Kir2.1に新規ミスセンス変異（L94P）を認める常染色体劣性遺伝形式のAndersen-Tawil症候群

Aim: Dominant negative mutations of the inwardly rectifying K+ channel Kir2.1 cause Andersen-Tawil syndrome, an autosomal dominant disorder. Here, we identified a novel Kir2.1 mutation causing autosomal recessive ATS, and explored the underlying mechanism. Methods: We sequenced the coding region of KCJN2. We assessed protein subcellular localization by transfecting cells with Kir2.1-enhanced green fluorescent protein fusions and observing them by confocal microscopy. We measured K+ currents using patch clamping. Results: We identified the novel Kir2.1 missense mutation L94P. L94P-EGFP was barely detected at the plasma membrane, in contrast to WT-EGFP and L94PEGFP+WT. The excitability of L94P-expressing cells was decreased compared with that of WT-expressing cells and L94P+WT-expressing cells (p < 0.001). Conclusions: Most L94P mutant Kir2.1 fails to reach the plasma membrane, but heterotetrameric channels comprising L94P+WT can traffic normally to the plasma membrane and generate currents. The L94P mutation is transmitted as an autosomal recessive trait.広島大学(Hiroshima University)博士(医学)Philosophy in Medical Sciencedoctora

Activity-dependent oligodendrocyte calcium dynamics and their changes in Alzheimer’s disease

Oligodendrocytes (OCs) form myelin around axons, which is dependent on neuronal activity. This activity-dependent myelination plays a crucial role in training and learning. Previous studies have suggested that neuronal activity regulates proliferation and differentiation of oligodendrocyte precursor cells (OPCs) and myelination. In addition, deficient activity-dependent myelination results in impaired motor learning. However, the functional response of OC responsible for neuronal activity and their pathological changes is not fully elucidated. In this research, we aimed to understand the activity-dependent OC responses and their different properties by observing OCs using in vivo two-photon microscopy. We clarified that the Ca2+ activity in OCs is neuronal activity dependent and differentially regulated by neurotransmitters such as glutamate or adenosine triphosphate (ATP). Furthermore, in 5-month-old mice models of Alzheimer’s disease, a period before the appearance of behavioral abnormalities, the elevated Ca2+ responses in OCs are ATP dependent, suggesting that OCs receive ATP from damaged tissue. We anticipate that our research will help in determining the correct therapeutic strategy for neurodegenerative diseases beyond the synapse

Hydrolyzed collagen intake increases bone mass of growing rats trained with running exercise

BackgroundSome studies have shown that dietary hydrolyzed collagen peptides (HC) effectively prevent age-related bone loss. However, it is not known whether the intake of HC also has positive effect on bone mass or strength when combined with exercise during growth phase.MethodsWe examined the effects of 11 weeks of HC intake and running exercise on bone mass and strength in growing rats. Rats were randomized into four groups, the 20% casein group (Casein20), the 40% casein group (Casein40), the 20% HC group (HC20), and the 40% HC group (HC40). Each group was further divided into exercise groups (Casein20 + Ex, Casein40 + Ex, HC20 + Ex, HC40 + Ex) and non-exercise group (Casein20, Casein40, HC20, HC40). In the HC intake groups, 30% of casein protein was replaced with HC. Exercise group rats were trained 6 days per week on a treadmill (25–30 m/min, 60 min) for 60 days. After being sacrificed, their bone mineral content (BMC) and bone strength were evaluated.ResultsExercise and dietary HC effects were observed in the adjusted BMC of lumbar spine and tibia among the 20% protein groups (p < 0.001 for exercise; p < 0.05 for dietary HC, respectively). These effects were also noted in the adjusted wet weight and dry weight of femur among the 20% protein groups (p < 0.001, p < 0.01 for exercise; p < 0.01, p < 0.001 for dietary HC, respectively). On the other hand, in adjusted bone breaking force and energy, dietary HC effect was not significant. Among the 40% protein groups, similar results were obtained in the adjusted BMC, femoral weight, bone breaking force, and energy. There were no differences between the 20% protein groups and the 40% protein groups.ConclusionsThe present study demonstrated that moderate HC intake (where the diet contains 20% protein, of which 30% is HC) increased bone mass during growth period and further promoted the effect of running exercise. On the other hand, a higher HC intake (where the diet contains 40% protein, of which 30% is HC) had no more beneficial effect on bone mass than the moderate HC intake

Functional Expression of Heme Oxygenase-1 in Human Differentiated Epidermis and Its Regulation by Cytokines

