretion with direct analysis of WPB exocytosis at reduced temperature, that proregion is a more reliable marker for WPB exocytosis at reduced temperature, where VWF-EC adhesion is increased

Differential cargo mobilisation within Weibel-Palade bodies after transient fusion with the plasma membrane.

Inflammatory chemokines can be selectively released from Weibel-Palade bodies (WPBs) during kiss-and-run exocytosis. Such selectivity may arise from molecular size filtering by the fusion pore, however differential intra-WPB cargo re-mobilisation following fusion-induced structural changes within the WPB may also contribute to this process. To determine whether WPB cargo molecules are differentially re-mobilised, we applied FRAP to residual post-fusion WPB structures formed after transient exocytosis in which some or all of the fluorescent cargo was retained. Transient fusion resulted in WPB collapse from a rod to a spheroid shape accompanied by substantial swelling (>2 times by surface area) and membrane mixing between the WPB and plasma membranes. Post-fusion WPBs supported cumulative WPB exocytosis. To quantify diffusion inside rounded organelles we developed a method of FRAP analysis based on image moments. FRAP analysis showed that von Willebrand factor-EGFP (VWF-EGFP) and the VWF-propolypeptide-EGFP (Pro-EGFP) were immobile in post-fusion WPBs. Because Eotaxin-3-EGFP and ssEGFP (small soluble cargo proteins) were largely depleted from post-fusion WPBs, we studied these molecules in cells preincubated in the weak base NH4Cl which caused WPB alkalinisation and rounding similar to that produced by plasma membrane fusion. In these cells we found a dramatic increase in mobilities of Eotaxin-3-EGFP and ssEGFP that exceeded the resolution of our method (∼ 2.4 µm2/s mean). In contrast, the membrane mobilities of EGFP-CD63 and EGFP-Rab27A in post-fusion WPBs were unchanged, while P-selectin-EGFP acquired mobility. Our data suggest that selective re-mobilisation of chemokines during transient fusion contributes to selective chemokine secretion during transient WPB exocytosis. Selective secretion provides a mechanism to regulate intravascular inflammatory processes with reduced risk of thrombosis

The interplay between the Rab27A effectors Slp4-a and MyRIP controls hormone-evoked Weibel-Palade body exocytosis.

Weibel-Palade body (WPB) exocytosis underlies hormone-evoked VWF secretion from endothelial cells (ECs). We identify new endogenous components of the WPB: Rab3B, Rab3D, and the Rab27A/Rab3 effector Slp4-a (granuphilin), and determine their role in WPB exocytosis. We show that Rab3B, Rab3D, and Rab27A contribute to Slp4-a localization to WPBs. siRNA knockdown of Slp4-a, MyRIP, Rab3B, Rab3D, Rab27A, or Rab3B/Rab27A, or overexpression of EGFP-Slp4-a or EGFP-MyRIP showed that Slp4-a is a positive and MyRIP a negative regulator of WPB exocytosis and that Rab27A alone mediates these effects. We found that ECs maintain a constant amount of cellular Rab27A irrespective of the WPB pool size and that Rab27A (and Rab3s) cycle between WPBs and a cytosolic pool. The dynamic redistribution of Rab proteins markedly decreased the Rab27A concentration on individual WPBs with increasing WPB number per cell. Despite this, the probability of WPB release was independent of WPB pool size showing that WPB exocytosis is not determined simply by the absolute amount of Rab27A and its effectors on WPBs. Instead, we propose that the probability of release is determined by the fractional occupancy of WPB-Rab27A by Slp4-a and MyRIP, with the balance favoring exocytosis

Classification of Dengue Fever Patients Based on Gene Expression Data Using Support Vector Machines

Background: Symptomatic infection by dengue virus (DENV) can range from dengue fever (DF) to dengue haemorrhagic fever (DHF), however, the determinants of DF or DHF progression are not completely understood. It is hypothesised that host innate immune response factors are involved in modulating the disease outcome and the expression levels of genes involved in this response could be used as early prognostic markers for disease severity. Methodology/Principal Findings: mRNA expression levels of genes involved in DENV innate immune responses were measured using quantitative real time PCR (qPCR). Here, we present a novel application of the support vector machines (SVM) algorithm to analyze the expression pattern of 12 genes in peripheral blood mononuclear cells (PBMCs) of 28 dengue patients (13 DHF and 15 DF) during acute viral infection. The SVM model was trained using gene expression data of these genes and achieved the highest accuracy of ,85% with leave-one-out cross-validation. Through selective removal of gene expression data from the SVM model, we have identified seven genes (MYD88, TLR7, TLR3, MDA5, IRF3, IFN-a and CLEC5A) that may be central in differentiating DF patients from DHF, with MYD88 and TLR7 observed to be the most important. Though the individual removal of expression data of five other genes had no impact on the overall accuracy, a significant combined role was observed when the SVM model of the two main genes (MYD88 and TLR7) was re-trained to include the five genes, increasing the overall accuracy to ,96%. Conclusions/Significance: Here, we present a novel use of the SVM algorithm to classify DF and DHF patients, as well as to elucidate the significance of the various genes involved. It was observed that seven genes are critical in classifying DF and DHF patients: TLR3, MDA5, IRF3, IFN-a, CLEC5A, and the two most important MYD88 and TLR7. While these preliminary results are promising, further experimental investigation is necessary to validate their specific roles in dengue disease

Health impacts of parental migration on left-behind children and adolescents: a systematic review and meta-analysis.

