Deciphering the relative roles of matrix metalloproteinase‐ and plasmin‐mediated matrix degradation during capillary morphogenesis using engineered hydrogels

Extracellular matrix (ECM) remodeling is essential for the process of capillary morphogenesis. Here we employed synthetic poly(ethylene glycol) (PEG) hydrogels engineered with proteolytic specificity to either matrix metalloproteinases (MMPs), plasmin, or both to investigate the relative contributions of MMP‐ and plasmin‐mediated ECM remodeling to vessel formation in a 3D‐model of capillary self‐assembly analogous to vasculogenesis. We first demonstrated a role for both MMP‐ and plasmin‐mediated mechanisms of ECM remodeling in an endothelial‐fibroblast co‐culture model of vasculogenesis in fibrin hydrogels using inhibitors of MMPs and plasmin. When this co‐culture model was employed in engineered PEG hydrogels with selective protease sensitivity, we observed robust capillary morphogenesis only in MMP‐sensitive matrices. Fibroblast spreading in plasmin‐selective hydrogels confirmed this difference was due to protease preference by endothelial cells, not due to limitations of the matrix itself. In hydrogels engineered with crosslinks that were dually susceptible to MMPs and plasmin, capillary morphogenesis was unchanged. These findings highlight the critical importance of MMP‐mediated degradation during vasculogenesis and provide strong evidence to justify the preferential selection of MMP‐degradable peptide crosslinkers in synthetic hydrogels used to study vascular morphogenesis and promote vascularization. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B:2507–2516, 2019.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/151850/1/jbmb34341_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/151850/2/jbmb34341.pd

Bacterial Thymidine Kinase as a Non-Invasive Imaging Reporter for Mycobacterium tuberculosis in Live Animals

Bacteria can be selectively imaged in experimentally-infected animals using exogenously administered 1-(2'deoxy-2'-fluoro-beta-D-arabinofuranosyl)-5-[(125)I]-iodouracil ([(125)I]-FIAU), a nucleoside analog substrate for bacterial thymidine kinase (TK). Our goal was to use this reporter and develop non-invasive methods to detect and localize Mycobacterium tuberculosis.We engineered a M. tuberculosis strain with chromosomally integrated bacterial TK under the control of hsp60 -- a strong constitutive mycobacterial promoter. [(125)I]FIAU uptake, antimicrobial susceptibilities and in vivo growth characteristics were evaluated for this strain. Using single photon emission computed tomography (SPECT), M. tuberculosis P(hsp60) TK strain was evaluated in experimentally-infected BALB/c and C3HeB/FeJ mice using the thigh inoculation or low-dose aerosol infection models. M. tuberculosis P(hsp60) TK strain actively accumulated [(125)I]FIAU in vitro. Growth characteristics of the TK strain and susceptibility to common anti-tuberculous drugs were similar to the wild-type parent strain. M. tuberculosis P(hsp60) TK strain was stable in vivo and SPECT imaging could detect and localize this strain in both animal models tested.We have developed a novel tool for non-invasive assessment of M. tuberculosis in live experimentally-infected animals. This tool will allow real-time pathogenesis studies in animal models of TB and has the potential to simplify preclinical studies and accelerate TB research

Single Dose Pharmacokinetics of Oral Tenofovir in Plasma, Peripheral Blood Mononuclear Cells, Colonic Tissue, and Vaginal Tissue

