Epstein-Barr Virus Infection in Chronically Inflamed Periapical Granulomas

Periapical granulomas are lesions around the apex of a tooth caused by a polymicrobial infection. Treatment with antibacterial agents is normally performed to eliminate bacteria from root canals; however, loss of the supporting alveolar bone is typically observed, and tooth extraction is often selected if root canal treatment does not work well. Therefore, bacteria and other microorganisms could be involved in this disease. To understand the pathogenesis of periapical granulomas more precisely, we focused on the association with Epstein-Barr virus (EBV) using surgically removed periapical granulomas (n = 32). EBV DNA was detected in 25 of 32 periapical granulomas (78.1%) by real-time PCR, and the median number of EBV DNA copies was approximately 8,688.01/μg total DNA. In contrast, EBV DNA was not detected in healthy gingival tissues (n = 10); the difference was statistically significant according to the Mann-Whitney U test (p = 0.0001). Paraffin sections were also analyzed by in situ hybridization to detect EBV-encoded small RNA (EBER)-expressing cells. EBER was detected in the cytoplasm and nuclei of B cells and plasma cells in six of nine periapical granulomas, but not in healthy gingival tissues. In addition, immunohistochemical analysis for latent membrane protein 1 (LMP-1) of EBV using serial tissue sections showed that LMP-1-expressing cells were localized to the same areas as EBER-expressing cells. These data suggest that B cells and plasma cells in inflamed granulomas are a major source of EBV infection, and that EBV could play a pivotal role in controlling immune cell responses in periapical granulomas.     

Schisandrin B Ameliorates ICV-Infused Amyloid β Induced Oxidative Stress and Neuronal Dysfunction through Inhibiting RAGE/NF-κB/MAPK and Up-Regulating HSP/Beclin Expression

Amyloid β (Aβ)-induced neurotoxicity is a major pathological mechanism of Alzheimer’s disease (AD). Our previous studies have demonstrated that schisandrin B (Sch B), an antioxidant lignan from Schisandra chinensis, could protect mouse brain against scopolamine- and cisplatin-induced neuronal dysfunction. In the present study, we examined the protective effect of Sch B against intracerebroventricular (ICV)-infused Aβ-induced neuronal dysfunction in rat cortex and explored the potential mechanism of its action. Our results showed that 26 days co-administration of Sch B significantly improved the behavioral performance of Aβ (1–40)-infused rats in step-through test. At the same time, Sch B attenuated Aβ-induced increases in oxidative and nitrosative stresses, inflammatory markers such as inducible nitric oxide syntheses, cyclooxygenase-2, interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α, and DNA damage. Several proteins such as receptor for advanced glycation end products (RAGE), nuclear factor-κB, mitogen-activated protein kinases, and apoptosis markers were over expressed in Aβ-infused rats but were significantly inhibited by Sch B treatment. Furthermore, Sch B negatively modulated the Aβ level with simultaneous up-regulation of HSP70 and beclin, autophagy markers in Aβ-infused rats. The aforementioned effects of Sch B suggest its protective role against Aβ-induced neurotoxicity through intervention in the negative cycle of RAGE-mediated Aβ accumulation during AD patho-physiology.     

mTOR regulates the nucleoplasmic diffusion of Xrn2 under conditions of heat stress

Stress induces various responses, including translational suppression and tRNA degradation in mammals. Previously, we showed that heat stress induces degradation of initiator tRNA^Met (iMet) through 5′–3′ exoribonuclease Xrn1 and Xrn2, respectively. In addition, we found that rapamycin inhibits the degradation of iMet under heat stress conditions. Here, we report that the mammalian target of rapamycin (mTOR) regulates the diffusion of Xrn2 from the nucleolus to the nucleoplasm, facilitating the degradation of iMet under conditions of heat stress. Our results suggest a mechanism of translational suppression through mTOR-regulated iMet degradation in mammalian cells.     

Direct Measurement of Helical Cell Motion of the Spirochete Leptospira

Leptospira are spirochete bacteria distinguished by a short-pitch coiled body and intracellular flagella. Leptospira cells swim in liquid with an asymmetric morphology of the cell body; the anterior end has a long-pitch spiral shape (S-end) and the posterior end is hook-shaped (H-end). Although the S-end and the coiled cell body called the protoplasmic cylinder are thought to be responsible for propulsion together, most observations on the motion mechanism have remained qualitative. In this study, we analyzed the swimming speed and rotation rate of the S-end, protoplasmic cylinder, and H-end of individual Leptospira cells by one-sided dark-field microscopy. At various viscosities of media containing different concentrations of Ficoll, the rotation rate of the S-end and protoplasmic cylinder showed a clear correlation with the swimming speed, suggesting that these two helical parts play a central role in the motion of Leptospira. In contrast, the H-end rotation rate was unstable and showed much less correlation with the swimming speed. Forces produced by the rotation of the S-end and protoplasmic cylinder showed that these two helical parts contribute to propulsion at nearly equal magnitude. Torque generated by each part, also obtained from experimental motion parameters, indicated that the flagellar motor can generate torque >4000 pN nm, twice as large as that of Escherichia coli. Furthermore, the S-end torque was found to show a markedly larger fluctuation than the protoplasmic cylinder torque, suggesting that the unstable H-end rotation might be mechanically related to changes in the S-end rotation rate for torque balance of the entire cell. Variations in torque at the anterior and posterior ends of the Leptospira cell body could be transmitted from one end to the other through the cell body to coordinate the morphological transformations of the two ends for a rapid change in the swimming direction.     

