Amyloid-ß specific regulatory T cells attenuate Alzheimer's disease pathobiology in APP/PS1 mice.

BACKGROUND: Regulatory T cells (Tregs) maintain immune tolerance. While Treg-mediated neuroprotective activities are now well-accepted, the lack of defined antigen specificity limits their therapeutic potential. This is notable for neurodegenerative diseases where cell access to injured brain regions is required for disease-specific therapeutic targeting and improved outcomes. To address this need, amyloid-beta (Aß) antigen specificity was conferred to Treg responses by engineering the T cell receptor (TCR) specific for Aß (TCRAß). The TCRAb were developed from disease-specific T cell effector (Teff) clones. The ability of Tregs expressing a transgenic TCRAß (TCRAß -Tregs) to reduce Aß burden, transform effector to regulatory cells, and reverse disease-associated neurotoxicity proved beneficial in an animal model of Alzheimer's disease. METHODS: TCRAß -Tregs were generated by CRISPR-Cas9 knockout of endogenous TCR and consequent incorporation of the transgenic TCRAb identified from Aß reactive Teff monoclones. Antigen specificity was confirmed by MHC-Aß-tetramer staining. Adoptive transfer of TCRAß-Tregs to mice expressing a chimeric mouse-human amyloid precursor protein and a mutant human presenilin-1 followed measured behavior, immune, and immunohistochemical outcomes. RESULTS: TCRAß-Tregs expressed an Aß-specific TCR. Adoptive transfer of TCRAß-Tregs led to sustained immune suppression, reduced microglial reaction, and amyloid loads. 18F-fluorodeoxyglucose radiolabeled TCRAß-Treg homed to the brain facilitating antigen specificity. Reduction in amyloid load was associated with improved cognitive functions. CONCLUSIONS: TCRAß-Tregs reduced amyloid burden, restored brain homeostasis, and improved learning and memory, supporting the increased therapeutic benefit of antigen specific Treg immunotherapy for AD.

LncRNA XR_001779380 Primes Epithelial Cells for IFN-γ-Mediated Gene Transcription and Facilitates Age-Dependent Intestinal Antimicrobial Defense.

Interferon (IFN) signaling is key to mucosal immunity in the gastrointestinal tract, but cellular regulatory elements that determine interferon gamma (IFN-γ)-mediated antimicrobial defense in intestinal epithelial cells are not fully understood. We report here that a long noncoding RNA (lncRNA), GenBank accession no. XR_001779380, was increased in abundance in murine intestinal epithelial cells following infection by Cryptosporidium, an important opportunistic pathogen in AIDS patients and a common cause of diarrhea in young children. Expression of XR_001779380 in infected intestinal epithelial cells was triggered by TLR4/NF-κB/Cdc42 signaling and epithelial-specific transcription factor Elf3. XR_001779380 primed epithelial cells for IFN-γ-mediated gene transcription through facilitating Stat1/Swi/Snf-associated chromatin remodeling. Interactions between XR_001779380 and Prdm1, which is expressed in neonatal but not adult intestinal epithelium, attenuated Stat1/Swi/Snf-associated chromatin remodeling induced by IFN-γ, contributing to suppression of IFN-γ-mediated epithelial defense in neonatal intestine. Our data demonstrate that XR_001779380 is an important regulator in IFN-γ-mediated gene transcription and age-associated intestinal epithelial antimicrobial defense. IMPORTANCE Epithelial cells along the mucosal surface provide the front line of defense against luminal pathogen infection in the gastrointestinal tract. These epithelial cells represent an integral component of a highly regulated communication network that can transmit essential signals to cells in the underlying intestinal mucosa that, in turn, serve as targets of mucosal immune mediators. LncRNAs are recently identified long noncoding transcripts that can regulate gene transcription through their interactions with other effect molecules. In this study, we demonstrated that lncRNA XR_001779380 was upregulated in murine intestinal epithelial cells following infection by a mucosal protozoan parasite Cryptosporidium. Expression of XR_001779380 in infected cells primed host epithelial cells for IFN-γ-mediated gene transcription, relevant to age-dependent intestinal antimicrobial defense. Our data provide new mechanistic insights into how intestinal epithelial cells orchestrate intestinal mucosal defense against microbial infection.

m6A mRNA Methylation Regulates Epithelial Innate Antimicrobial Defense Against Cryptosporidial Infection.

Increasing evidence supports that N6-methyladenosine (m6A) mRNA modification may play an important role in regulating immune responses. Intestinal epithelial cells orchestrate gastrointestinal mucosal innate defense to microbial infection, but underlying mechanisms are still not fully understood. In this study, we present data demonstrating significant alterations in the topology of host m6A mRNA methylome in intestinal epithelial cells following infection by Cryptosporidium parvum, a coccidian parasite that infects the gastrointestinal epithelium and causes a self-limited disease in immunocompetent individuals but a life-threatening diarrheal disease in AIDS patients. Altered m6A methylation in mRNAs in intestinal epithelial cells following C. parvum infection is associated with downregulation of alpha-ketoglutarate-dependent dioxygenase alkB homolog 5 and the fat mass and obesity-associated protein with the involvement of NF-кB signaling. Functionally, m6A methylation statuses influence intestinal epithelial innate defense against C. parvum infection. Specifically, expression levels of immune-related genes, such as the immunity-related GTPase family M member 2 and interferon gamma induced GTPase, are increased in infected cells with a decreased m6A mRNA methylation. Our data support that intestinal epithelial cells display significant alterations in the topology of their m6A mRNA methylome in response to C. parvum infection with the involvement of activation of the NF-кB signaling pathway, a process that modulates expression of specific immune-related genes and contributes to fine regulation of epithelial antimicrobial defense.

Knockdown of m6A methyltransferase METTL3 in gastric cancer cells results in suppression of cell proliferation.

N6-methyladenosine (m6A) RNA modification regulates multiple biological functions. Methyltransferase like 3 (METTL3), one of the major N6-methyltransferases, is highly expressed in gastric cancer, but its potential role in disease is unclear. The current study knocked out METTL3 (METTL3-KO) in human gastric cancer AGS cells using CRISPR/Cas9. METTL3-KO AGS cells exhibited decreased m6A methylation levels. A significant inhibition of cell proliferation was observed in METTL3-KO AGS cells. Silencing METTL3 in AGS cells altered the expression profile of many effector molecules that were previously demonstrated to serve key roles in AGS cell proliferation, including the suppressor of cytokine signaling (SOCS) family of proteins. The results further demonstrated that SOCS2 upregulation in METTL3-KO AGS cells was associated with a decreased RNA decay rate. Furthermore, SOCS2 KO or SOCS2 overexpression caused a significant increase and decrease in AGS cell proliferation, respectively. The current data suggested that METTL3-KO in gastric cancer cells resulted in the suppression of cell proliferation by inducing SOCS2, suggesting a potential role of elevated METTL3 expression in gastric cancer progression.

Directional migration of cancer cells induced by a blue light intensity gradient.

We used a spatial light modulator to project an optical micropattern of 473 nm light with a quartic intensity gradient on a single lung cancer cell. We observed that the intracellular amounts of reactive oxygen species (ROS) of the cancer cells were proportional to the intensity of the blue light, and the blue light intensity gradients could drive directional cell migration. This optically induced directional cell migration was inhibited by a ROS scavenger in the culture medium in a dose-dependent manner. In contrast, the ROS levels in fibroblasts were saturated by the blue light at low intensity and therefore the fibroblasts did not exhibit directional migration in the intensity gradient.