Self-oxygenation of engineered living tissues orchestrates osteogenic commitment of mesenchymal stem cells.

Oxygenating biomaterials can alleviate anoxic stress, stimulate vascularization, and improve engraftment of cellularized implants. However, the effects of oxygen-generating materials on tissue formation have remained largely unknown. Here, we investigate the impact of calcium peroxide (CPO)-based oxygen-generating microparticles (OMPs) on the osteogenic fate of human mesenchymal stem cells (hMSCs) under a severely oxygen deficient microenvironment. To this end, CPO is microencapsulated in polycaprolactone to generate OMPs with prolonged oxygen release. Gelatin methacryloyl (GelMA) hydrogels containing osteogenesis-inducing silicate nanoparticles (SNP hydrogels), OMPs (OMP hydrogels), or both SNP and OMP (SNP/OMP hydrogels) are engineered to comparatively study their effect on the osteogenic fate of hMSCs. OMP hydrogels associate with improved osteogenic differentiation under both normoxic and anoxic conditions. Bulk mRNAseq analyses suggest that OMP hydrogels under anoxia regulate osteogenic differentiation pathways more strongly than SNP/OMP or SNP hydrogels under either anoxia or normoxia. Subcutaneous implantations reveal a stronger host cell invasion in SNP hydrogels, resulting in increased vasculogenesis. Furthermore, time-dependent expression of different osteogenic factors reveals progressive differentiation of hMSCs in OMP, SNP, and SNP/OMP hydrogels. Our work demonstrates that endowing hydrogels with OMPs can induce, improve, and steer the formation of functional engineered living tissues, which holds potential for numerous biomedical applications, including tissue regeneration and organ replacement therapy.

Aberrant Expression of ACO1 in Vasculatures Parallels Progression of Idiopathic Pulmonary Fibrosis.

Rationale: Idiopathic pulmonary fibrosis (IPF) is characterized by mitochondrial dysfunction. However, details about the non-mitochondrial enzymes that sustain the proliferative nature of IPF are unclear. Aconitases are a family of enzymes that sustain metabolism inside and outside mitochondria. It is hypothesized that aconitase 1 (ACO1) plays an important role in the pathogenesis of IPF given that ACO1 represents an important metabolic hub in the cytoplasm. Objectives: To determine if ACO1 expression in IPF lungs shows specific patterns that may be important in the pathogenesis of IPF. To determine the similarities and differences in ACO1 expression in IPF, bleomycin-treated, and aging lungs. Methods: ACO1 expression in IPF lungs were characterized and compared to non-IPF controls by western blotting, immunostaining, and enzymatic activity assay. ACO1-expressing cell types were identified by multicolor immunostaining. Using similar methods, the expression profiles of ACO1 in IPF lungs versus bleomycin-treated and aged mice were investigated. Measurements and main results: Lower lobes of IPF lungs, unlike non-IPF controls, exhibit significantly high levels of ACO1. Most of the signals colocalize with von Willebrand factor (vWF), a lineage marker for vascular endothelial cells. Bleomycin-treated lungs also show high ACO1 expressions. However, most of the signals colocalize with E-cadherin and/or prosurfactant protein C, representative epithelial cell markers, in remodeled areas. Conclusions: A characteristic ACO1 expression profile observed in IPF vasculatures may be a promising diagnostic target. It also may give clues as to how de novo angiogenesis contributes to the irreversible nature of IPF.

COMMD3:BMI1 Fusion and COMMD3 Protein Regulate C-MYC Transcription: Novel Therapeutic Target for Metastatic Prostate Cancer.

