SORL1 stabilizes ABCB1 to promote cisplatin resistance in ovarian cancer.

Ovarian cancer (OC) has the worst prognosis among gynecological malignancies. Cisplatin (CDDP) is one of the most commonly used treatments for OC, but recurrence and metastasis are common due to endogenous or acquired resistance. High expression of ATP-binding cassette (ABC) transporters is an important mechanism of resistance to OC chemotherapy, but targeting ABC transporters in OC therapy remains a challenge. The expression of sortilin-related receptor 1 (SORL1; SorLA) in the response of OC to CDDP was determined by analysis of TCGA and GEO public datasets. Immunohistochemistry and western blotting were utilized to evaluate the expression levels of SORL1 in OC tissues and cells that were sensitive or resistant to CDDP treatment. The in vitro effect of SORL1 on OC cisplatin resistance was proven by CCK-8 and cell apoptosis assays. The subcutaneous xenotransplantation model verified the in vivo significance of SORL1 in OC. Finally, the molecular mechanism by which SORL1 regulates OC cisplatin resistance was revealed by coimmunoprecipitation, gene set enrichment analysis and immunofluorescence analysis. This study demonstrated that SORL1 is closely related to CDDP resistance and predicts a poor prognosis in OC. In vivo xenograft experiments showed that SORL1 knockdown significantly enhanced the effect of CDDP on CDDP-resistant OC cells. Mechanistically, silencing of SORL1 inhibits the early endosomal antigen 1 (EEA1) pathway, which impedes the stability of ATP-binding cassette B subfamily member 1 (ABCB1), sensitizing CDDP-resistant OC cells to CDDP. The findings of this study suggest that targeting SORL1 may represent a promising therapeutic approach for overcoming CDDP resistance in OC.

F-spondin inhibits differentiation of clastic precursors via lipoprotein receptor-related protein 8 (LRP8).

BACKGROUND: F-spondin, known to be a secreted neuronal glycoprotein, is highly expressed on the tooth root surface. The authors previously reported that F-spondin is one of the specific markers of cementoblasts in periodontal tissue. In chronic periodontitis, significant cemental resorption rarely occurs on the root side, although alveolar bone resorption by osteoclasts is one of the major pathologic changes. Thus, it was hypothesized that secretory F-spondin from cementoblasts might be involved in differentiation of clastic cells on the root surface. The authors studied effects of secretory F-spondin from F-spondin-expressing cells and its pathway on receptor activator of nuclear factor-κB ligand (RANKL)-mediated differentiation of clastic cells. METHODS: Osteoclast precursors were used in this study. With a chamber assay, the authors examined effects of secretory molecules from F-spondin-expressing cells of transgenic mice on RANKL-induced clastic cell differentiation. RESULTS: Secretory molecules from F-spondin-overexpressing cells significantly inhibited the RANKL-mediated tartrate-resistant acid phosphatase (TRAP)-positive cells from primary progenitor cells with the chamber system. F-spondin suppressed RANKL-mediated nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1); TRAP; cathepsin K; and dendritic cell-specific transmembrane protein (DC-STAMP) expression in the cells. The suppressive effect of F-spondin on RANKL-induced differentiation of clastic cells was partially blocked by knockdown of low-density lipoprotein receptor-related protein 8 (LRP8). CONCLUSIONS: These findings indicate that secretory factors from F-spondin-expressing cells, including F-spondin, downregulate differentiation of clastic precursors. Moreover, F-spondin inhibits RANKL-mediated differentiation of clastic cells partially via LRP8. It is suggested that secretory F-spondin may act protectively from cemental resorption partially via LRP8 in periodontal tissue.

Direct inhibition of GSK3beta by the phosphorylated cytoplasmic domain of LRP6 in Wnt/beta-catenin signaling.

Wnt/beta-catenin signaling plays a central role in development and is also involved in a diverse array of diseases. Binding of Wnts to the coreceptors Frizzled and LRP6/5 leads to phosphorylation of PPPSPxS motifs in the LRP6/5 intracellular region and the inhibition of GSK3beta bound to the scaffold protein Axin. However, it remains unknown how GSK3beta is specifically inhibited upon Wnt stimulation. Here, we show that overexpression of the intracellular region of LRP6 containing a Ser/Thr rich cluster and a PPPSPxS motif impairs the activity of GSK3beta in cells. Synthetic peptides containing the PPPSPxS motif strongly inhibit GSK3beta in vitro only when they are phosphorylated. Microinjection of these peptides into Xenopus embryos confirms that the phosphorylated PPPSPxS motif potentiates Wnt-induced second body axis formation. In addition, we show that the Ser/Thr rich cluster of LRP6 plays an important role in LRP6 binding to GSK3beta. These observations demonstrate that phosphorylated LRP6/5 both recruits and directly inhibits GSK3beta using two distinct portions of its cytoplasmic sequence, and suggest a novel mechanism of activation in this signaling pathway.

