The microRNA-212/132 cluster regulates B cell development by targeting Sox4.

MicroRNAs have emerged as key regulators of B cell fate decisions and immune function. Deregulation of several microRNAs in B cells leads to the development of autoimmune disease and cancer in mice. We demonstrate that the microRNA-212/132 cluster (miR-212/132) is induced in B cells in response to B cell receptor signaling. Enforced expression of miR-132 results in a block in early B cell development at the prepro-B cell to pro-B cell transition and induces apoptosis in primary bone marrow B cells. Importantly, loss of miR-212/132 results in accelerated B cell recovery after antibody-mediated B cell depletion. We find that Sox4 is a target of miR-132 in B cells. Co-expression of SOX4 with miR-132 rescues the defect in B cell development from overexpression of miR-132 alone, thus suggesting that miR-132 may regulate B lymphopoiesis through Sox4. In addition, we show that the expression of miR-132 can inhibit cancer development in cells that are prone to B cell cancers, such as B cells expressing the c-Myc oncogene. We have thus uncovered miR-132 as a novel contributor to B cell development.

MicroRNA-132 enhances transition from inflammation to proliferation during wound healing.

Wound healing is a complex process that is characterized by an initial inflammatory phase followed by a proliferative phase. This transition is a critical regulatory point; however, the factors that mediate this process are not fully understood. Here, we evaluated microRNAs (miRs) in skin wound healing and characterized the dynamic change of the miRNome in human skin wounds. miR-132 was highly upregulated during the inflammatory phase of wound repair, predominantly expressed in epidermal keratinocytes, and peaked in the subsequent proliferative phase. TGF-ß1 and TGF-ß2 induced miR-132 expression in keratinocytes, and transcriptome analysis of these cells revealed that miR-132 regulates a large number of immune response- and cell cycle-related genes. In keratinocytes, miR-132 decreased the production of chemokines and the capability to attract leukocytes by suppressing the NF-κB pathway. Conversely, miR-132 increased activity of the STAT3 and ERK pathways, thereby promoting keratinocyte growth. Silencing of the miR-132 target heparin-binding EGF-like growth factor (HB-EGF) phenocopied miR-132 overexpression in keratinocytes. Using mouse and human ex vivo wound models, we found that miR-132 blockade delayed healing, which was accompanied by severe inflammation and deficient keratinocyte proliferation. Together, our results indicate that miR-132 is a critical regulator of skin wound healing that facilitates the transition from the inflammatory to the proliferative phase.

The MicroRNA-132 and MicroRNA-212 Cluster Regulates Hematopoietic Stem Cell Maintenance and Survival with Age by Buffering FOXO3 Expression.

MicroRNAs are critical post-transcriptional regulators of hematopoietic cell-fate decisions, though little remains known about their role in aging hematopoietic stem cells (HSCs). We found that the microRNA-212/132 cluster (Mirc19) is enriched in HSCs and is upregulated during aging. Both overexpression and deletion of microRNAs in this cluster leads to inappropriate hematopoiesis with age. Enforced expression of miR-132 in the bone marrow of mice led to rapid HSC cycling and depletion. A genetic deletion of Mirc19 in mice resulted in HSCs that had altered cycling, function, and survival in response to growth factor starvation. We found that miR-132 exerted its effect on aging HSCs by targeting the transcription factor FOXO3, a known aging associated gene. Our data demonstrate that Mirc19 plays a role in maintaining balanced hematopoietic output by buffering FOXO3 expression. We have thus identified it as a potential target that might play a role in age-related hematopoietic defects.

The BTB-ZF transcription factor Zbtb20 is driven by Irf4 to promote plasma cell differentiation and longevity.

The transcriptional network regulating antibody-secreting cell (ASC) differentiation has been extensively studied, but our current understanding is limited. The mechanisms of action of known "master" regulators are still unclear, while the participation of new factors is being revealed. Here, we identify Zbtb20, a Bcl6 homologue, as a novel regulator of late B cell development. Within the B cell lineage, Zbtb20 is specifically expressed in B1 and germinal center B cells and peaks in long-lived bone marrow (BM) ASCs. Unlike Bcl6, an inhibitor of ASC differentiation, ectopic Zbtb20 expression in primary B cells facilitates terminal B cell differentiation to ASCs. In plasma cell lines, Zbtb20 induces cell survival and blocks cell cycle progression. Immunized Zbtb20-deficient mice exhibit curtailed humoral responses and accelerated loss of antigen-specific plasma cells, specifically from the BM pool. Strikingly, Zbtb20 induction does not require Blimp1 but depends directly on Irf4, acting at a newly identified Zbtb20 promoter in ASCs. These results identify Zbtb20 as an important player in late B cell differentiation and provide new insights into this complex process.

The miRNA-212/132 family regulates both cardiac hypertrophy and cardiomyocyte autophagy.

Pathological growth of cardiomyocytes (hypertrophy) is a major determinant for the development of heart failure, one of the leading medical causes of mortality worldwide. Here we show that the microRNA (miRNA)-212/132 family regulates cardiac hypertrophy and autophagy in cardiomyocytes. Hypertrophic stimuli upregulate cardiomyocyte expression of miR-212 and miR-132, which are both necessary and sufficient to drive the hypertrophic growth of cardiomyocytes. MiR-212/132 null mice are protected from pressure-overload-induced heart failure, whereas cardiomyocyte-specific overexpression of the miR-212/132 family leads to pathological cardiac hypertrophy, heart failure and death in mice. Both miR-212 and miR-132 directly target the anti-hypertrophic and pro-autophagic FoxO3 transcription factor and overexpression of these miRNAs leads to hyperactivation of pro-hypertrophic calcineurin/NFAT signalling and an impaired autophagic response upon starvation. Pharmacological inhibition of miR-132 by antagomir injection rescues cardiac hypertrophy and heart failure in mice, offering a possible therapeutic approach for cardiac failure.

Differential immune activation following encapsulation of immunostimulatory CpG oligodeoxynucleotide in nanoliposomes.

The immunogenicity of a vaccine formulation is closely related to the effective internalization by the innate immune cells that provide prolonged and simultaneous delivery of antigen and adjuvant to relevant antigen presenting cells. Endosome associated TLR9 recognizes microbial unmethylated CpG DNA. Clinical applications of TLR9 ligands are significantly hampered due to their pre-mature in vivo digestion and rapid clearance. Liposome encapsulation is a powerful tool to increase in vivo stability as well as enhancing internalization of its cargo to relevant immune cells. The present study established that encapsulating CpG motifs in different liposomes having different physicochemical properties altered not only encapsulation efficiency, but also the release and delivery rates that ultimately impacted in vitro and ex-vivo cytokine production rates and types. Moreover, different liposomes encapsulating CpG ODN significantly increased Th1-biased cytokines and chemokines gene transcripts Additional studies demonstrated that co-stimulatory and surface marker molecules significantly upregulated upon liposome/CpG injection. Finally, co-encapsulating model antigen ovalbumin with CpG ODN adjuvant in nanoliposomes profoundly augmented Th1 and cell mediated anti-Ova specific immune response. Collectively, this work established an unappreciated immunoregulatory property of nanoliposomes mediating immunity against protein antigen and could be harnessed to design more effective therapeutic vaccines or stand alone immunoprotective agents targeting infectious diseases, as well as cancer or allergy.