The presence of micronuclei in mammalian cells is related to several mutagenetic stresses. In order to understand how micronuclei emerge, behave in cells, and affect cell fate, we performed extensive time-lapse microscopy of HeLa H2B-GFP cells in the presence of hydroxyurea at low concentration. Micronuclei formed after mitosis from lagging chromatids or chromatin bridges between anaphase chromosomes and were stably maintained in the cells for up to one cell cycle. Nuclear buds also formed from chromatin bridges or during interphase. If the micronuclei-bearing cells entered mitosis, they either produced daughter cells without micronuclei or, more frequently, produced cells with additional micronuclei. Low concentrations of hydroxyurea efficiently induced multipolar mitosis, which generated lagging chromatids or chromatin bridges, and also generated multinuclear cells that were tightly linked to apoptosis. We found that the presence of micronuclei is related to apoptosis but not to multipolar mitosis. Furthermore, the structural heterogeneity among micronuclei, with respect to chromatin condensation or the presence of lamin B, derived from the mechanism of micronuclei formation. Our study reinforces the notion that micronucleation has important implications in the genomic plasticity of tumor cells.
BackgroundAdenosine A2b receptor (ADORA2B) encodes an adenosine receptor that is a member of the G protein-coupled receptor superfamily. This integral membrane protein stimulates adenylate cyclase activity in the presence of adenosine. Little is known about the relevance of ADORA2B to human malignancy including oral squamous cell carcinoma (OSCC). We aimed to characterize the expression state and function of ADORA2B in OSCC.MethodsThe ADORA2B expression levels in nine OSCC-derived cells were analyzed by quantitative reverse transcriptase-polymerase chain reaction and immunoblotting analyses. Using an ADORA2B knockdown model, we assessed cellular proliferation and expression of hypoxia-inducible factor1α (HIF-1α). We examined the adenosine receptor expression profile under both normoxic and hypoxic conditions in the OSCC-derived cells. In addition to in vitro data, the clinical correlation between the ADORA2B expression levels in primary OSCCs (n = 100 patients) and the clinicopathological status by immunohistochemistry (IHC) also was evaluated.ResultsADORA2B mRNA and protein were up-regulated significantly (p < 0.05) in seven OSCC-derived cells compared with human normal oral keratinocytes. Suppression of ADORA2B expression with shRNA significantly (p < 0.05) inhibited cellular proliferation compared with the control cells. HIF-1α also was down-regulated in ADORA2B knockdown OSCC cells. During hypoxia, ADORA2B expression was induced significantly (p < 0.05) in the mRNA and protein after 24 hours of incubation in OSCC-derived cells. IHC showed that ADORA2B expression in primary OSCCs was significantly (p < 0.05) greater than in the normal oral counterparts and that ADORA2B-positive OSCCs were correlated closely (p < 0.05) with tumoral size.ConclusionOur results suggested that ADORA2B controls cellular proliferation via HIF-1α activation, indicating that ADORA2B may be a key regulator of tumoral progression in OSCCs.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-015-1577-2) contains supplementary material, which is available to authorized users.
Joint angle kinematics of the throwing limb from the early-cocking phase to ball release were investigated for the fastball (FB) and curveball (CB) baseball pitches through use of a three-dimensional film analysis technique. Small sticks were fixed to the hand and forearm to permit rotations of the radioulnar and wrist joint to be calculated. The actions were very similar for two pitches within one subject. There were no differences in the motions of the shoulder and elbow joints or in the temporal sequences between FB and CB pitches. However, there was a significant difference (p<0.05) in the motion of supination/pronation of the forearm and dorsiflexion/palmar flexion of the wrist prior to the ball release; the forearm was supinated more in the CB (maximum supination: 39.9 ±6.0° at 0.072 ±0.045 s before ball release, or BRL) compared to the FB (19.4 ±8.5° at 0.076 ±0.046 s before BRL), whereas the wrist was dorsiflexed more in the FB (maximum dorsiflexion: 41.7 ±6.5° at 0.039 ±0.011 s before BRL) compared to the CB (31.2 ±4.7° at 0.036 ±0.013 s before BRL) during late-cocking and acceleration phases leading to the ball release.
Treatment resistance in CSCs seems to be regulated by various mechanisms, and, therefore, additional treatment strategies to target CSCs are required in patients with HNSCC.
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