Biology subjects

Hansen, K. C.

Publications and source records attributed to Hansen, K. C..

23 records · Page 2Linked to original sources

bioRxiv2020

Evaluation and refinement of sample preparation methods for extracellular matrix proteome coverage

The extracellular matrix is a key component of tissues, yet it is under-represented in proteomic datasets. Identification and evaluation of proteins in the extracellular matrix (ECM) has proved challenging due to the insolubility of many ECM proteins in traditional protein extraction buffers. Here we separate the decellularization and ECM extraction steps of several prominent methods for evaluation under real-world conditions. The results are used to optimize a two-fraction ECM extraction method. Approximately one dozen additional parameters are tested and recommendations for analysis based on overall ECM coverage or specific ECM classes are given. Compared to a standard in-solution digest, the optimized method yielded a 4-fold improvement in unique ECM peptide identifications. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=103 SRC="FIGDIR/small/391946v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@7e356aorg.highwire.dtl.DTLVardef@1e4f196org.highwire.dtl.DTLVardef@1b0e10forg.highwire.dtl.DTLVardef@187e040_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2020

Injectable Myocardial Matrix Hydrogel Mitigates Negative Left Ventricular Remodeling in a Chronic Myocardial Infarction Model

A first-in-man clinical study on a myocardial-derived decellularized extracellular matrix (ECM) hydrogel yielded evidence for potential efficacy in ischemic heart failure (HF) patients. However, little is understood about the mechanism of action in chronic myocardial infarction (MI). In this study we investigated efficacy and mechanism by which the myocardial matrix hydrogel can mitigate negative left ventricular (LV) remodeling in a chronic model of MI. Assessment of cardiac function via magnetic resonance imaging (MRI) demonstrated preservation of LV volumes and apical wall thickening. Differential gene expression analyses showed the matrix is able to prevent worsening HF in a small animal chronic MI model through modulation of the immune response, downregulation of pathways involved in HF progression and fibrosis, and upregulation of genes important for cardiac muscle contraction.

bioengineering↗

bioRxiv2020

Tissue specific muscle extracellular matrix hydrogel improves skeletal muscle regeneration in vivo over non-matched tissue source

Decellularized extracellular matrix (ECM) hydrogels present a novel, clinical intervention for a myriad of regenerative medicine applications. The source of ECM is typically the same tissue to which the treatment is applied; however, the need for tissue specific ECM sources has not been rigorously studied. We hypothesized that tissue specific ECM would improve regeneration through preferentially stimulating physiologically relevant processes (e.g. progenitor cell proliferation and differentiation). One of two decellularized hydrogels (tissue specific skeletal muscle or non mesoderm-derived lung) or saline were injected intramuscularly two days after notexin injection in mice (n=7 per time point) and muscle was harvested at days 5 and 14 for histological and gene expression analysis. Both injectable hydrogels were decellularized using the same detergent and were controlled for donor characteristics (i.e. species, age). At day 5, the skeletal muscle ECM hydrogel significantly increased the density of Pax7+ satellite cells in the muscle. Gene expression analysis at day 5 showed that skeletal muscle ECM hydrogels increased expression of genes implicated in muscle contractility. By day 14, skeletal muscle ECM hydrogels improved muscle regeneration over saline and lung ECM hydrogels as shown through a shift in fiber cross sectional area distribution towards larger fibers. This data indicates a potential role for muscle-specific regenerative capacity of decellularized, injectable muscle hydrogels. Further transcriptomic analysis of whole muscle mRNA indicates the mechanism of tissue specific ECM-mediated tissue repair may be immune and metabolism pathway-driven. Taken together, this suggests there is benefit in using tissue specific ECM for regenerative medicine applications.Competing Interest StatementKLC is co-founder, board member, consultant, receives income, and has equity interest in Ventrix, Inc.View Full Text

bioengineering↗

bioRxiv2020

Inflammation promotes tumor aggression by stimulating stromal cell-dependent collagen crosslinking and stromal stiffening

Collagen deposition and stromal stiffening accompany malignancy, compromise treatment, and promote tumor aggression. Clarifying the molecular nature of and the factors that regulate extracellular matrix stiffening in tumors should identify biomarkers to stratify patients for therapy and therapeutic interventions to improve outcome. We profiled lysyl hydroxylase- and lysyl oxidase-mediated collagen crosslinks and quantified the greatest abundance of total and complex collagen crosslinks in more aggressive human breast cancer subtypes with the stiffest stroma. These tissues also harbored the highest number of tumor-associated macrophages (TAM), whose therapeutic ablation not only reduced metastasis, but also concomitantly decreased accumulation of collagen crosslinks and stromal stiffening. Epithelial-targeted expression of the crosslinking enzyme lysyl oxidase had no impact on collagen crosslinking in PyMT mammary tumors, whereas stromal cell targeting did. Consistently, stromal cells in microdissected human tumors expressed the highest level of collagen crosslinking enzymes. Immunohistochemical analysis of a cohort of breast cancer patient biopsies revealed that stromal expression of lysyl hydroxylase two, an enzyme that induces hydroxylysine aldehyde-derived collagen crosslinks and stromal stiffening correlated significantly disease specific mortality. The findings link tissue inflammation, stromal cell-mediated collagen crosslinking and stiffening to tumor aggression and identify lysyl hydroxylase two as a novel stromal biomarker. SignificanceWe show infiltrating macrophages induce stromal fibroblast, and not epithelial, expression of collagen crosslinking enzymes that drive tumor stiffening. Stromal enzyme LH2 is significantly upregulated in breast cancer patients with the stiffest stroma, the most trivalent HLCCs and the worst prognosis, underscoring its potential as a biomarker and therapeutic target.

cancer biology↗

bioRxiv2018

Targeting glutamine metabolism and redox state for leukemia therapy

Acute myeloid leukemia (AML) is a hematological malignancy characterized by the accumulation of immature myeloid precursor cells. AML is poorly responsive to conventional genotoxic chemotherapy and a diagnosis of AML is usually fatal. More effective and less toxic forms of therapy are desperately needed. AML cells are known to be highly dependent on the amino acid glutamine for their survival. Here, we show that blocking glutamine metabolism through the use of a glutaminase inhibitor (CB-839) significantly impairs antioxidant glutathione production in multiple types of AML, resulting in accretion of mitochondrial reactive oxygen species (mitoROS) and apoptotic cell death. Moreover, glutaminase inhibition makes AML cells susceptible to adjuvant drugs that further perturb mitochondrial redox state, such as arsenic trioxide (ATO) and homoharringtonine (HHT). Indeed, the combination of ATO or HHT with CB-839 exacerbates mitoROS and apoptosis, and leads to more complete cell death in AML cell lines, primary AML patient samples and in vivo using mouse models of AML. In addition, these redox-targeted combination therapies are effective in eradicating acute lymphoblastic leukemia cells in vitro and in vivo. Thus, targeting glutamine metabolism in combination with drugs that perturb mitochondrial redox state represents an effective and potentially widely applicable therapeutic strategy for treating multiple types of leukemia.\n\nKey PointsO_LIGlutaminase inhibition commonly impairs glutathione metabolism and induces mitochondrial oxidative stress in acute myeloid leukemia cells\nC_LIO_LIA glutaminase inhibitor synergizes with pro-oxidant drugs in inducing apoptosis and eliminating leukemia cells in vitro and in vivo\nC_LI

cancer biology↗