Background. Endometrial regenerative cells (ERCs) have been identified to ameliorate colitis in mice; however, whether exosomes derived from ERCs (ERC-exos) own similar effects on colitis remains unclear. Ferroptosis, an iron-dependent cell programmed death form, has been reported to promote inflammation in UC. Thus, in this study, whether ERC-exos can treat colitis and regulate intestine ferroptosis will be explored. Methods. In this study, iron, malondialdehyde (MDA) production, glutathione (GSH) synthesis, and acyl-CoA synthetase long-chain family member (ACSL) 4 and glutathione peroxidase 4 (GPX4) expressions were measured in colon samples from healthy people and UC patients to explore the effects of ferroptosis. In vitro, ERC-exos were cocultured with ferroptosis inducer erastin-treated NCM460 human intestinal epithelial cell line, and ferroptotic parameters were measured. In vivo, colitis was induced by 3% dextran sulfate sodium (DSS) in BALB/c mice, and animals were randomly assigned to normal, untreated, and ERC-exos-treated groups. The Disease Activity Index (DAI) score, histological features, tissue iron, MDA, GSH, ACSL4, and GPX4 were measured to verify the role of ERC-exos in attenuating UC. Results. Compared with healthy people, UC samples exhibited higher levels of iron, MDA, and ACSL4, while less levels of GSH and GPX4. In vitro, the CCK-8 assay showed that ERC-exos rescued erastin-induced cell death, and ERC-exos treatment significantly increased the levels of GSH and expression of GPX4, while markedly decreasing the levels of iron, MDA, and expression of ACSL4. In vivo, ERC-exos treatment effectively reduced DAI score, ameliorated colon pathological damage, and improved disease symptoms. Moreover, ERC-exos treatment further enhanced the levels of GSH and the expression of GPX4 but reduced the levels of iron, MDA, and expression of ACSL4 in the colon of colitis mice. Conclusions. Ferroptosis was involved in the pathogenesis of UC, and ERC-exos attenuated DSS-induced colitis through downregulating intestine ferroptosis. This study may provide a novel insight into treating UC in the future.

Background. Traditional interventions can play a certain role in attenuating ulcerative colitis (UC), known as one type of inflammatory bowel diseases, but sometimes are not effective. Endometrial regenerative cells (ERCs) have been shown to exert immunosuppressive effects in different models of inflammation, and stem cell-derived conditioned media (CM) have advantages over cell therapy in terms of easy access and direct action. However, whether ERC-CM could alleviate colitis remains unclear and will be explored in this study. Methods. Menstrual blood was collected from healthy female volunteers to obtain ERCs and ERC-CM. Acute colitis was induced by 3% dextran sodium sulfate (DSS), and ERC-CM was injected on days 4, 6, and 8, respectively, after induction. The disease activity index was calculated through the record of weight change, bleeding, and fecal viscosity during the treatment process. Histological features, macrophage and CD4+ T cell in the spleen and colon, and cytokine profiles in the sera and colon were measured. In addition, an in vitro lymphocyte proliferation assay was measured by using a CCK-8 kit in this study. Results. ERC-CM treatment significantly improved the symptoms and histological changes in colitis mice. ERC-CM increased the percentage of Tregs in the spleen and colon but decreased the percentages of M1 macrophages and Th1 and Th17 cells in the spleen and decreased the population of Th17 cells in the colon. In addition, ERC-CM treatment decreased the local expression of TNF-α, IL-6, and iNOS in the colon. Furthermore, ERC-CM increased the levels of anti-inflammatory cytokines IL-10 and IL-27 but decreased proinflammatory cytokines IL-6 and IL-17 in the sera. In addition, ERC-CM significantly inhibited ConA-induced mouse lymphocyte proliferation in vitro. Conclusion. The results suggest that ERC-CM can exert similar therapeutic effects as ERCs and could be explored for future application of cell-free therapy in the treatment of colitis.

Background We have demonstrated that endometrial regenerative cells (ERCs) are mesenchymal-like stromal cells and can attenuate experimental colitis, however, its underlying mechanism needs further investigation. Dickkopf-1 (DKK1), a glucoprotein secreted by mesenchymal stromal cells (MSCs), is a classical inhibitor of Wnt/β-catenin pathway which is closely associated with the development of colitis. Therefore, the objective of this study was to investigate whether ERCs could also secret DKK1, and whether the downregulation of DKK1 (DKK1low-ERCs) would enhance the therapeutic effects of ERCs in attenuation of experimental colitis. Methods BALB/c mice were given 3% dextran sodium sulfate (DSS) for 7 consecutive days and free tap water for 3 days sequentially to induce experimental colitis. Unmodified ERCs, IL-1β-treated ERCs (DKK1low-ERCs) and glucocorticoid-treated ERCs (DKK1high-ERCs) were injected (1 million/mouse/day, i.v.) on day 2, 5 and 8 respectively. Colonic and splenic samples were harvested on day 10 after DSS-induction. Results It was found that DKK1low-ERC treatment markedly attenuated colonic damage, body weight loss and colon-length shortening in colitis mice. Compared with other treatments, cell populations of CD4+IL-4+Th2, CD4+CD25+FOXP3+Treg, and CD68+CD206+macrophages in spleens were also significantly upregulated in DKK1low-ERC group (p < 0.05). In addition, lower expression of pro-inflammatory (TNF-α and IFN-γ), but higher levels of anti-inflammatory cytokines (IL-4 and IL-10) and β-catenin were detected in colons in DKK1low-ERC group (p < 0.01 vs. other groups). Conclusions DKK1low-ERCs display augmented immunoregulatory ability and therapeutic effects in DSS-induced colitis.