Helical image reconstruction of the outward-open human erythrocyte band 3 membrane domain in tubular crystals

The C-terminal membrane domain of erythrocyte band 3 functions as an anion exchanger. Here, we report the three-dimensional (3D) structure of the membrane domain in an inhibitor-stabilized, outward-open conformation at 18 Å resolution. Unstained, frozen-hydrated tubular crystals containing the membrane domain of band 3 purified from human red blood cells (hB3MD) were examined using cryo-electron microscopy and iterative helical real-space reconstruction (IHRSR). The 3D image reconstruction of the tubular crystals showed the molecular packing of hB3MD dimers with dimensions of 60 × 110 Å in the membrane plane and a thickness of 70 Å across the membrane. Immunoelectron microscopy and carboxyl-terminal digestion demonstrated that the intracellular surface of hB3MD was exposed on the outer surface of the tubular crystal. A 3D density map revealed that hB3MD consists of at least two subdomains and that the outward-open form is characterized by a large hollow area on the extracellular surface and continuous density on the intracellular surface.     

Transplanted neural progenitor cells expressing mutant NT3 promote myelination and partial hindlimb recovery in the chronic phase after spinal cord injury

Neutrotrophin-3 (NT3) plays a protective role in injured central nervous system tissues through interaction with trk receptors. To enhance the regeneration of damaged tissue, a combination therapy with cell transplantation and neurotrophins has been under development. We examined whether the transplantation of neural progenitor cells (NPCs) secreting NT3/D15A, a multi-neurotrophin with the capacity to bind both trkB and trkC, would enhance the repair of damaged tissues and the functional recovery in a chronic phase of spinal cord injury. The cultured NPCs with lentiviral vector containing either GFP or NT3/D15A were transplanted into the contused spinal cord at 6 weeks after the initial thoracic injury. Eight weeks after the transplantation, the NT3/D15A transplants displayed better survival than the GFP transplants, and they exhibited enhanced myelin formation and partial improvement of hindlimb function. Our study revealed that NT3/D15A produced positive effects in injured spinal cords even in the chronic phase. These effects suggest an enhanced neurotrophin-trk signaling by NT3/D15A.     

Important roles of Tyr43 at the putative heme distal side in the oxygen recognition and stability of the Fe(II)-O2 complex of YddV, a globin-coupled heme-based oxygen sensor diguanylate cyclase

YddV from Escherichia coli (Ec) is a novel globin-coupled heme-based oxygen sensor protein displaying diguanylate cyclase activity in response to oxygen availability. In this study, we quantified the turnover numbers of the active [Fe(III), 0.066 min(-1); Fe(II)-O(2) and Fe(II)-CO, 0.022 min(-1)] [Fe(III), Fe(III)-protoporphyrin IX complex; Fe(II), Fe(II)-protoporphyrin IX complex] and inactive forms [Fe(II) and Fe(II)-NO, <0.01 min(-1)] of YddV for the first time. Our data indicate that the YddV reaction is the rate-determining step for two consecutive reactions coupled with phosphodiesterase Ec DOS activity on cyclic di-GMP (c-di-GMP) [turnover number of Ec DOS-Fe(II)-O(2), 61 min(-1)]. Thus, O(2) binding and the heme redox switch of YddV appear to be critical factors in the regulation of c-di-GMP homeostasis. The redox potential and autoxidation rate of heme of the isolated heme domain of YddV (YddV-heme) were determined to be -17 mV versus the standard hydrogen electrode and 0.0076 min(-1), respectively. The Fe(II) complexes of Y43A and Y43L mutant proteins (residues at the heme distal side of the isolated heme-bound globin domain of YddV) exhibited very low O(2) affinities, and thus, their Fe(II)-O(2) complexes were not detected on the spectra. The O(2) dissociation rate constant of the Y43W protein was >150 s(-1), which is significantly larger than that of the wild-type protein (22 s(-1)). The autoxidation rate constants of the Y43F and Y43W mutant proteins were 0.069 and 0.12 min(-1), respectively, which are also markedly higher than that of the wild-type protein. The resonance Raman frequencies representing ν(Fe-O(2)) (559 cm(-1)) of the Fe(II)-O(2) complex and ν(Fe-CO) (505 cm(-1)) of the Fe(II)-CO complex of Y43F differed from those (ν(Fe-O(2)), 565 cm(-1); ν(Fe-CO), 495 cm(-1)) of the wild-type protein, suggesting that Tyr43 forms hydrogen bonds with both O(2) and CO molecules. On the basis of the results, we suggest that Tyr43 located at the heme distal side is important for the O(2) recognition and stability of the Fe(II)-O(2) complex, because the hydroxyl group of the residue appears to interact electrostatically with the O(2) molecule bound to the Fe(II) complex in YddV. Our findings clearly support a role of Tyr in oxygen sensing, and thus modulation of overall conversion from GTP to pGpG via c-di-GMP catalyzed by YddV and Ec DOS, which may be applicable to other globin-coupled oxygen sensor enzymes.     

Influence of predator‐specific defense adaptation on intraguild predation

The persistence of intraguild predation (IGP), the prey–predator interaction between competing species, is puzzling because simple IGP models readily predict species extinction. In this study, we explored a mathematical model incorporating predator‐specific defense adaptation of basal prey against intraguild prey and intraguild predator. The model explicitly described the dynamics of the defense effort against each predator under the assumption that anti‐predator defense was associated with reducing effort allocated to reproduction. The model predicted that defense adaptation (i.e. the ability to reallocate defense effort) would facilitate coexistence, particularly when system productivity is high; at low productivity, coexistence would be facilitated or inhibited depending on initial effort allocation prior to defense adaptation. In addition, we found that three‐species dynamics became more stable at higher adaptation rates. The results suggest that common behavioral changes, such as predator‐specific defense adaptation, have significant implications for the community structure and dynamics of IGP systems.