Bone metabolism is regulated by the cooperative activity between bone-forming osteoblasts and bone-resorbing osteoclasts. However, the mechanisms mediating the switch between the osteoblastic and ...osteoclastic phases have not been fully elucidated. Here, we identify a specific subset of mature osteoblast-derived extracellular vesicles that inhibit bone formation and enhance osteoclastogenesis. Intravital imaging reveals that mature osteoblasts secrete and capture extracellular vesicles, referred to as small osteoblast vesicles (SOVs). Co-culture experiments demonstrate that SOVs suppress osteoblast differentiation and enhance the expression of receptor activator of NF-κB ligand, thereby inducing osteoclast differentiation. We also elucidate that the SOV-enriched microRNA miR-143 inhibits Runt-related transcription factor 2, a master regulator of osteoblastogenesis, by targeting the mRNA expression of its dimerization partner, core-binding factor β. In summary, we identify SOVs as a mode of cell-to-cell communication, controlling the dynamic transition from bone-forming to bone-resorbing phases in vivo.
Abstract
Blood pressure has a daily pattern, with higher values in the active period. Its elevation at the onset of the active period substantially increases the risk of fatal cardiovascular events. ...Renin secretion stimulated by renal sympathetic neurons is considered essential to this process; however, its regulatory mechanism remains largely unknown. Here, we show the importance of transient receptor potential melastatin-related 6 (TRPM6), a Mg
2+
-permeable cation channel, in augmenting renin secretion in the active period. TRPM6 expression is significantly reduced in the distal convoluted tubule of hypotensive
Cnnm2
-deficient mice. We generate kidney-specific
Trpm6
-deficient mice and observe a decrease in blood pressure and a disappearance of its circadian variation. Consistently, renin secretion is not augmented in the active period. Furthermore, renin secretion after pharmacological activation of β-adrenoreceptor, the target of neuronal stimulation, is abrogated, and the receptor expression is decreased in renin-secreting cells. These results indicate crucial roles of TRPM6 in the circadian regulation of blood pressure.
The biological significance of micro (mi)RNAs has traditionally been evaluated according to their RNA expression levels based on the assumption that miRNAs recognize and regulate their targets in an ...unvarying fashion. Here we show that a fraction of mature miRNAs including miR-17-5p, -21-5p, and -200c-3p and let-7a-5p harbor methyl marks that potentially alter their stability and target recognition. Importantly, methylation of these miRNAs was significantly increased in cancer tissues as compared to paired normal tissues. Furthermore, miR-17-5p methylation level in serum samples distinguished early pancreatic cancer patients from healthy controls with extremely high sensitivity and specificity. These findings provide a basis for diagnostic strategies for early-stage cancer and add a dimension to our understanding of miRNA biology.
Osteoclasts have a unique bone-destroying capacity, playing key roles in steady-state bone remodeling and arthritic bone erosion. Whether the osteoclasts in these different tissue settings arise from ...the same precursor states of monocytoid cells is presently unknown. Here, we show that osteoclasts in pannus originate exclusively from circulating bone marrow-derived cells and not from locally resident macrophages. We identify murine CX
CR1
Ly6C
F4/80
I-A
/I-E
macrophages (termed here arthritis-associated osteoclastogenic macrophages (AtoMs)) as the osteoclast precursor-containing population in the inflamed synovium, comprising a subset distinct from conventional osteoclast precursors in homeostatic bone remodeling. Tamoxifen-inducible Foxm1 deletion suppressed the capacity of AtoMs to differentiate into osteoclasts in vitro and in vivo. Furthermore, synovial samples from human patients with rheumatoid arthritis contained CX
CR1
HLA-DR
CD11c
CD80
CD86
cells that corresponded to mouse AtoMs, and human osteoclastogenesis was inhibited by the FoxM1 inhibitor thiostrepton, constituting a potential target for rheumatoid arthritis treatment.