Gene rearrangement is reported to be associated to the aggressive phenotype and poor prognosis in prostate cancer. We identified a gene fusion between a transcription repressor (BMI1) and transcriptional factor (COMMD3) in human prostate cancer. We show that COMMD3:BMI1 fusion expression is significantly increased in prostate cancer disease in an order: normal tissue < primary < metastatic tumors (Mets). Although elevated TMPRSS-ERG/ETV fusion is reported in prostate cancer, we identified a subtype of Mets exhibiting low TMPRSS:ETV and high COMMD3:BMI1 We delineated the mechanism and function of COMMD3 and COMMD3:BMI1 in prostate cancer. We show that COMMD3 level is elevated in prostate cancer cell models, PDX models (adenocarcinoma, NECaP), and Mets. The analysis of TCGA/NIH/GEO clinical data showed a positive correlation between increased COMMD3 expression to the disease recurrence and poor survival in prostate cancer. We show that COMMD3 drives proliferation of normal cells and promotes migration/invasiveness of neoplastic cells. We show that COMMD3:BMI1 and COMMD3 regulate C-MYC transcription and C-MYC downstream pathway. The ChIP analysis showed that COMMD3 protein is recruited at the promoter of C-MYC gene. On the basis of these data, we investigated the relevance of COMMD3:BMI1 and COMMD3 as therapeutic targets using in vitro and xenograft mouse models. We show that siRNA-mediated targeting of COMMD3:BMI1 and COMMD3 significantly decreases (i) C-MYC expression in BRD/BET inhibitor-resistant cells, (ii) proliferation/invasion in vitro, and (iii) growth of prostate cancer cell tumors in mice. The IHC analysis of tumors confirmed the targeting of COMMD3-regulated molecular pathway under in vivo conditions. We conclude that COMMD3:BMI1 and COMMD3 are potential progression biomarkers and therapeutic targets of metastatic prostate cancer.

Atomic resolution crystal structure of glutaredoxin 1 from Plasmodium falciparum and comparison with other glutaredoxins.

Glutaredoxins (Grxs) are redox proteins that use glutathione ((γ)Glu-Cys-Gly; GSH) as a cofactor. Plasmodium falciparum has one classic dithiol (CXXC) glutaredoxin (glutaredoxin 1; PfGrx1) and three monothiol (CXXS) Grx-like proteins (GLPs), which have five residue insertions prior to the active-site Cys. Here, the crystal structure of PfGrx1 has been determined by the sulfur single-wavelength anomalous diffraction (S-SAD) method utilizing intrinsic protein and solvent S atoms. Several residues were modelled with alternate conformations, and an alternate position was refined for the active-site Cys29 owing to radiation damage. The GSH-binding site is occupied by water polygons and buffer molecules. Structural comparison of PfGrx1 with other Grxs and Grx-like proteins revealed that the GSH-binding motifs (CXXC/CXXS, TVP, CDD, Lys26 and Gln/Arg63) are structurally conserved. Both the monothiol and dithiol Grxs possess three conserved water molecules; two of these were located in the GSH-binding site. PfGrx1 has several polar and charged amino-acid substitutions that provide structurally important additional hydrogen bonds and salt bridges missing in other Grxs.

Malaria parasite tyrosyl-tRNA synthetase secretion triggers pro-inflammatory responses.

Malaria infection triggers pro-inflammatory responses in humans that are detrimental to host health. Parasite-induced enhancement in cytokine levels correlate with malaria-associated pathologies. Here we show that parasite tyrosyl-tRNA synthetase (PfTyrRS), a housekeeping protein translation enzyme, induces pro-inflammatory responses from host immune cells. PfTyrRS exits from the parasite cytoplasm into the infected red blood cell (iRBC) cytoplasm, from where it is released into the extracellular medium on iRBC lysis. Using its ELR peptide motif, PfTyrRS specifically binds to and internalizes into host macrophages, leading to enhanced secretion of the pro-inflammatory cytokines TNF-α and IL-6. PfTyrRS-macrophage interaction also augments expression of adherence-linked host endothelial receptors ICAM-1 and VCAM-1. Our description of PfTyrRS as a parasite-secreted protein that triggers pro-inflammatory host responses, along with its atomic resolution crystal structure in complex with tyrosyl-adenylate, provides a novel platform for targeting PfTyrRS in anti-parasitic strategies.