HIV seroconversion outcomes in preexposure prophylaxis (PrEP) trials of oral tenofovir (TFV)-containing regimens are highly sensitive to drug concentration, yet less-than-daily dosing regimens are under study. Description of TFV and its active moiety, TFV diphosphate (TFV-DP), in blood, vaginal tissue, and colon tissue may guide the design and interpretation of PrEP clinical trials. Six healthy women were administered a single oral dose of 300 mg tenofovir disoproxil fumarate (TDF) and 4.3 mg (12.31 MBq, 333 μCi) [superscript 14]C-TDF slurry. Blood was collected every 4 h for the first 24 h, then at 4, 8, 11, and 15 days postdosing. Colonic and vaginal samples (tissue, total and CD4+ cells, luminal fluid and cells) were collected 1, 8 and 15 days postdose. Samples were analyzed for TFV and TFV-DP. Plasma TFV demonstrated triphasic decay with terminal elimination half-life median [interquartile range (IQR)] 69 h (58–77). Peripheral blood mononuclear cell (PBMC) TFV-DP demonstrated biphasic peaks (median 12 h and 96 h) followed by a terminal 48 h (38–76) half-life; C[subscript max] was 20 fmol/million cells (2–63). One day postdose, the TFV-DP paired colon:vaginal tissue concentration ratio was 1 or greater in all subjects' tissue homogenates, median 124 (range 1–281), but was not sustained. The ratio was lower and more variable in cells extracted from tissue. Among all sample types, TFV and TFV-DP half-life ranged from 23 to 139 h. PBMC TFV-DP rose slowly in the hours after dosing indicating that success with exposure-driven dosing regimens may be sensitive to timing of the dose prior to exposure. Colonic tissue homogenate TFV-DP concentrations were greater than in vaginal homogenate at 24 h, but not in cells extracted from tissue. These and the other pharmacokinetic findings will guide the interpretation and design of future PrEP trials.National Center for Advancing Translational Sciences (U.S.) (Grant UL1RR025005)National Institutes of Health (U.S.)National Institutes of Health (U.S.) (Roadmap for Medical Research

Adaptive step ODE algorithms for the 3D simulation of electric heart activity with graphics processing units

In this paper we studied the implementation and performance of adaptive step methods for large systems of ordinary differential equations systems in graphics processing units, focusing on the simulation of three-dimensional electric cardiac activity. The Rush-Larsen method was applied in all the implemented solvers to improve efficiency. We compared the adaptive methods with the fixed step methods, and we found that the fixed step methods can be faster while the adaptive step methods are better in terms of accuracy and robustness. (c) 2013 Elsevier Ltd. All rights reserved.This work has been partially funded by Universitat Politecnica de Valencia through Programa de Apoyo a la InvestigaciOn y Desarrollo (PAID-06-11) and (PAID-05-12), by Generalitat Valenciana through projects PROMETEO/2009/013 and Ayudas para la realizacion de proyectos de I+D para grupos de investigacion emergentes GV/2012/039, and by Ministerio Espafiol de Economia y Competitividad through project TEC2012-38142-004.García Mollá, VM.; Liberos Mascarell, A.; Vidal Maciá, AM.; Guillem Sánchez, MS.; Millet Roig, J.; González Salvador, A.; Martínez Zaldívar, FJ.... (2014). Adaptive step ODE algorithms for the 3D simulation of electric heart activity with graphics processing units. Computers in Biology and Medicine. 44:15-26. https://doi.org/10.1016/j.compbiomed.2013.10.023S15264

Near-infrared molecular imaging of tumors via chemokine receptors CXCR4 and CXCR7

The chemokine CXCL12/SDF-1 and its receptors CXCR4 and CXCR7 play a major role in tumor invasion, proliferation and metastasis. Since both receptors are overexpressed on distinct tumor cells and on the tumor vasculature, we evaluated their potential as targets for detection of cancers by molecular imaging. We synthesized conjugates of CXCL12 and the near-infrared (NIR) fluorescent dye IRDye®800CW, tested their selectivity, sensitivity and biological activity in vitro and their feasibility to visualize tumors in vivo. Purified CXCL12-conjugates detected in vitro as low as 500 A764 human glioma cells or MCF-7 breast cancer cells that express CXCR7 alone or together with CXCR4. Binding was time- and concentration-dependent, and the label could be competitively displaced by the native peptide. Control conjugates with bovine serum albumin or lactalbumin failed to label the cells. In mice, the conjugate distributed rapidly. After 1–92 h, subcutaneous tumors of human MCF-7 and A764 cells in immunodeficient mice were detected with high sensitivity. Background was observed in particular in liver within the first 24 h, but also skull and hind limbs yielded some background. Overall, fluorescent CXCL12-conjugates are sensitive and selective probes to detect solid and metastatic tumors by targeting tumor cells and tumor vasculature