Lung cancer is a common type of cancer that represents a health problem worldwide; lung adenocarcinoma (LUAD) is a major subtype of lung cancer. Although several treatments for LUAD have been ...developed, the mortality rate remains high because of uncontrollable progression. Further biological and clinicopathological studies are therefore needed. Here, we investigated the role of family with sequence similarity 111 member B (FAM111B), which is highly expressed in papillary‐predominant LUAD; however, its role in cancer is unclear. An immunohistochemical analysis confirmed that papillary‐predominant adenocarcinomas exhibited higher expression of FAM111B, compared with lepidic‐predominant adenocarcinomas. Additionally, FAM111B expression was significantly correlated with clinical progression. In vitro functional analyses using FAM111B‐knockout cells demonstrated that FAM111B plays an important role in proliferation and cell cycle progression of KRAS‐driven LUAD under serum‐starvation conditions. Furthermore, FAM111B regulated cyclin D1‐CDK4‐dependent cell cycle progression by degradation of p16. In summary, we revealed the clinical importance of FAM111B in human tumor tissues, as well as its function as a degradative enzyme. Therefore, FAM111B has potential as a clinicopathological prognostic marker for LUAD.
We screened family with sequence similarity 111 member B (FAM111B), the precise functional role of which in cancers is not clear, as the molecule is highly expressed in papillary‐predominant lung adenocarcinoma. Immunohistochemical analysis confirmed that papillary‐predominant adenocarcinoma had higher expression of FAM111B compared with lepidic‐predominant adenocarcinoma, and FAM111B expression levels were significantly correlated with patients’ clinical progression. In vitro functional analyses using FAM111B‐knockout (FAM111B‐KO) cells demonstrated that FAM111B plays an important role in proliferation and cell cycle progression of LUAD with the KRAS mutation, specifically under the conditions of serum starvation, and it was revealed that FAM111B negatively regulates CyclinD1‐CDK4‐dependent cell cycle progression by functioning as a degrading enzyme to control p16 expression level.
The frontal craniofacial skeleton derived from neural crest cells is vital for facial structure and masticatory functions. The exact role of Indian hedgehog (Ihh) in facial and masticatory ...development has not been fully explored. In this study, we generated craniofacial neural crest cells‐specific Ihh deletion mice (Wnt1‐Cre;Ihhfl/fl;Tomatofl/+) and found the gradual dwarfism without perinatal lethality. Morphological and histological analyses revealed unambiguous craniofacial phenotypes in mutants, where we observed skeletal malocclusion accompanied by markedly hypoplastic nasomaxillary complex and reversed incisor occlusion. Both the replacement of nasal concha cartilage by turbinate bones and the endochondral ossification of nasal septum ethmoid bone were substantially delayed. We also observed hypoplastic mandibles in mutants where the mandibular ramus was unexpectedly the most affected. Both the condylar process and mandibular angle cartilages were distorted. However, dental examination showed no significant changes in teeth and dentition. Finally, a comprehensive RNA sequence analysis utilizing condylar cartilage identified Ihh‐associated gene network including several cell cycle genes and 16 genes related to the extracellular matrix, sulfate transporters, transcription factors, receptors, a ciliogenesis factor, and an adhesion molecule. Our data provide direct in vivo evidence that Ihh plays crucial roles in midface and masticatory system formation, likely by activating key genes.
The non‐receptor tyrosine kinase c‐Src is frequently activated during progression of colon cancers. In this study, we found that among the c‐Src‐regulated microRNAs (miRNAs), miR‐27b is also ...repressed by activation of K‐Ras/H‐Ras. Inhibitor studies suggested that the phosphatidylinositol 3‐kinase pathway is involved in the repression of miR‐27b. MicroRNA‐27b was repressed in various colon cancer cell lines and tumor tissues. Re‐expression of miR‐27b in human colon cancer HCT116 cells caused morphological changes and suppressed tumor growth, cell adhesion, and invasion. We also identified ARFGEF1 and paxillin as novel targets of miR‐27b, and found that miR‐27b‐mediated regulation of ARFGEF1 is crucial for controlling anchorage‐independent growth, and that of paxillin is important for controlling cell adhesion and invasion. Re‐expression of miR‐27b suppressed the activation of c‐Src induced by integrin‐mediated cell adhesion, suggesting that repression of miR‐27b may contribute to c‐Src activation in cancer cells. These findings show that miR‐27b functions as a tumor suppressor by controlling ARFGEF1 and the paxillin/c‐Src circuit at focal adhesions.
miR‐27b functions as a tumor suppressor by controlling ARFGEF1 and the paxillin/c‐Src circuit at focal adhesions.
