Nucling is an Apaf1‐binding proapoptotic protein involved in apoptosome‐mediated apoptosis. Luciferase assays have revealed that the activation of nuclear factor‐κB induced by tumor necrosis factor‐α, interleukin‐1β and lipopolysaccharide is downregulated by the overexpression of Nucling in HEK293 cells. Moreover, the expression of endogenous cyclooxygenase 2, tumor necrosis factor‐α and galectin‐3, the end‐point molecules in the pathway for the activation of nuclear factor‐κB, as well as nuclear factor‐κB (p65) itself, is upregulated in Nucling gene‐deficient mouse embryonic fibroblasts, suggesting that nuclear factor‐κB is a target of Nucling. Subsequent study has revealed that Nucling physically interacts with nuclear factor‐κB (p65 and p50) and that the binding domain of Nucling is its amino‐terminal region (amino acids 1–466) containing ankyrin repeats. Overexpression of Nucling prevents the translocation of nuclear factor‐κB into the nucleus. In addition, the cytoplasmic retention of endogenous nuclear factor‐κB in resting cells is not observed in Nucling gene‐deficient mouse embryonic fibroblasts. These results reveal a novel function of Nucling as a suppressor of nuclear factor‐κB, mediated by its cytoplasmic retention through physical interaction.

Nucling is a stress‐inducible protein associated with apoptosomes. The cytochrome c‐triggered formation of apoptosomes represents a key‐initiating event in apoptosis. We have recently reported that Nucling regulates the apoptotic pathway by controlling the activation of NF‐κB as well. Here we show that hepatocellular carcinoma (HCC) arising spontaneously against a background of hepatitis occurred more frequently in Nucling‐knockout (KO) mice than wild‐type (WT) mice. Biochemical serum testing revealed potential liver dysfunction with hypercholesterolemia in Nucling‐KO males. In the background of Nucling‐KO mice, we observed the up‐regulation of TNFα, spontaneous NF‐κB‐activation and the induction of galectin‐3 expression in liver. In addition, we observed a decrease in the number of Kupffer cells (KCs) in the KO mice. KCs are important for the hepatic immune system, acting as phagocytes or antigen‐presenting cells (APCs). We found that KCs in Nucling‐KO mice were apoptotic possibly through the up‐regulation of TNFα. These observations indicate that Nucling is important for the regulation of NF‐κB signals in liver. We propose that Nucling deficiency could be a powerful tool to reveal the NF‐κB‐related molecular networks leading to hepatitis and HCC development.

Nucling is a proapoptotic protein that regulates the apoptosome and nuclear factor-kappa B (NF-κB) signalling pathways. Strong stimuli, such as Gram-negative bacterial lipopolysaccharide (LPS), induce the simultaneous secretion of cytokines following the activation of NF-κB. Proinflammatory cytokines can induce liver damage through several mechanisms such as increases in oxidative stress and apoptotic reactions leading to tissue necrosis. Herein, we show that Nucling-knockout (KO) mice are resistant to LPS that consistently caused mortality in wild-type (WT) counterparts. Although serum levels of cytokines such as tumour necrosis factor (TNF)-α, interleukin (IL)-1β and IL-6 did not differ significantly between WT and Nucling-KO mice after the LPS challenge, hepatocytes of Nucling-KO mice were refractory to LPS- or TNF-α-induced cell death. These results were consistent with the decreased expression of proapoptotic proteins including apoptosis-inducing factor and cleaved form of poly (ADP-ribose) polymerase and terminal deoxynucleotidyl transferase dUTP nick end-labelling positive cells in the liver of Nucling-KO mice after the administration of a lethal dose of LPS. Moreover, the upregulation of NF-κB-regulated anti-apoptotic molecules including cellular inhibitor of apoptosis (cIAP) 1 and cIAP2 was observed in the liver of Nucling-KO mice after LPS treatment. These findings indicate that the Nucling deficiency leads to resistance to apoptosis in liver. We propose that Nucling is important for the induction of apoptosis in cells damaged by cytotoxic stressors through the NF-κB signalling pathway.

Nucling is identified as a novel regulator of apoptosis. In this study, we investigated the nuclear factor-κB (NF-κB)-dependent mechanism of Nucling expression, as well as the degradation pathways mediated by proteasome system and caspase activation. Using chromatin immunoprecipitation assay in wild-type mouse embryonic fibroblasts (WT MEFs), we found that NF-κB p65 could bind to the κB motifs in the promoter regions of Nucling gene at four putative-binding sites. By real-time PCR and immunoblot, we confirmed that Nucling expression was up-regulated by tumour necrosis factor-α (TNF-α) stimulation in WT MEFs, but not in NF-κB p50 knockout MEFs. On the other hand, we investigated the degradation of Nucling in connection with proteasome and caspase by using cycloheximide chase. The results showed that Nucling is a short-lived protein, and its degradation was recovered by proteasome inhibitor MG132. Moreover, under TNF-α stimulation, degradation of Nucling was recovered by pan-caspase inhibitor zVAD-fmk. Taken together, we propose a mechanism of Nucling intracellular metabolism. Nucling expression is induced by canonical NF-κB signalling pathway, whereas Nucling is undergoing proteasome degradation, as well as being cleaved by caspase system under stress conditions. This opens a new perspective for studying the NF-κB dependent regulation mechanism of cell death and survival.

Mothers and family members of young female students play important roles for guiding their self-care strategies for menstruation-related symptoms; which often affect their daily life and academic life. The aim of this study is to clarify the differences in menstruation-related symptoms before and during menstruation in university students living alone and university students living with their family in Japan. We conducted a cross-sectional online survey to assess menstruation-related symptoms before and during menstruation using the menstrual distress questionnaire (MDQ). Among 135 students; the proportion of students living alone was 60.7% and the proportion of students living with their family was 39.3%. Before menstruation; the MDQ total score and the scores for negative affect and behavior change were significantly higher in students living alone than in students living with their family. During menstruation; scores for negative affect and impaired concentration were also significantly higher in students living alone. In addition; before menstruation; scores for an increase in appetite and craving for sweets were significantly higher in students living alone. Thus; living alone affected the psychological aspects of menstruation-related symptoms in young women. The results suggest that university students who live alone should be aware of the importance of talking about their menstruation problems with family members and seeking their advice

Menstruation-related symptoms in young women are associated with lifestyle factors such as stress. This study aimed to examine menstrual conditions, menstruation-related symptoms, and types of stress in Japanese female students from their first to second years of university and the association between menstruation-related symptoms and stress levels. This prospective longitudinal cohort study utilized a self-reported web survey with the Menstrual Distress Questionnaire (MDQ) at three timepoints (2, 8, and 14 months) following admission and continuing into the second year. Data from 33 out of 102 students, who were followed throughout the study period, were used for analysis. Results indicated a trend towards changes in menstrual regularity, with the lowest (48.5%) at timepoint 1 and the highest (72.7%) at timepoint 3, and significant changes in perceived bleeding amount. Stress related to study and academic performance was consistently highest, with significant differences across timepoints. Intra-menstrual MDQ scores were highest at timepoint 1, with significant variations in impaired concentration severity across timepoints. Significant correlations were found between the MDQ scores and stress related to health and personality at all timepoints, and to future prospects, friendships, study, and academic performance at different timepoints. In short, stress related to academic performance, along with a strong correlation between MDQ scores and specific stress types, such as health and personality, was observed across all timepoints. This implies the need to provide new students with health education regarding menstruation and stress management before specific timepoints.