A liquid fraction of extracellular matrix inhibits glioma cell viability in vitro and in vivo.

Suppressive effects of extracellular matrix (ECM) upon various cancers have been reported. Glioblastoma multiforme has poor prognosis and new therapies are desired. This work investigated the effects of a saline-soluble fraction of urinary bladder ECM (ECM-SF) upon glioma cells. Viability at 24 hours in 1, 5, or 10 mg/mL ECM-SF-spiked media was evaluated in primary glioma cells (0319, 1015, 1119), glioma cell lines (A172, T98G, U87MG, C6), and brain cell lines (HCN-2, HMC3). Viability universally decreased at 5 and 10 mg/mL with U87MG, HCN-2, and HCM3 being least sensitive. Apoptosis in 0319 and 1119 cells was confirmed via NucView 488. Bi-weekly intravenous injection of ECM-SF (120 mg/kg) for 10 weeks in Sprague-Dawley rats did not affect weight, temperature, complete blood count, or multi-organ histology (N = 5). Intratumoral injection of ECM-SF (10 uL of 30 mg/mL) at weeks 2-4 post C6 inoculation in Wistar rats increased median survival from 24.5 to 51 days (hazard ratio for death 0.22) and decreased average tumor volume at time of death from 349 mm3 to 90 mm3 over 10 weeks (N = 6). Mass spectrometry identified 2,562 protein species in ECM-SF, parent ECM, and originating tissue. These results demonstrate the suppressive effects of ECM on glioma and warrant further study.

Lipidomics and RNA sequencing reveal a novel subpopulation of nanovesicle within extracellular matrix biomaterials.

Biomaterials composed of extracellular matrix (ECM) provide both mechanical support and a reservoir of constructive signaling molecules that promote functional tissue repair. Recently, matrix-bound nanovesicles (MBVs) have been reported as an integral component of ECM bioscaffolds. Although liquid-phase extracellular vesicles (EVs) have been the subject of intense investigation, their similarity to MBV is limited to size and shape. Liquid chromatography-mass spectrometry (LC-MS)-based lipidomics and redox lipidomics were used to conduct a detailed comparison of liquid-phase EV and MBV phospholipids. Combined with comprehensive RNA sequencing and bioinformatic analysis of the intravesicular cargo, we show that MBVs are a distinct and unique subpopulation of EV and a distinguishing feature of ECM-based biomaterials. The results begin to identify the differential biologic activities mediated by EV that are secreted by tissue-resident cells and deposited within the ECM.

An unbiased approach elucidates variation in (S)-(+)-linalool, a context-specific mediator of a tri-trophic interaction in wild tobacco.

Plant volatile organic compounds (VOCs) mediate many interactions, and the function of common VOCs is especially likely to depend on ecological context. We used a genetic mapping population of wild tobacco, Nicotiana attenuata, originating from a cross of 2 natural accessions from Arizona and Utah, separated by the Grand Canyon, to dissect genetic variation controlling VOCs. Herbivory-induced leaf terpenoid emissions varied substantially, while green leaf volatile emissions were similar. In a field experiment, only emissions of linalool, a common VOC, correlated significantly with predation of the herbivore Manduca sexta by native predators. Using quantitative trait locus mapping and genome mining, we identified an (S)-(+)-linalool synthase (NaLIS). Genome resequencing, gene cloning, and activity assays revealed that the presence/absence of a 766-bp sequence in NaLIS underlies the variation of linalool emissions in 26 natural accessions. We manipulated linalool emissions and composition by ectopically expressing linalool synthases for both enantiomers, (S)-(+)- and (R)-(-)-linalool, reported to oppositely affect M. sexta oviposition, in the Arizona and Utah accessions. We used these lines to test ovipositing moths in increasingly complex environments. The enantiomers had opposite effects on oviposition preference, but the magnitude of the effect depended strongly both on plant genetic background, and complexity of the bioassay environment. Our study reveals that the emission of linalool, a common VOC, differs by orders-of-magnitude among geographically interspersed conspecific plants due to allelic variation in a linalool synthase, and that the response of a specialist herbivore to linalool depends on enantiomer, plant genotype, and environmental complexity.

Human Testis Extracellular Matrix Enhances Human Spermatogonial Stem Cell Survival In Vitro.

