Manganese overexposure results in ferroptosis through the HIF-1α/p53/SLC7A11 pathway in ICR mouse brain and PC12 cells.

Manganese (Mn) overexposure has been associated with the development of neurological damage reminiscent of Parkinson's disease, while the underlying mechanisms have yet to be fully characterized. This study aimed to investigate the mechanisms leading to injury in dopaminergic neurons induced by Mn and identify novel treatment approaches. In the in vivo and in vitro models, ICR mice and dopaminergic neuron-like PC12 cells were exposed to Mn, respectively. We treated them with anti-ferroptotic agents ferrostatin-1 (Fer-1), deferoxamine (DFO), HIF-1α activator dimethyloxalylglycine (DMOG) and inhibitor LW6. We also used p53-siRNA to verify the mechanism underlying Mn-induced neurotoxicity. Fe and Mn concentrations increased in ICR mice brains overexposed to Mn. Additionally, Mn-exposed mice exhibited movement impairment and encephalic pathological changes, with decreased HIF-1α, SLC7A11, and GPX4 proteins and increased p53 protein levels. Fer-1 exhibited protective effects against Mn-induced both behavioral and biochemical changes. Consistently, in vitro, Mn exposure caused ferroptosis-related changes and decreased HIF-1α levels, all ameliorated by Fer-1. Upregulation of HIF-1α by DMOG alleviated the Mn-associated ferroptosis, while LW6 exacerbated Mn-induced neurotoxicity through downregulating HIF-1α. p53 knock-down also rescued Mn-induced ferroptosis without altering HIF-1α protein expression. Mn overexposure resulted in ferroptosis in dopaminergic neurons, mediated through the HIF-1α/p53/SLC7A11 pathway.

T-cell-specific Sel1L deletion exacerbates EAE by promoting Th1/Th17-cell differentiation.

Multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE) are demyelinating neuroinflammatory diseases identified by the accumulation and aggregation of misfolded proteins in the brain. The Sel1L-Hrd1 complex comprising endoplasmic reticulum associated degradation (ERAD) is an ER-protein quality control system (ERQC) in the cell. Unfortunately, the contribution of ERAD to the development of these diseases has not been well explored. In this study, we used mice with a conditional deletion (KO) of Sel1L in T cells to dissect the role of ERAD on T cells and its contribution to the development of EAE. The results showed that Sel1L KO mice developed more severe EAE than the control wild type (WT) mice. Although, no obvious effects on peripheral T cells in steady state, more CD44-CD25+ double-negative stage 3 (DN3) cells were detected in the thymus. Moreover, Sel1L deficiency promoted the differentiation of Th1 and Th17 cells and upregulated the proliferation and apoptosis of CD4 T cells in vitro. Regarding the mechanism analyzed by RNA sequencing, 437 downregulated genes and 271 upregulated genes were detected in Sel1L deletion CD4 T cells, which covered the activation, proliferation, differentiation and apoptosis of these T cells. Thus, this study declared that the dysfunction of Sel1L in ERAD in T cells exacerbated the severity of EAE and indicated the important role of ERQC in maintaining immune homeostasis in the central nervous system.

Regulation of thymic T regulatory cell differentiation by TECs in health and disease.

The thymus produces self-limiting and self-tolerant T cells through the interaction between thymocytes and thymus epithelial cells (TECs), thereby generating central immune tolerance. The TECs are composed of cortical and medullary thymic epithelial cells, which regulate the positive and negative selection of T cells, respectively. During the process of negative selection, thymocytes with self-reactive ability are deleted or differentiated into regulatory T cells (Tregs). Tregs are a subset of suppressor T cells that are important for maintaining immune homeostasis. The differentiation and development of Tregs depend on the development of TECs and other underlying molecular mechanisms. Tregs regulated by thymic epithelial cells are closely related to human health and are significant in autoimmune diseases, thymoma and pregnancy. In this review, we summarize the current molecular and transcriptional regulatory mechanisms by which TECs affect the development and function of thymic Tregs. We also review the pathophysiological models of thymic epithelial cells regulating thymic Tregs in human diseases and specific physiological conditions.

Endoplasmic Reticulum Quality Control in Immune Cells.