HIF-1 induces LDH to decrease pyruvate in Mtb-infected macrophages
Abstract
Macrophages are major components of tuberculosis (TB) granulomas and are responsible for host defenses against the ...intracellular pathogen, Mycobacterium tuberculosis. We herein showed the strong expression of hypoxia-inducible factor-1α (HIF-1α) in TB granulomas and more rapid death of HIF-1α-conditional knockout mice than wild-type (WT) mice after M. tuberculosis infection. Although interferon-γ (IFN-γ) is a critical host-protective cytokine against intracellular pathogens, HIF-1-deficient macrophages permitted M. tuberculosis growth even after activation with IFN-γ. These results prompted us to investigate the role of HIF-1α in host defenses against infection. We found that the expression of lactate dehydrogenase-A (LDH-A) was controlled by HIF-1α in M. tuberculosis-infected macrophages IFN-γ independently. LDH-A is an enzyme that converts pyruvate to lactate and we found that the intracellular level of pyruvate in HIF-1α-deficient bone marrow-derived macrophages (BMDMs) was significantly higher than in WT BMDMs. Intracellular bacillus replication was enhanced by an increase in intracellular pyruvate concentrations, which were decreased by LDH-A. Mycobacteria in phagosomes took up exogenous pyruvate more efficiently than glucose, and used it as the feasible carbon source for intracellular growth. These results demonstrate that HIF-1α prevents the hijacking of pyruvate in macrophages, making it a fundamental host-protective mechanism against M. tuberculosis.
The mammalian target of rapamycin complex 1 (mTORC1) signaling pathway is activated by intracellular nutritional sufficiency and extracellular growth signals. It has been reported that mTORC1 acts as ...a hub that integrates these inputs to orchestrate a number of cellular responses, including translation, nucleotide synthesis, lipid synthesis, and lysosome biogenesis. However, little is known about specific control of mTORC1 signaling downstream of this complex. Here, we demonstrate that Ragulator, a heteropentameric protein complex required for mTORC1 activation in response to amino acids, is critical for inhibiting the nuclear translocation of transcription factor EB (TFEB). We established a unique RAW264.7 clone that lacked Ragulator but retained total mTORC1 activity. In a nutrition-sufficient state, the nuclear translocation of TFEB was markedly enhanced in the clone despite total mTORC1 kinase activity. In addition, as a cellular phenotype, the number of lysosomes was increased by tenfold in the Ragulator-deficient clone compared with that of control cells. These findings indicate that mTORC1 essentially requires the Ragulator complex for regulating the subcellular distribution of TFEB. Our findings also suggest that other scaffold proteins may be associated with mTORC1 for the specific regulation of downstream signaling.
Osteoclastic bone resorption and osteoblastic bone formation/replenishment are closely coupled in bone metabolism. Anabolic parathyroid hormone (PTH), which is commonly used for treating ...osteoporosis, shifts the balance from osteoclastic to osteoblastic, although it is unclear how these cells are coordinately regulated by PTH. Here, we identify a serine protease inhibitor, secretory leukocyte protease inhibitor (SLPI), as a critical mediator that is involved in the PTH-mediated shift to the osteoblastic phase. Slpi is highly upregulated in osteoblasts by PTH, while genetic ablation of Slpi severely impairs PTH-induced bone formation. Slpi induction in osteoblasts enhances its differentiation, and increases osteoblast-osteoclast contact, thereby suppressing osteoclastic function. Intravital bone imaging reveals that the PTH-mediated association between osteoblasts and osteoclasts is disrupted in the absence of SLPI. Collectively, these results demonstrate that SLPI regulates the communication between osteoblasts and osteoclasts to promote PTH-induced bone anabolism.
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