CAFs shape myeloid-derived suppressor cells to promote stemness of intrahepatic cholangiocarcinoma through 5-lipoxygenase.

BACKGROUND AND AIMS: We previously demonstrated that cancer-associated fibroblasts (CAFs) promote tumor growth through recruitment of myeloid-derived suppressor cells (MDSCs). 5-lipoxygenase (5-LO) is highly expressed in myeloid cells and is critical for synthesizing leukotriene B4 (LTB4), which is involved in tumor progression by activating its receptor leukotriene B4 receptor type 2 (BLT2). In this study, we investigated whether and how CAFs regulate MDSC function to enhance cancer stemness, the driving force of the cancer aggressiveness and chemotherapy refractoriness, in highly desmoplastic intrahepatic cholangiocarcinoma (ICC). APPROACH AND RESULTS: RNA-sequencing analysis revealed enriched metabolic pathways but decreased inflammatory pathways in cancer MDSCs compared with blood MDSCs from patients with ICC. Co-injection of ICC patient-derived CAFs promoted cancer stemness in an orthotopic ICC model, which was blunted by MDSC depletion. Conditioned media (CM) from CAF-educated MDSCs drastically promoted tumorsphere formation efficiency and stemness marker gene expression in ICC cells. CAF-CM stimulation increased expression and activity of 5-LO in MDSCs, while 5-LO inhibitor impaired the stemness-enhancing capacity of MDSCs in vitro and in vivo. Furthermore, IL-6 and IL-33 primarily expressed by CAFs mediated hyperactivated 5-LO metabolism in MDSCs. We identified the LTB4-BLT2 axis as the critical downstream metabolite signaling of 5-LO in promoting cancer stemness, as treatment with LTB4 was elevated in CAF-educated MDSCs, or blockade of BLT2 (which was preferentially expressed in stem-like ICC cells) significantly reduced stemness-enhancing effects of CAF-educated MDSCs. Finally, BLT2 blockade augmented chemotherapeutic efficacy in ICC patient-derived xenograft models. CONCLUSIONS: Our study reveals a role for CAFs in orchestrating the optimal cancer stemness-enhancing microenvironment by educating MDSCs, and suggests the 5-LO/LTB4-BLT2 axis as promising therapeutic targets for ICC chemoresistance by targeting cancer stemness.

Effect of Lignin Modifier on Engineering Performance of Bituminous Binder and Mixture.

Lignin accounts for approximately 30% of the weight of herbaceous biomass. Utilizing lignin in asphalt pavement industry could enhance the performance of pavement while balancing the construction cost. This study aims to evaluate the feasibility of utilizing lignin as a bitumen performance improver. For this purpose, lignin derived from aspen wood chips (labeled as KL) and corn stalk residues (labeled as CL) were selected to prepare the lignin modified bituminous binder. The properties of the lignin modified binder were investigated through rheological, mechanical and chemical tests. The multiple stress creep recovery (MSCR) test results indicated that adding lignin decreased the Jnr of based binder by a range of 8% to 23% depending on the stress and lignin type. Lignin showed a positive effect on the low temperature performance of asphalt binder, because at -18 °C, KL and CL were able to reduce the stiffness of base binder from 441 MPa to 369 MPa and 378 MPa, respectively. However, lignin was found to deteriorate the fatigue life and workability of base binder up to 30% and 126%. With bituminous mixture, application of lignin modifiers improved the Marshall Stability and moisture resistance of base mixture up to 21% and 13%, respectively. Although, adding lignin modifiers decreased the molecular weight of asphalt binder according to the gel permeation chromatography (GPC) test results. The Fourier-transform infrared spectroscopy (FTIR) test results did not report detectable changes in functional group of based binder.

Fibroblastic FAP promotes intrahepatic cholangiocarcinoma growth via MDSCs recruitment.