The endoplasmic reticulum quality control (ERQC) system, including endoplasmic reticulum-associated degradation (ERAD), the unfolded protein response (UPR), and autophagy, presides over cellular protein secretion and maintains proteostasis in mammalian cells. As part of the immune system, a variety of proteins are synthesized and assembled correctly for the development, activation, and differentiation of immune cells, such as dendritic cells (DCs), macrophages, myeloid-derived-suppressor cells (MDSCs), B lymphocytes, T lymphocytes, and natural killer (NK) cells. In this review, we emphasize the role of the ERQC in these immune cells, and also discuss how the imbalance of ER homeostasis affects the immune response, thereby suggesting new therapeutic targets for immunotherapy.

Effect of idebenone on bone marrow mesenchymal stem cells in vitro.

In recent years, stem cell research has continued to benefit from its crossover with chemistry, particularly the investigation of small molecular drugs modulating specific targets to regulate stem cell fate. Idebenone (IDB) is a yellow crystalline powder that is used in the treatment of chronic cerebrovascular diseases. The objective of the present study was to examine whether IDB had an influence on bone marrow­derived mesenchymal stem cells (BMSCs) extracted from the bone marrow of Sprague­Dawley rats. The effects of IDB on cell proliferation, cell cloning and migration were investigated. Cell cycle, apoptosis, DAPI nuclear staining and senescence­associated ß­galactosidase (SA­ß­gal) staining were also examined. The results revealed that IDB at suitable concentrations enhanced cell cloning capacity, promoted the proliferation of BMSCs, delayed cellular senescence, and inhibited cell apoptosis and migration. Western blot analysis indicated that IDB increased the expression of B­cell lymphoma 2 (Bcl­2), signal transducer and activator of transcription­3, Nanog, octamer­binding transcription factor 4, E­cadherin, proliferating cell nuclear antigen, cyclinD1 and cyclinD3, and decreased the expression of Bcl­2­associated X protein, cleaved caspase­3, N­cadherin, vimentin and α­smooth muscle actin. In conclusion, these experiments confirmed that IDB in low doses had no toxic effect and may exert protective effects on BMSCs.

Nicotine-enhanced stemness and epithelial-mesenchymal transition of human umbilical cord mesenchymal stem cells promote tumor formation and growth in nude mice.

Cigarette smoking is a well-known risk factor in the development and progression of malignant diseases. Nicotine, the major constituent in cigarette smoke, has also shown negative effects on stem cells. Mesenchymal stem cells (MSCs) have been widely demonstrated to migrate into tumors and play key roles in cancer progression. However, the mechanisms by which nicotine impacts MSCs and tumorigenesis of lung cancer are still undetermined. In this study we investigated the effects of nicotine on human umbilical cord mesenchymal stem cells (hUC-MSCs) and the impacts of nicotine-treated hUC-MSCs on tumor formation and progression. We found that nicotine has a toxic effect on hUC-MSCs and changes the morphology, inhibits proliferation and promotes apoptosis of hUC-MSCs in a dose-dependent manner. Nicotine-treated hUC-MSCs produce higher level of IL-6. Moreover, nicotine promotes migration, stemness and epithelial-mesenchymal transition (EMT) of hUC-MSCs by inhibiting E-cadherin expression and upregulating mesenchymal markers such as N-cadherin and Vimentin, leading to the induction of stem cell markers Sox2, Nanog, Sall4, Oct4 and CD44. Migration and proliferation of non-small cell lung cancer A549 cells and breast cancer MCF-7 cells are promoted after their coculture with nicotine-treated hUC-MSCs in a cell-cell contact-independent manner. Furthermore, nicotine-treated hUC-MSCs promote tumor formation and growth of A549 cells in nude mice. These studies demonstrated that the enhanced stemness and EMT of hUC-MSCs induced by nicotine are critical for the development of tobacco-related cancers.

Are Sertoli cells a kind of mesenchymal stem cells?