AMPK Modulation Ameliorates Dominant Disease Phenotypes of CTRP5 Variant in Retinal Degeneration

Late-onset retinal degeneration (L-ORD) is an autosomal dominant disorder caused by a missense substitution in CTRP5. Distinctive clinical features include sub-retinal pigment epithelium (RPE) deposits, choroidal neovascularization, and RPE atrophy. In induced pluripotent stem cells-derived RPE from L-ORD patients (L-ORD-iRPE), we show that the dominant pathogenic CTRP5 variant leads to reduced CTRP5 secretion. In silico modeling suggests lower binding of mutant CTRP5 to adiponectin receptor 1 (ADIPOR1). Downstream of ADIPOR1 sustained activation of AMPK renders it insensitive to changes in AMP/ATP ratio resulting in defective lipid metabolism, reduced Neuroprotectin D1(NPD1) secretion, lower mitochondrial respiration, and reduced ATP production. These metabolic defects result in accumulation of sub-RPE deposits and leave L-ORD-iRPE susceptible to dedifferentiation. Gene augmentation of L-ORD-iRPE with WT CTRP5 or modulation of AMPK, by metformin, re-sensitize L-ORD-iRPE to changes in cellular energy status alleviating the disease cellular phenotypes. Our data suggests a mechanism for the dominant behavior of CTRP5 mutation and provides potential treatment strategies for L-ORD patients. © 2021, This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply

Detection of paralytic shellfish toxins in mussels and oysters using the qualitative neogen lateral-flow immunoassay: an interlaboratory study

Paralytic shellfish toxins (PSTs) in bivalve molluscs represent a public health risk and are controlled via compliance with a regulatory limit of 0.8 mg saxitoxin (STX)center dot 2HCl equivalents per kilogram of shellfish meat (eq/kg). Shellfish industries would benefit from the use of rapid immunological screening tests for PSTs to be used for regulation, but to date none have been fully validated. An interlaboratory study involving 16 laboratories was performed to determine the suitability of the Neogen test to detect PSTs in mussels and oysters. Participants performed the standard protocol recommended by the manufacturer and a modified protocol with a conversion step to improve detection of gonyautoxin 1&4. The statistical analysis showed that the protocols had good homogeneity across all laboratories, with satisfactory repeatability, laboratory, and reproducibility variation near the regulatory level. The mean probability of detection (POD) at 0.8 mg STX center dot 2HCl eq/kg using the standard protocol in mussels and oysters was 0.966 and 0.997, respectively, and 0.968 and 0.966 using the modified protocol. The estimated LOD in mussels was 0.316 mg STX center dot 2HCl eq/kg with the standard and 0.682 mg STX center dot 2HCl eq/kg with the modified protocol, and 0.710 and 0.734 mg STX center dot 2HCl eq/kg for oysters, respectively. The Neogen test may be acceptable for regulatory purposes for oysters in accordance with European Commission directives in which the standard protocol provides, at the regulatory level, a probability of a negative response of 0.033 on 95% of occasions. Its use for mussels is less consistent at the regulatory level due to the wide prediction interval around the POD

Advances and Prospect of Nanotechnology in Stem Cells

In recent years, stem cell nanotechnology has emerged as a new exciting field. Theoretical and experimental studies of interaction between nanomaterials or nanostructures and stem cells have made great advances. The importance of nanomaterials, nanostructures, and nanotechnology to the fundamental developments in stem cells-based therapies for injuries and degenerative diseases has been recognized. In particular, the effects of structure and properties of nanomaterials on the proliferation and differentiation of stem cells have become a new interdisciplinary frontier in regeneration medicine and material science. Here we review some of the main advances in this field over the past few years, explore the application prospects, and discuss the issues, approaches and challenges, with the aim of improving application of nanotechnology in the stem cells research and development