Improving the immunomodulatory efficacy of mesenchymal stem cells (MSCs) through pretreatment with pro-inflammatory cytokines is an evolving field of investigation. However, the underlying mechanisms have not been fully clarified. Here, we pretreated human umbilical cord-derived MSCs with interleukin-1β (IL-1β) and evaluated their therapeutic effects in a cecal ligation and puncture-induced sepsis model. We found that systemic administration of IL-1β-pretreated MSCs (βMSCs) ameliorated the symptoms of murine sepsis more effectively and increased the survival rate compared with naïve MSCs. Furthermore, βMSCs could more effectively induce macrophage polarization toward an anti-inflammatory M2 phenotype through the paracrine activity. Mechanistically, we demonstrated that βMSC-derived exosomes contributed to the enhanced immunomodulatory properties of βMSCs both in vitro and in vivo. Importantly, we found that miR-146a, a well-known anti-inflammatory microRNA, was strongly upregulated by IL-1β stimulation and selectively packaged into exosomes. This exosomal miR-146a was transferred to macrophages, resulted in M2 polarization, and finally led to increased survival in septic mice. In contrast, inhibition of miR-146a through transfection with miR-146a inhibitors partially negated the immunomodulatory properties of βMSC-derived exosomes. Taken together, IL-1β pretreatment effectively enhanced the immunomodulatory properties of MSCs partially through exosome-mediated transfer of miR-146a. Therefore, we believe that IL-1β pretreatment may provide a new modality for better therapeutic application of MSCs in inflammatory disorders. Stem Cells 2017;35:1208-1221.

Stromal carcinoma-related fibroblasts (CAFs) are the main type of non-immune cells in the tumor microenvironment (TME). CAFs interact with cancer cells to promote tumor proliferation. Long non-coding RNAs (lncRNAs) are known to regulate cell growth, apoptosis and metastasis of cancer cells, but their role in stromal cells is unclear. Using RNA sequencing, we identified a stromal lncRNA signature during the transformation of CAFs from normal fibroblasts (NFs) in oral squamous cell carcinoma (OSCC). We uncovered an uncharacterized lncRNA, FLJ22447, which was remarkably up-regulated in CAFs, referred to LncRNA-CAF (Lnc-CAF) hereafter. Interleukin-33 (IL-33) was mainly located in the stroma and positively co-expressed with Lnc-CAF to elevate the expression of CAF markers (α-SMA, vimentin and N-cadherin) in fibroblasts. In a co-culture system, IL-33 knockdown impaired Lnc-CAF-mediated stromal fibroblast activation, leading to decreased proliferation of tumor cells. Mechanistically, Lnc-CAF up-regulated IL-33 levels and prevented p62-dependent autophagy-lysosome degradation of IL-33, which was independent of LncRNA-protein scaffold effects. Treatment with the autophagy inducer, rapamycin, impaired the proliferative effect of Lnc-CAF/IL-33 by promoting IL-33 degradation. In turn, tumor cells further increased Lnc-CAF levels in stromal fibroblasts via exosomal Lnc-CAF. In patients with OSCC, high Lnc-CAF/IL-33 expression correlated with high TNM stage (n = 140). Moreover, high Lnc-CAF expression predicted poor prognosis. In vivo, Lnc-CAF knockdown restricted tumor growth and was associated with decreased Ki-67 expression and α-SMA+ CAF in the stroma. In conclusion, we identified a stromal lncRNA signature, which reprograms NFs to CAFs via Lnc-CAF/IL-33 and promotes OSCC development.

Saliva is a noninvasive biofluid that can contain metabolite signatures of oral squamous cell carcinoma (OSCC). Conductive polymer spray ionization mass spectrometry (CPSI-MS) is employed to record a wide range of metabolite species within a few seconds, making this technique appealing as a point-of-care method for the early detection of OSCC. Saliva samples from 373 volunteers, 124 who are healthy, 124 who have premalignant lesions, and 125 who are OSCC patients, were collected for discovering and validating dysregulated metabolites and determining altered metabolic pathways. Metabolite markers were reconfirmed at the primary tissue level by desorption electrospray ionization MS imaging (DESI-MSI), demonstrating the reliability of diagnoses based on saliva metabolomics. With the aid of machine learning (ML), OSCC and premalignant lesions can be distinguished from the normal physical condition in real time with an accuracy of 86.7%, on a person by person basis. These results suggest that the combination of CPSI-MS and ML is a feasible tool for accurate, automated diagnosis of OSCC in clinical practice.

CD68 has been widely used as a pan-macrophage marker for tumor-associated macrophages (TAM) which always involve in carcinogenesis. But the correlations between CD68(+) TAMs and prognosis of patients show to be inconsistent, which might due to lack of specific markers of TAMs. We here found that the microlocalization of CD68(+) TAMs also played a unique role in prognosis of patients with oral squamous cell carcinoma (OSCC). CD68(+) TAMs were identified in paraffin-embedded OSCC specimens (n = 91) by using immunohistochemistry. The number of CD68(+) TAMs was remarkably increased from adjacent none-neoplasia tissues (NT) to tumor nest (TN), but tumor stroma (TS) was infiltrated with highest frequency of CD68(+) TAMs (P < 0.0001). Unexpectedly, more CD68(+) TAMs in TS, but not NT or TN, were associated with high tumor grade (P = 0.033), lymph node metastasis (P = 0.034), and shorter 10-year overall survival time, disease free survival. Considering TAMs was derived from monocytes in peripheral blood, we assessed the relationship between leukocytes in peripheral blood and CD68(+) TAMs in OSCC and found that more CD68(+) TAMs in TS were accompanied with decreased monocytes and lymphocytes in peripheral blood (P < 0.05). Although Cox regression analysis revealed that CD68(+) TAMs in TS were not an independent prognostic factor for OSCC patients, we raised a possibility that the microlocalization of CD68(+) TAMs was an indispensable factor for the advance of OSCC.