OBJECTIVE: Sertoli cells (SCs) are a major component of testis which secrete a variety of cytokines and immunosuppressive factors, providing nutritional support and immune protection for sperm growth and development. The purpose of this study was to investigate the relationship between SCs and bone marrow mesenchymal stem cells (BMSCs) in order to provide a theoretical basis for better application of SCs. METHODS: We used the adherence method to isolate Sprague-Dawley rat SCs and BMSCs. Cells surface markers were detected by flow cytometry. The capacity of cells to differentiate was determined by osteogenic and adipogenic induction. Assessment of cell proliferation was performed by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2-H-tetrazolium bromide] assay. Changes in the nucleus were analyzed by Hoechst nuclear staining. Cell aging was observed with ß-galactosidase, which is a biological marker of senescence. RT-PCR was employed to detect the expression of cytokines. RESULTS: From the aforementioned experiments, we found that the surface markers of SCs and BMSCs were almost exactly the same. Proliferation of SCs, as well as osteogenic and adipogenic differentiation, were weaker than in BMSCs. Compared with BMSCs, Hoechst nuclear staining showed that the chromatin of SCs began to aggregate and was slightly larger. ß-galactosidase staining showed that SCs were in a slightly aging state. The secretion of cytokines from SCs was slightly less than the secretion from BMSCs. CONCLUSION: SCs are a kind of mesenchymal stem cells which have begun the process of differentiation.

miR-145 inhibits proliferation and migration of breast cancer cells by directly or indirectly regulating TGF-ß1 expression.

Studies have demonstrated low expression of miR-145 associated with cell proliferation and migration in a wide variety of tumors. Here, we studied the expression of miR-145 in relation to the occurrence and development of breast cancer. Total RNA from breast cancer tissue and corresponding adjacent normal tissue was extracted and used to detect miR-145 expression by quantitative real-time polymerase chain reaction (qRT-PCR). We also transfected breast cancer cells with hsa-miR-145 mimics, hsa-miR-145 inhibitor, mimics negative control (mimics NC) or inhibitor negative control (inhibitor NC). Cell proliferation was analyzed by colony formation assays and methyl thiazolyl tetrazolium assays. Cell proliferation in breast cancer cells was decreased after overexpression of miR-145 and increased following miR-145 suppression. Cell migration and invasion were assessed using Transwell and wound healing assays, respectively, and were also decreased after overexpression of miR-145 and increased after miR-145 suppression in breast cancer cells. Finally, western blot assays showed that overexpression of miR-145 inhibited expression of transforming growth factor-ß1 (TGF-ß1). Collectively, these data suggest that miR-145 may inhibit TGF-ß1 protein expression which may in turn contribute to tumor formation.

MicroRNA-124 inhibits cell proliferation and migration by regulating SNAI2 in breast cancer.

MicroRNA (miRNA) is a type of endogenous non­coding RNA implicated in various cellular processes. Studies have shown that miR-124 is involved in the malignant progression of cancer, but little is known concerning its potential role in breast cancer. Therefore, the purpose of this study was to conduct a functional analysis of miR-124 in breast cancer, and to identify its target genes in this disease. To this end, we used quantitative real-time PCR to examine the expression level of miR-124 in breast cancer tissue specimens and cell lines. To study the functional significance of miR-124, we overexpressed miR-124 with miR-124 mimics and observed breast cancer cell proliferation, colony formation, migration, and invasion abilities by in vitro cell culture experiments. Target prediction algorithms and luciferase reporter gene assays were used to identify the target genes of miR-124. We also knocked down miR-124 targets using short hairpin RNA (shRNA) constructs, and observed associated breast cancer cell characteristics by in vitro cell culture experiments. We found that miR-124 expression significantly decreased in breast cancer tissues and cells compared to normal tissues and cells. In addition, cell proliferation, colony formation, migration, and invasion were decreased after overexpression of miR-124 in breast cancer cells. Furthermore, we used several algorithms to identify the snail family zinc finger 2 (SNAI2) as a potential target gene of miR-124. The protein expression level and luciferase activity of the 3'-untranslated region of SNAI2 were significantly decreased in breast cancer cells transfected with miR-124 mimics. Cell proliferation, colony formation, migration, and invasion were also decreased after knockdown of SNAI2 by shRNA. In conclusion, our data suggest that miR-124 expression is decreased in breast cancer and plays an important role as a tumor suppressor gene by targeting SNAI2. These findings may reveal novel perspectives for clinical treatments against breast cancer.