Osteoclasts seem to be metabolic active during their differentiation and bone-resorptive activation. However, the functional role of lactate dehydrogenase (LDH), a tetrameric enzyme consisting of an A and/or B subunit that catalyzes interconversion of pyruvate to lactate, in RANKL-induced osteoclast differentiation is not known. In this study, RANKL treatment induced gradual gene expression and activation of the LDH A2B2 isotype during osteoclast differentiation as well as the LDH A1B3 and B4 isotypes during osteoclast maturation after pre-osteoclast formation. Glucose consumption and lactate production in growth media were accelerated during osteoclast differentiation, together with enhanced expression of H+-lactate co-transporter and increased extracellular acidification, demonstrating that glycolytic metabolism was stimulated during differentiation. Further, oxygen consumption via mitochondria was stimulated during osteoclast differentiation. On the contrary, depletion of LDH-A or LDH-B subunit suppressed both glycolytic and mitochondrial metabolism, resulting in reduced mature osteoclast formation via decreased osteoclast precursor fusion and down-regulation of the osteoclastogenic critical transcription factor NFATc1 and its target genes. Collectively, our findings suggest that RANKL-induced LDH activation stimulates glycolytic and mitochondrial respiratory metabolism, facilitating mature osteoclast formation via osteoclast precursor fusion and NFATc1 signaling.

The cell proliferation process of aerial lateral organs, such as leaves and flowers, is coordinated by complex genetic networks that, in general, converge on the cell cycle. The Arabidopsis thaliana NGATHA (AtNGA) family comprises four members that belong to the B3-type transcription factor superfamily, and has been suggested to be involved in growth and development of aerial lateral organs, although its role in the cell proliferation and expansion processes remains to be resolved in more detail. In order to clarify the role of AtNGAs in lateral organ growth, we took a systematic approach using both the loss- and gain-of-functional mutants of all four members. Our results showed that overexpressors of AtNGA1 to AtNGA4 developed small, narrow lateral organs, whereas the nga1 nga2 nga3 nga4 quadruple mutant produced large, wide lateral organs. We found that cell numbers of the lateral organs were significantly affected: a decrease in overexpressors and, inversely, an increase in the quadruple mutant. Kinematic analyses on leaf growth revealed that, compared with the wild type, the overexpressors displayed a lower activity of cell proliferation and yet the mutant a higher activity. Changes in expression of cell cycle-regulating genes were well in accordance with the cell proliferation activities, establishing that the AtNGA transcription factors act as bona fide negative regulators of the cell proliferation of aerial lateral organs.

In an effort to elucidate biological functions of transcription factors of Brassica rapa L. (ssp. pekinensis), an NGATHA homolog, BrNGA1, that belongs to the B3-type transcription factor superfamily was identified and expressed in Arabidopsis thaliana under the control of the cauliflower mosaic virus (CaMV) 35S promoter. Arabidopsis plants overexpressing BrNGA1, named BrNGA1ox, displayed markedly reduced organ growth compared with the wild type: lateral organs, such as leaves, flowers and cotyledons, were small and distinctively narrow, and their root growth was also severely retarded. Reduced sizes of BrNGA1ox organs were mainly due to reduction in cell numbers. Kinematic analysis of leaf growth revealed that both the rate and duration of cell proliferation declined during organogenesis, which was consistent with the reduced expression of cyclin genes. Reduction in organ growth was strongly correlated with the small size of meristematic cell pools in the shoot and root meristems. Taken together, these data indicate that BrNGA1 acts as a negative regulator of cell proliferation and may do so, in part, by regulating the size of the meristematic cell pool.

Intestinal fibrosis induced by chronic and recurrent colitis, which is exacerbated by bowel stenosis, stricture, and obstruction, is challenging to treat. Toll-like receptor 4 (TLR4) stimulates innate and acquired immunity in response to specific microbial components, but the role of TLR4 in intestinal fibrosis is largely unknown. We investigated its role in intestinal fibrosis using not only a murine fibrosis model but also human myofibroblasts and intestinal epithelial cells. Colon fibrosis was induced in TLR4-deficient (TLR4−/−) mice and its wild-type counterparts with 3% dextran sulfate sodium. Absence of TLR4 gene attenuated chronic inflammation and colonic macrophages infiltration; intestinal fibrosis and collagen deposition were suppressed. Also, the production of tumor necrosis factor-α, interleukin-12p40, and transforming growth factor-β was reduced in TLR4-deficient peritoneal macrophages. TLR4 was silenced in CCD-18Co cells by small interfering RNA (siRNA), and matrix metalloproteinase-1, tissue inhibitor of metalloproteinase, and collagen α1 expression was evaluated. Role of TLR4 in epithelial-mesenchymal transition (EMT) was evaluated in HCT116 cells. Suppression of TLR4 transcription by siRNAs affected myofibroblasts activity, collagen synthesis, and EMT in the human cancer cell line. Thus, we suggest that TLR4 can be an essential mediator in intestinal chronic inflammation and fibrosis, indicating that TLR4 signaling is a potential therapeutic target for intestinal fibrosis.