Background : Ischemia reperfusion injury (IRI) is the major cause of intestinal damage in clinic. Although either mesenchymal stromal cells (MSCs) or interleukin 37 (IL-37) showed some beneficial roles to ameliorate IRI, their effects are limited. In this study, the protective effects of IL-37 gene-modified MSCs (IL-37-MSCs) for better prevention of intestinal IRI are investigated. Methods: Intestinal IRI model was established by occluding the superior mesenteric artery for 30min and then reperfusing for 72 hours in rats. Forty adult male SD rats were randomly divided into sham control, IL-37-MSC-treated, MSC-treated, recombinant IL-37 (rIL-37)-treated and untreated groups. Intestinal damage was assessed by H&E staining. The levels of gut barrier function factors (diamine oxidase and D-Lactate) and inflammation reactivity cytokine IL-1β were assayed by ELISA. The expressions of tissue damage-related NLRP3 inflammasome and relative proteins including clevead caspase-1, IL-1β and IL-18 were detected by western blot. As downstream of IL-1β and IL-18, the mRNA levels of proinflammatory mediators IL-6 and TNF-α were determined by qPCR. Data were analyzed by one-way analysis of variance among groups. Results : IL-37-MSCs were able to migrate to the damaged tissue and significantly inhibit intestinal IRI. As compared with MSCs or rIL-37 monotherapy group, IL-37-MSC treatment not only improved gut barrier function but also decreased local and systemic inflammation reactivity cytokine IL-1β level in IRI rats. In addition, tissue damage-related NLRP3 and relative proteins (cleaved caspase-1, IL-1β and IL-18) were significantly decreased in IRI rats treated with IL-37-MSCs. Furthermore, IL-1β and IL-18 related proinflammatory mediators IL-6 and TNF-α mRNA expressions were markedly decreased following IL-37-MSC treatment. Conclusion : The results suggest that IL-37 gene modification significantly enhance the protective effects of MSCs against intestinal IRI. In addition, NLRP3-related signaling pathways could be associated with IL-37-MSC mediated protection.

Objectives This study aimed to discover the ceRNAs network in pathophysiological development of human colorectal cancer (CRC) and to screen biomarkers for target therapy and prognosis by using integrated bioinformatics analysis.Methods Data on gene expressions of mRNAs, circRNAs, and miRNAs and clinical information were downloaded from The Cancer Genome Atlas and Gene Expression Omnibus database respectively. Differentially expressed mRNAs (DEGs) were identified by using DESeq2 package of R software. Functional enrichment analysis was conducted by ClusterProfiler package of R software. Protein-protein interaction (PPI) network was shown by STRING website. Survival analysis of hub genes was performed by survival package in R software. Interactions among hub gene, DEmiRNAs and DEcircRNAs were used to construct ceRNAs network.Results A total of 412 DEGs (including 82 upregulated and 330 downregulated) were screened out between 473 CRC and 41 normal samples. 260 DEcircRNAs (including 253 upregulated and 7 downregulated) were altered. A ceRNAs and PPI network was successfully constructed and TIMP1 associated with prognosis were employed.Conclusion The present study identifies a novel ceRNAs network, which imply that TIMP1 is a potential biomarker underlying the development of CRC, providing new insights for survival prediction and therapeutic target.