Desmoplasia is a hallmark of intrahepatic cholangiocarcinoma (ICC), which constitutes a barrier to infiltration of lymphocyte, but not myeloid cells. Given that dense desmoplastic stroma has been reported to be a barrier to infiltration of lymphocyte, but not myeloid cells. We here investigated whether fibroblastic FAP influenced ICC progression via non-T cell-related immune mechanisms. We demonstrated fibroblastic FAP expression was critical for STAT3 activation and CCL2 production, and ICC-CAFs were the primary source of CCL2 in human ICC microenvironment by using ICC-Fbs from six ICC patients. Fibroblastic knockdown of FAP significantly impaired the ability of ICC-CAFs to promote ICC growth, MDSCs infiltration and angiogenesis, which was restored by adding exogenous CCL2. Furthermore, interestingly, the tumor-promoting effect of fibroblastic FAP is dependent on MDSCs via secretion of CCL2, as depletion of Gr-1+ cells reversed the restoring effects of exogenous CCL2 on tumor growth and angiogenesis. In vitro migration assay confirmed that exogenous CCL2 could rescue the impaired ability of ICC-Fbs to attract Gr-1+ cells caused by fibroblastic FAP knockdown. In contrast, fibroblastic FAP knockdown had no effect on ICC cell proliferation and apoptotic resistance. Depletion MDSCs by anti-Gr-1 monoclonal antibody in subcutaneous transplanted tumor model abrogated tumor promotion by ICC-CAFs suggested that the pro-tumor function of Fibroblastic FAP relied on MDSCs. Mechanical, flow cytometry and chamber migration assay were conducted to find Fibroblastic FAP was required by the ability of ICC-CAFs to promote MDSC migration directly. Moreover, fibroblastic FAP knockdown had no effect on cell proliferation and apoptotic resistance. Here, we revealed the T-cell independent mechanisms underlying the ICC-promoting effect of fibroblastic FAP by attracting MDSCs via CCL2, which was mainly attributed to the ability of FAP to attract MDSCs and suggests that specific targeting fibroblastic FAP may represent a promising therapeutic strategy against ICC.

Inhibition of retinal angiogenesis by gold nanoparticles via inducing autophagy.

AIM: To investigate the effect of gold nanoparticles on retinal angiogenesis in vitro and in vivo, and to reveal the possible mechanism. METHODS: Seed growth method was used to synthesize gold nanoparticles (GNPs). The size, zeta potential, absorption spectrum and morphology of GNPs were identified using Malvern Nano-ZS, multimode reader (BioTek synergy2) and transmission electron microscope. Cell viability was analyzed using cell counting kit-8 method and cell growth was assessed with EdU kit. Transwell chamber was used to investigate cell migration. Tube formation method was used to assess the angiogenic property in vitro. Oxygen induced retinopathy (OIR) model was used to investigate the effect of GNPs on retinal angiogenesis. Confocal microscope and Western blot were used to study the possible mechanism of GNPs inhibited angiogenesis. RESULTS: The GNPs synthesized were uniform and well dispersed. GNPs of 10 µg/mL and 20 µg/mL were able to inhibit human umbilical vein endothelial cells proliferation (50% and 72% separately, P<0.001), migration (54% and 83% separately, P<0.001) and tube formation (52% and 90% separately, P<0.001). Further data showed that GNPs were able to improve the retinopathy in an OIR model. The possible mechanism might be that GNPs were able to induce autophagy significantly (P<0.05). CONCLUSION: The present study suggests that GNPs are able to inhibit retinal neovascularization in vitro and in vivo. GNPs might be a potential nanomedicine for the treatment of retinal angiogenesis.

Biocompatible PEGylated Gold nanorods function As cytokinesis inhibitors to suppress angiogenesis.

Pathological angiogenesis is driven by uncontrolled growth of endothelial cells (ECs), which could lead to retinopathy, tumor and rheumatoid arthritis, etc. ECs must experience multiple cell division process to grow, and cytokinesis is the final step. The present study shows that PEGylated GNRs (PEG-GNRs) specifically target ECs cytokinesis process which results in high ratio of binucleated cells, and these binucleated ECs lose the ability to proliferate. Further data show that PEG-GNRs do not induce toxicity in vitro and in vivo. PEG-GNRs could inhibit ECs proliferation, migration, tube formation and inhibit angiogenesis in ex vivo model. Oxygen induced retinopathy and tumor angiogenesis model further show that PEG-GNRs can inhibit angiogenesis in vivo. Gene expression profiles reveal that PEG-GNRs mainly affect ECs cell division process, and PEG-GNRs treated ECs are arrested in G2/M phase. The mechanism is that PEG-GNRs could disrupt TGFß pathway, and subsequently suppress the assembly of actin filaments in contractile ring site. These findings indicate that PEG-GNR is a novel cytokinesis inhibitor which can be used to interfere with retinal angiogenesis and tumor.