IMPACT STATEMENT: This study developed and characterized human testis extracellular matrix (htECM) and porcine testis ECM (ptECM) for testing in human spermatogonial stem cell (hSSC) culture. Results confirmed the hypothesis that ECM from the homologous species (human) and homologous tissue (testis) is optimal for maintaining hSSCs. We describe a simplified feeder-free, serum-free condition for future iterative testing to achieve the long-term goal of stable hSSC cultures. To facilitate analysis and understand the fate of hSSCs in culture, we describe a multiparameter, high-throughput, quantitative flow cytometry approach to rapidly count undifferentiated spermatogonia, differentiated spermatogonia, apoptotic spermatogonia, and proliferative spermatogonia in hSSC cultures.

Cytocompatibility and mechanical properties of surgical sealants for cardiovascular applications.

OBJECTIVES: The present study compared physical, mechanical, and biologic characteristics of 4 clinically available surgical sealants for cardiovascular repair. METHODS: BioGlue (Cryolife Inc, Kennesaw, Ga), PreveLeak (Mallinckrodt Pharmaceuticals, St Louis, Mo), Tridyne VS (BD, Franklin Lakes, NJ), and Coseal (Baxter Healthcare Corporation, Westlake Village, Calif) were compared for the following properties: hydrated swelling, cytocompatibility, burst strength, biaxial stretching (elasticity), and in vitro degradation. RESULTS: Sealants showed a wide range of swelling upon hydration. By gravimetric and volumetric measurement, swelling was greatest for Coseal followed by Tridyne VS, BioGlue, and PreveLeak. Tridyne VS was the most cytocompatible based on Alamar Blue assay results, supporting 85% cell survival compared with 36% to 39% survival with the other sealants. All sealants withstood pressure above mean arterial pressure (70-110 mm Hg) and physiologic systolic blood pressure (90-140 mm Hg) in an ex vivo arterial flow burst model; lowest peak pressure at failure was PreveLeak at 235 ± 48 mm Hg, and highest peak pressure at failure was BioGlue at 596 ± 72 mm Hg. Biaxial tensile testing showed no differences in elasticity between ex vivo porcine aorta and carotid arteries and Tridyne VS or Coseal, and BioGlue and PreveLeak were significantly stiffer. In vitro degradation time for Coseal was 6 days and 21 days for Tridyne VS. No degradation was observed in BioGlue or PreveLeak for 30 days. CONCLUSIONS: Although all sealants withstood supraphysiologic arterial pressure, there were differences in characteristics that may be important in clinical outcome. Coseal degradation time was short compared with other sealants, whereas BioGlue and PreveLeak showed a significant compliance mismatch with native porcine carotid artery. Tridyne VS was significantly more cytocompatible than the other 3 sealants.

Isolation and Host Range of Bacteriophage with Lytic Activity against Methicillin-Resistant Staphylococcus aureus and Potential Use as a Fomite Decontaminant.

Staphylococcus aureus (SA) is a commensal bacterium and opportunistic pathogen commonly associated with humans and is capable of causing serious disease and death including sepsis, pneumonia, and meningitis. Methicillin-resistant SA (MRSA) isolates are typically resistant to many available antibiotics with the common exception of vancomycin. The presence of vancomycin resistance in some SA isolates combined with the current heavy use of vancomycin to treat MRSA infections indicates that MRSA may achieve broad resistance to vancomycin in the near future. New MRSA treatments are clearly needed. Bacteriophages (phages) are viruses that infect bacteria, commonly resulting in death of the host bacterial cell. Phage therapy entails the use of phage to treat or prevent bacterial infections. In this study, 12 phages were isolated that can replicate in human SA and/or MRSA isolates as a potential way to control these infections. 5 phage were discovered through mitomycin C induction of prophage and 7 others as extracellular viruses. Primary SA strains were also isolated from environmental sources to be used as tools for phage discovery and isolation as well as to examine the target cell host range of the phage isolates by spot testing. Primary isolates were tested for susceptibility to oxacillin in order to determine which were MRSA. Experiments were performed to assess the host range and killing potential of newly discovered phage, and significant reductions in bacterial load were detected. We explored the utility of some phage to decontaminate fomites (glass and cloth) and found a significant reduction in colony forming units of MRSA following phage treatment, including tests of a phage cocktail against a cocktail of MRSA isolates. Our findings suggest that phage treatment can be used as an effective tool to decontaminate human MRSA from both hard surfaces and fabrics.