N-methyl-D-aspartate receptor inhibition by an apolipoprotein E-derived peptide relies on low-density lipoprotein receptor-associated protein.

The effects of a synthetic apoE peptide, viz., residues 133-149 (apoE[133-149]), a mimetic that comprises the apoE receptor binding domain, on N-methyl-D-aspartate (NMDA)/glycine-induced ion flow through NMDA receptor (NMDAR) channels, have been investigated. The activity of apoE[133-149] was found to depend on the low-density lipoprotein receptor-related protein (LRP). Competition experiments with receptor-associated protein (RAP) and activated alpha(2)-macroglobulin (alpha(2)M*), two proteins that compete for apoE binding to LRP, demonstrate that apoE[133-149] inhibition of NMDAR function is mediated at a locus in LRP that overlaps with the binding sites of RAP and alpha(2)M*. A coreceptor of LRP, cell surface heparin sulfate proteoglycan, did not function in this system. Additional electrophysiology experiments demonstrated that the inhibitory potency of apoE[133-149] was threefold greater for NMDAR-transfected wild-type Chinese hamster ovary (CHO) cells compared with NMDAR-transfected CHO cells deficient in LRP. Studies with truncation and replacement variants of the apoE peptide demonstrated that the NMDAR inhibitory properties of these peptides correlate with their binding affinities for LRP. These novel results indicate that apoE functions as an inhibitor of NMDAR ion channels indirectly via LRP, and are suggestive of a participatory role for LRP in NMDAR-based neuropathies.

The Wnt antagonist Dickkopf-1 and its receptors are coordinately regulated during early human adipogenesis.

Secretion of Wnts by adipose cells has an important role in the control of murine adipogenesis. We present the first evidence that a Wnt antagonist, Dickkopf 1 (Dkk1), is secreted by human preadipocytes and promotes adipogenesis. DKK1 mRNA increases six hours after onset of human adipogenesis and this is followed by an increase in Dkk1 protein. With further differentiation, the mRNA and protein levels progressively decline such that they are undetectable in mature adipocytes. The transient induction in DKK1 correlates with downregulation of cytoplasmic and nuclear beta-catenin levels, this being a surrogate marker of canonical Wnt signalling, and Wnt/beta-catenin transcriptional activity. In addition, constitutive expression of Dkk1 in 3T3-L1 preadipocytes promotes their differentiation, further supporting the functional significance of increased Dkk1 levels during human adipogenesis. Concomitant downregulation of the Dkk1 receptors LRP5 and LRP6 is likely to potentiate the ability of Dkk1 to inhibit Wnt signalling and promote differentiation. Notably, Dkk1 is not expressed in primary murine preadipocytes or cell lines. The involvement of Dkk1 in human but not murine adipogenesis indicates that inter-species differences exist in the molecular control of this process. Given the public health importance of disorders of adipose mass, further knowledge of the pathways involved specifically in human adipocyte differentiation might ultimately be of clinical relevance.

Human triglyceride-rich lipoproteins impair glucose metabolism and insulin signalling in L6 skeletal muscle cells independently of non-esterified fatty acid levels.

AIMS/HYPOTHESIS: Elevated fasting and postprandial plasma levels of triglyceride-rich lipoproteins (TGRLs), i.e. VLDL/remnants and chylomicrons/remnants, are a characteristic feature of insulin resistance and are considered a consequence of this state. The aim of this study was to investigate whether intact TGRL particles are capable of inducing insulin resistance. METHODS: We studied the effect of highly purified TGRLs on glycogen synthesis, glycogen synthase activity, glucose uptake, insulin signalling and intramyocellular lipid (IMCL) content using fully differentiated L6 skeletal muscle cells. RESULTS: Incubation with TGRLs diminished insulin-stimulated glycogen synthesis, glycogen synthase activity, glucose uptake and insulin-stimulated phosphorylation of Akt and glycogen synthase kinase 3. Insulin-stimulated tyrosine phosphorylation of IRS-1, and IRS-1- and IRS-2-associated phosphatidylinositol 3-kinase (PI3K) activity were not impaired by TGRLs, suggesting that these steps were not involved in the lipoprotein-induced effects on glucose metabolism. The overall observed effects were time- and dose-dependent and paralleled IMCL accumulation. NEFA concentration in the incubation media did not increase in the presence of TGRLs indicating that the effects observed were solely due to intact lipoprotein particles. Moreover, co-incubation of TGRLs with orlistat, a potent active-site inhibitor of various lipases, did not alter TGRL-induced effects, whereas co-incubation with receptor-associated protein (RAP), which inhibits interaction of TGRL particles with members of the LDL receptor family, reversed the TGRL-induced effects on glycogen synthesis and insulin signalling. CONCLUSIONS/INTERPRETATION: Our data suggest that the accumulation of TGRLs in the blood stream of insulin-resistant patients may not only be a consequence of insulin resistance but could also be a cause for it.