Secretory granules (SGs) of mast cells are lysosome-related organelles that contain various inflammatory molecules such as histamine, which are stored in the cytoplasm. Mast cell degranulation is the regulated exocytosis of SGs in response to external stimuli, such as the antigen-mediated cross-linking of the high-affinity IgE receptor, FcεRI. Upon stimulation, SGs undergo priming to become fusion-competent prior to fusing with the plasma membrane, which is mediated by Munc13-4, one of the five members of the vesicle-priming Munc13 protein family. Although Munc13-4 is shown to be crucial for mast cell degranulation, the functional involvement of other Munc13 isoform(s) remains unknown. Herein, this was investigated using the RBL-2H3 mast cell line. We found that Munc13-1 and Munc13-4 are the only Munc13 isoforms that are expressed in the RBL-2H3 cells, and Munc13-1 is distributed in the cytoplasm, but highly concentrated on the late endosome and/or lysosome. Unexpectedly, antigen-induced degranulation was considerably increased by Munc13-1 knockdown, but decreased by its overexpression. Further, we found that the hypersecretion phenotype of the Munc13-1-knockdown cells was attenuated by simultaneous Munc13-4 knockdown. These results suggested that Munc13-1 has an inhibitory role in antigen-induced mast cell degranulation, which is performed in a Munc13-4-dependent manner.
The present study aimed to examine the changes in the expression of Forkhead box protein O (FOXO) and proinflammatory cytokines in both slow-twitch soleus and fast-twitch plantaris muscles following cast immobilization. Male C57BL/6 mice were subjected to cast immobilization for 7 and 21 days. Cast immobilization increased FOXO3a mRNA and total protein expression in both the soleus and plantaris muscles. Although FOXO3a phosphorylation tended to increase in response to cast immobilization in both muscles, a significant increase was evident after 21 days of immobilization only in the soleus muscle. The degree of the response of FOXO3a was very different between the soleus and plantaris muscles; however, the kinetics of FOXO3a in both muscles were similar. Thus, the regulation of muscle atrophy by FOXO might act via a common mechanism in both slow-twitch soleus and fast-twitch plantaris muscles. Gene expression of proinflammatory cytokines tended to increase in response to cast immobilization, and a significant increase was evident after 21 days of immobilization in the soleus muscle. However, in the plantaris muscle, proinflammatory cytokine gene expression remained unchanged throughout the immobilization period; nevertheless, immobilization induced greater reduction in muscle fiber cross-sectional area in the plantaris than in the soleus muscle. Thus, these observations indicate that regulation of muscle atrophy by proinflammatory cytokines might contribute to muscle fiber type-specific mechanisms.
Iron is an essential trace metal in almost all organisms and plays an important role in the redox system. We previously reported that iron deficiency activated autophagy and nuclear factor erythroid 2-related factor 2 (Nrf2) signaling for oxidative stress. However, regulatory mechanisms underlying the association between autophagy and Nrf2 signaling are unclear. In this study, we found that treatment of cells with an iron-specific chelator deferoxamine (DFO) increased reactive oxidative species (ROS) production by elevating the expression of p47phox and p67phox compared with that in untreated cells. The DFO treatment also induced protein aggregation and formed aggresome, which is a cellular response to misfolded protein. In addition, DFO treatment upregulated the expression of the autophagic gene p62/SQSTM1, which in turn activated intracellular proteolysis during autophagy. DFO treatment phosphorylated p62/SQSTM1 (Thr351) to activate Nrf2. However, silencing of p62/SQSTM1 followed by DFO treatment attenuated Nrf2 activation and resulted in the accumulation of carboxyl proteins compared with DFO treatment alone. These results indicated that iron deficiency activates Nrf2 signaling by modulating p62/SQSTM1 during autophagy.
Fish protein is a source of animal protein that is consumed worldwide. Although it has been reported that the intake of Alaska pollack protein (APP) reduces body fat accumulation and increases muscle weight in rats, the mechanisms underlying these effects are poorly understood. As a possibility, peptides released from APP in the gastrointestinal tract are important to the functions of APP. In the present study, we examined the effects of APP hydrolysate digested artificially with pepsin and pancreatin on white adipose tissue and skeletal muscle. We found that APP hydrolysate group shows significantly lower weight of white adipose tissue and higher weight of soleus muscle than the control group. We also found that APP hydrolysate group reduces food intake and mRNA expressions of neuropeptide Y and agouti-related protein in the hypothalamus compared with the control group. These results may imply that APP hydrolysate exhibits anti-obesity activity by the reduction of appetite and the enhancement of basal energy expenditure by skeletal muscle hypertrophy in rats. The downregulation of orexigenic gene by APP hydrolysate in the hypothalamus may contribute to the reduction of appetite. These results suggest that the effect of APP on anti-obesity and muscle hypertrophy may be induced by peptides released from APP in the gastrointestinal tract.
Noradrenergic modulation has been frequently discussed in the context of neural activities that are related to pelvic organs. The sacral preganglionic nucleus (SPN) is a spinal nucleus containing parasympathetic preganglionic neurons that send fibers to pelvic nerves. In spite of the abundant presence of noradrenergic fibers around the SPN, the effects of noradrenaline (NA) remain obscure. To explore this issue, NA (50 μM) was applied to parasympathetic preganglionic neurons in the SPN during whole-cell patch clamp recording. The SPN was labeled with the retrograde tracer, DiI. These neurons demonstrated two classes of firing patterns (delayed and regular) in terms of initiation of firing. Independent of these firing patterns, NA induced inward (56%) or outward (32%) currents in labeled SPN neurons. Phenylephrine, an α1 receptor agonist, induced an inward current, and clonidine, an α2 receptor agonist, induced an outward current, indicating the existence of both α1 and α2 adrenoreceptors in DiI-labeled SPN neurons. NA also modulated synaptic currents according to the firing patterns. In delayed firing neurons, NA inhibited both spontaneous excitatory post-synaptic currents (sEPSCs) and spontaneous inhibitory post-synaptic currents (sIPSCs). Hence, NA facilitated sEPSCs and sIPSCs in about a half of regular firing neurons. Bath application of phenylephrine facilitated sEPSCs and sIPSCs, and clonidine inhibited them. These results support the hypothesis of multiple effects of NA in the SPN, and may suggest functional differences among SPN neurons.
Interferon (IFN) has various side effects, including psychiatric symptoms. Event-related potentials are used as an electrophysiologic index of cognitive disorders. Auditory event-related potentials (P300) are often used in conditions in which cognitive ability is affected. In this study, we evaluated the association between P300, used to assess cognitive impairment, and neuropsychological side effects of IFN treatment in patients with chronic hepatitis C. Subjects were 20 patients with chronic hepatitis C; 13 patients were treated with peg IFN-α2b and ribavirin (riba group), and 7 patients were treated with peg IFN-α2a (alone group). P300 was performed on all patients before treatment and after 1 week, 4 weeks, 2 months, and 3 months of treatment. In addition, 10 patients of them completed the self-rating depression scale (SDS). P300 latency was significantly prolonged at all points of measurement during IFN treatment. No correlation between the change of SDS score and the change rate of P300 latency was shown. Six patients with neuropsychological symptom had a significantly increased change rate of P300 latency compared with patients without neuropsychological symptoms (P < 0.05). Based on P300 findings, this study suggests that patients with chronic hepatitis C treated with IFN may experience significant cognitive impairment.