Haowen Zhou

The precise control of messenger RNA (mRNA) translation is a crucial step in posttranscriptional gene regulation of cellular physiology. However, it remains a challenge to systematically study mRNA translation at the transcriptomic scale with spatial and single-cell resolution. Here, we report the development of ribosome-bound mRNA mapping (RIBOmap), a highly multiplexed three-dimensional in situ profiling method to detect cellular translatome. RIBOmap profiling of 981 genes in HeLa cells revealed cell cycle-dependent translational control and colocalized translation of functional gene modules. We mapped 5413 genes in mouse brain tissues, yielding spatially resolved single-cell translatomic profiles for 119,173 cells and revealing cell type-specific and brain region-specific translational regulation, including translation remodeling during oligodendrocyte maturation. Our method detected widespread patterns of localized translation in neuronal and glial cells in intact brain tissue networks.

BACKGROUND: Evidence has demonstrated conditioned medium (CM) from periodontal ligament stem cells (PDLSCs) improved periodontal regeneration. Gingival mesenchymal stem cells (GMSCs) have been considered an alternative strategy for regenerative medicine. To determine whether GMSC-CM could promote periodontal wound healing, we compared the effects of GMSC-CM and PDLSC-CM on periodontal regeneration and the underlying mechanisms in rat periodontal defects. METHODS: Cell-free CMs were collected from PDLSCs, GMSCs, and gingival fibroblasts (GFs) using ultracentrifugation (100-fold concentration). Periodontal defects were created on the buccal side of the first molar in the left mandible of 90 rats by a surgical method. Collagen membranes loaded with concentrated CMs (α-MEM, GF-CM, GMSC-CM, PDLSC-CM) were transplanted into periodontal defects. After 1, 2, and 4 weeks, the animals were sacrificed and specimens including the first molar and the surrounding tissues were separated and decalcified. Hematoxylin-eosin and Masson's trichrome staining were performed to evaluate periodontal regeneration. Immunohistochemical staining for tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-10 was conducted to analyze inflammation. Immunohistochemistry of BSP-II and Runx2 was performed to analyze osteoblast differentiation. RESULTS: Histological analysis showed the amount of newly formed periodontal tissue was significantly higher in both the GMSC-CM and PDLSC-CM groups than in the other groups, with no significant difference between these two groups. At 1 and 2 weeks, the expression levels of TNF-α and IL-1β were significantly lower in the GMSC-CM and PDLSC-CM groups than in the other three groups, while there was no significant difference between these two groups. IL-10 expression was significantly higher in the GMSC-CM group than in the PDLSC-CM group and the other three groups. At 1, 2, and 4 weeks, BSP-II and Runx2 expressions were significantly higher in the GMSC-CM and PDLSC-CM groups than in the other three groups, with no significant difference between the two groups. CONCLUSIONS: Our results demonstrate that GMSC-CM transplantation can significantly promote periodontal regeneration in rats and achieve the same effect as PDLSC-CM. The mechanism of periodontal regeneration may involve the regulation of inflammatory factors and the promotion of osteogenic differentiation of bone progenitor cells in the wound region by CMs from MSCs.

BACKGROUND: Aberrant activation of Wnt/beta-catenin signaling promotes the development of several cancers. It has been demonstrated that the Wnt signaling pathway is activated in chronic lymphocytic leukemia (CLL) cells, and that uncontrolled Wnt/beta-catenin signaling may contribute to the defect in apoptosis that characterizes this malignancy. Thus, the Wnt signaling pathway is an attractive candidate for developing targeted therapies for CLL. METHODOLOGY/PRINCIPAL FINDINGS: The diuretic agent ethacrynic acid (EA) was identified as a Wnt inhibitor using a cell-based Wnt reporter assay. In vitro assays further confirmed the inhibitory effect of EA on Wnt/beta-catenin signaling. Cell viability assays showed that EA selectively induced cell death in primary CLL cells. Exposure of CLL cells to EA decreased the expression of Wnt/beta-catenin target genes, including LEF-1, cyclin D1 and fibronectin. Immune co-precipitation experiments demonstrated that EA could directly bind to LEF-1 protein and destabilize the LEF-1/beta-catenin complex. N-acetyl-L-cysteine (NAC), which can react with the alpha, beta-unsaturated ketone in EA, but not other anti-oxidants, prevented the drug's inhibition of Wnt/beta-catenin activation and its ability to induce apoptosis in CLL cells. CONCLUSIONS/SIGNIFICANCE: Our studies indicate that EA selectively suppresses CLL survival due to inhibition of Wnt/beta-catenin signaling. Antagonizing Wnt signaling in CLL with EA or related drugs may represent an effective treatment of this disease.