BACKGROUND: Globally, a growing number of children and adolescents are left behind when parents migrate. We investigated the effect of parental migration on the health of left behind-children and adolescents in low-income and middle-income countries (LMICs). METHODS: For this systematic review and meta-analysis we searched MEDLINE, Embase, CINAHL, the Cochrane Library, Web of Science, PsychINFO, Global Index Medicus, Scopus, and Popline from inception to April 27, 2017, without language restrictions, for observational studies investigating the effects of parental migration on nutrition, mental health, unintentional injuries, infectious disease, substance use, unprotected sex, early pregnancy, and abuse in left-behind children (aged 0-19 years) in LMICs. We excluded studies in which less than 50% of participants were aged 0-19 years, the mean or median age of participants was more than 19 years, fewer than 50% of parents had migrated for more than 6 months, or the mean or median duration of migration was less than 6 months. We screened studies using systematic review software and extracted summary estimates from published reports independently. The main outcomes were risk and prevalence of health outcomes, including nutrition (stunting, wasting, underweight, overweight and obesity, low birthweight, and anaemia), mental health (depressive disorder, anxiety disorder, conduct disorders, self-harm, and suicide), unintentional injuries, substance use, abuse, and infectious disease. We calculated pooled risk ratios (RRs) and standardised mean differences (SMDs) using random-effects models. This study is registered with PROSPERO, number CRD42017064871. FINDINGS: Our search identified 10 284 records, of which 111 studies were included for analysis, including a total of 264 967 children (n=106 167 left-behind children and adolescents; n=158 800 children and adolescents of non-migrant parents). 91 studies were done in China and focused on effects of internal labour migration. Compared with children of non-migrants, left-behind children had increased risk of depression and higher depression scores (RR 1·52 [95% CI 1·27-1·82]; SMD 0·16 [0·10-0·21]), anxiety (RR 1·85 [1·36-2·53]; SMD 0·18 [0·11-0·26]), suicidal ideation (RR 1·70 [1·28-2·26]), conduct disorder (SMD 0·16 [0·04-0·28]), substance use (RR 1·24 [1·00-1·52]), wasting (RR 1·13 [1·02-1·24]) and stunting (RR 1·12 [1·00-1·26]). No differences were identified between left-behind children and children of non-migrants for other nutrition outcomes, unintentional injury, abuse, or diarrhoea. No studies reported outcomes for other infectious diseases, self-harm, unprotected sex, or early pregnancy. Study quality varied across the included studies, with 43% of studies at high or unclear risk of bias across five or more domains. INTERPRETATION: Parental migration is detrimental to the health of left-behind children and adolescents, with no evidence of any benefit. Policy makers and health-care professionals need to take action to improve the health of these young people. FUNDING: Wellcome Trust

Is interpregnancy interval associated with cardiovascular risk factors in later life? A cohort study

OBJECTIVES: Pregnancy represents a metabolic challenge to women; in a normal pregnancy, transient metabolic changes occur that support the needs of the growing fetus. It is possible that repeating this challenge within a relatively short amount of time may result in lasting damage to the woman's cardiovascular health. Conversely, it is also possible that a long interpregnancy interval (IPI) may reflect subfertility, which has been found to be associated with cardiovascular disease (CVD). We examine the associations of short and long IPI with measures of cardiovascular health. DESIGN: Prospective cohort. SETTING: Mothers of the Avon Longitudinal Study of Parents and Children (ALSPAC). PARTICIPANTS: Women with two live births in order to control for confounding by parity. OUTCOME MEASURES: Arterial distensibility, common carotid intima, adiposity, blood pressure, lipids, glucose, insulin, proinsulin, triglycerides, C reactive protein. RESULTS: 25% (n=3451) of ALSPAC mothers had provided sufficient data to determine full reproductive history—of these, 1477 had two live births, with 54% mothers having non-missing data on all variables required for our analyses. A total of 1268 mothers with IPI (interbirth interval minus 9 months’ gestation) had CVD risk factors measured/imputed at mean age 48 years. After adjusting for confounding, we found no association of either short (≤15 months) or long (>27 months) IPI and increased levels of cardiovascular risk factors. There was some suggestion that women with long and short IPIs had a more favourable lipid profile compared with women whose IPI was 16–27 months; however, the differences were small in magnitude and imprecisely estimated. CONCLUSIONS: This study does not support the hypothesis that either long or short IPI is a risk factor for later cardiovascular health

Is interpregnancy interval associated with cardiovascular risk factors in later life? A cohort study

OBJECTIVES: Pregnancy represents a metabolic challenge to women; in a normal pregnancy, transient metabolic changes occur that support the needs of the growing fetus. It is possible that repeating this challenge within a relatively short amount of time may result in lasting damage to the woman's cardiovascular health. Conversely, it is also possible that a long interpregnancy interval (IPI) may reflect subfertility, which has been found to be associated with cardiovascular disease (CVD). We examine the associations of short and long IPI with measures of cardiovascular health. DESIGN: Prospective cohort. SETTING: Mothers of the Avon Longitudinal Study of Parents and Children (ALSPAC). PARTICIPANTS: Women with two live births in order to control for confounding by parity. OUTCOME MEASURES: Arterial distensibility, common carotid intima, adiposity, blood pressure, lipids, glucose, insulin, proinsulin, triglycerides, C reactive protein. RESULTS: 25% (n=3451) of ALSPAC mothers had provided sufficient data to determine full reproductive history—of these, 1477 had two live births, with 54% mothers having non-missing data on all variables required for our analyses. A total of 1268 mothers with IPI (interbirth interval minus 9 months’ gestation) had CVD risk factors measured/imputed at mean age 48 years. After adjusting for confounding, we found no association of either short (≤15 months) or long (>27 months) IPI and increased levels of cardiovascular risk factors. There was some suggestion that women with long and short IPIs had a more favourable lipid profile compared with women whose IPI was 16–27 months; however, the differences were small in magnitude and imprecisely estimated. CONCLUSIONS: This study does not support the hypothesis that either long or short IPI is a risk factor for later cardiovascular health