Although heme oxygenase-1 (HO-1) is induced in keratinocytes after UV radiation, HO-1 expression during normal epidermal differentiation has not yet been reported. We showed by real-time PCR, western blotting, and ELISA that HO-1 mRNA and protein expression by cultured normal human keratinocytes was upregulated during epidermal differentiation induced by a high-calcium medium. Immunohistochemical staining and in situ hybridization showed the graduated expression of HO-1 in the upper epidermis, which was accompanied by suprabasal HO-1 mRNA expression, and the accumulation of bilirubin (BR) in the stratum corneum. We examined the activation of nuclear factor E2-related factor 2 (Nrf2), which is a pivotal transcription factor for HO-1 expression, by western blotting and by examining the mRNA expression of Nrf2 target genes, and excluded its role in HO-1 expression in epidermal differentiation. Next, we examined the regulation of HO-1 expression by inflammatory cytokines. IL-4 and IL-22 significantly reduced HO-1 mRNA and protein expression, whereas IL-1β, IL-17A, and tumor necrosis factor-α (TNF-α) increased it. Finally, immunohistochemical studies on psoriatic lesional skin showed that HO-1 expression was downregulated in the parakeratotic epidermis, whereas it was retained in the orthokeratotic epidermis. These studies demonstrate that HO-1 is functionally expressed by keratinocytes in parallel with epidermal differentiation and that its expression is independently affected by several cytokines

Structures and mechanisms of actin ATP hydrolysis

The major cytoskeleton protein actin undergoes cyclic transitions between the monomeric G-form and the filamentous F-form, which drive organelle transport and cell motility. This mechanical work is driven by the ATPase activity at the catalytic site in the F-form. For deeper understanding of the actin cellular functions, the reaction mechanism must be elucidated. Here, we show that a single actin molecule is trapped in the F-form by fragmin domain-1 binding and present their crystal structures in the ATP analog-, ADP-Pi-, and ADP-bound forms, at 1.15-Å resolutions. The G-to-F conformational transition shifts the side chains of Gln137 and His161, which relocate four water molecules including W1 (attacking water) and W2 (helping water) to facilitate the hydrolysis. By applying quantum mechanics/molecular mechanics calculations to the structures, we have revealed a consistent and comprehensive reaction path of ATP hydrolysis by the F-form actin. The reaction path consists of four steps: 1) W1 and W2 rotations; 2) PG–O3B bond cleavage; 3) four concomitant events: W1–PO3− formation, OH− and proton cleavage, nucleophilic attack by the OH− against PG, and the abstracted proton transfer; and 4) proton relocation that stabilizes the ADP-Pi–bound F-form actin. The mechanism explains the slow rate of ATP hydrolysis by actin and the irreversibility of the hydrolysis reaction. While the catalytic strategy of actin ATP hydrolysis is essentially the same as those of motor proteins like myosin, the process after the hydrolysis is distinct and discussed in terms of Pi release, F-form destabilization, and global conformational changes

The Confederate Stories of America: The Short-Story Cycle and the Representation of the American South

My dissertation examines the ways in which the short-story cycle has provided a unique generic framework for representing and investigating the complex interplay of contending forces that constitute what we think of as the American South. Often confused with a collection of disparate short stories or a novel, the short-story cycle is a collection of short stories in which each story is independent, but simultaneously interrelated to one another. Although the South has produced a number of short-story cycles or linked story collections, scholars have not paid much attention to the connection between the genre/form and the region. I consider, however, the genre/form has been an apt medium for writers to represent Southern society, which is a mosaic of diverse races and values, but certain actors and groups within it have attempted to imagine it and present it as having some sort of fundamental unity. The most substantial scholarship on the genre agrees that some of the precursors of the American short-story cycle are a collection of regional sketches written in the nineteenth century. In order to clarify the important role that the short-story cycle has played in the history of the South and its literary history, I examine not only a short-story cycle proper such as William Faulkner’s Go Down, Moses (1942) and Eudora Welty’s The Golden Apples (1949), but also the works that can be regarded as pioneers of the southern short-story cycle, such as George Washington Cable’s The Grandissimes (1880) and Charles Chesnutt’s The Conjure Woman (1899). I also analyze Jean Toomer’s Cane (1923) and Julia Peterkin’s Green Thursday (1924) not as a representative of literary modernism, for which scholars have considered the genre has a strong affinity, as an intersection of precursors of the cycle written in the nineteenth-century South and that written in the twentieth-century South. My dissertation explores how southern writers have been shaping the genre, adapting it to their own use, and redefining its limits so as to go beyond the idea and representation of “the South.