Background Endometrial regenerative cells (ERCs) play an important role in attenuation of acute allograft rejection, while their effects are limited. IL-37, a newly discovered immunoregulatory cytokine of the IL-1 family, can regulate both innate and adaptive immunity. Whether IL-37 overexpression can enhance the therapeutic effects of ERCs in inhibition of acute cardiac allograft rejection remains unknown and will be explored in this study. Methods C57BL/6 mice recipients receiving BALB/c mouse heterotopic heart allografts were randomly divided into the phosphate-buffered saline (untreated), ERC treated, negative lentiviral control ERC (NC-ERC) treated, and IL-37 overexpressing ERC (IL-37-ERC) treated groups. Graft pathological changes were assessed by H&E staining. The intra-graft cell infiltration and splenic immune cell populations were analyzed by immunohistochemistry and flow cytometry, respectively. The stimulatory property of recipient DCs was tested by an MLR assay. Furthermore, serum cytokine profiles of recipients were measured by ELISA assay. Results Mice treated with IL-37-ERCs achieved significantly prolonged allograft survival compared with the ERC-treated group. Compared with all the other control groups, IL-37-ERC-treated group showed mitigated inflammatory response, a significant increase in tolerogenic dendritic cells (Tol-DCs), regulatory T cells (Tregs) in the grafts and spleens, while a reduction of Th1 and Th17 cell population. Additionally, there was a significant upregulation of immunoregulatory IL-10, while a reduction of IFN-γ, IL-17A, IL-12 was detected in the sera of IL-37-ERC-treated recipients. Conclusion IL-37 overexpression can promote the therapeutic effects of ERCs to inhibit acute allograft rejection and further prolong graft survival. This study suggests that gene-modified ERCs overexpressing IL-37 may pave the way for novel therapeutic options in the field of transplantation.

Hypopharyngeal cancer is a subtype of the head and neck malignancies. We aimed to explore the role of lysine-specific demethylase 1 (LSD1/KDM1A) in the progression of hypopharyngeal cancer and to identify the potential mechanisms. First, LSD1 expression in head and neck squamous cell carcinoma (HNSCC) tissues and the correlation between LSD1 and the stage of HNSC were analyzed by the University of ALabama at Birmingham CANcer data analysis Portal (UALCAN). Following LSD1 silencing, proliferation of pharyngeal cancer cell line FaDu cells was evaluated by cell counting kit-8 and colony formation assays. Wounding healing and transwell assays were used to measure the capacities of migration and invasion. In addition, expression of proteins related to epithelial-to-mesenchymal transition (EMT), autophagy, and pyroptosis was tested by Western blot analysis or immunofluorescence. After treatment with autophagy inhibitor 3-methyladenine (3-MA) or NLR family pyrin domain containing 3 (NLRP3) inhibitor MCC950, the malignant biological properties were measured again. High LSD1 expression was observed in HNSC tissues, which was correlated with stage. LSD1 knockdown significantly suppressed the proliferation, migration, invasion, and EMT of hypopharyngeal cancer cells. Moreover, autophagy and pyroptosis were induced by LSD1 depletion, observed by the enhanced fluorescence intensity of LC3, gasdermin-D (GSDMD)-N, and apoptosis-associated speck-like protein containing a CARD (ASC), accompanied by upregulated expression of LC3II/LC3I, Beclin-1, NLRP3, cleaved-caspase 1, ASC, interleukin (IL)-1β, and IL-18 and downregulated expression of p62. Importantly, 3-MA or MCC950 addition obviously reversed the inhibitory effects of LSD1 silencing on the proliferation, migration, invasion, and EMT of hypopharyngeal cancer cells. To sum up, LSD1 silencing could restrain the progression of hypopharyngeal cancer cells by inducing autophagy and pyroptosis.

Background: To investigate whether RAS-selective lethal 3 (RSL3) combined with cisplatin can induce ferroptosis in cisplatin (DDP) resistant nasopharyngeal carcinoma (NPC) cells. Methods: The sensitivity of nasopharyngeal carcinoma cell lines HNE-1, CEN2Z, HONE-1 and cisplatin-resistant strain HNE-1/DDP to DDP and RAS was detected by CCK-8 method. After combined treatment of RSL3 and DDP, CCK-8 method was used. Changes of HNE-1/DDP cell activity, cell death and cellular reactive oxygen species (ROS) levels were detected by flow cytometry, cellular Fe2+ and MDA levels were detected by Fe2+ and malondialdehyde (MDA) kits, proteins linked to ferroptosis. Results: The sensitivity of HNE-1/DDP to DDP was much lower than that of normal HNE-1 cell line, and the IC50 of RAS was as high as (45.89±6.89) μmol/L. Flow cytometry results showed that when HNE-1/DDP was treated with RSL3 or DDP alone, the mortality rate did not exceed 30%, and the increase in ROS level did not exceed 15%. RSL3 combined with DDP could increase the cell death rate of HNE-1/DDP to 89.69% ± 9.48% and the ROS level to 18.72% ± 3.05%. Combined treatment raised intracellular Fe2+ and MDA levels, reduced the expression of the ferroptosis-related protein glutathione peroxidase 4 (GPX4), and reversed RSL3 combination by the ferroptosis inhibitor Ferrostatin-1. DDP-induced death of cell inhibited the increase of ROS, Fe2+ and MDA levels. Conclusions: RSL3 combined with DDP can induce ferroptosis in cisplatin-resistant strain HNE-1/DDP.