A high-resolution phase-contrast microscopy system for label-free imaging in living cells.

Cell biologists have long sought the ability to observe intracellular structures in living cells without labels. This study presents procedures to adjust a commercially available apodized phase-contrast (APC) microscopy system for better visualizing the dynamic behaviors of various subcellular organelles in living cells. By harnessing the versatility of this technique to capture sequential images, we could observe morphological changes in cellular geometry after virus infection in real time without probes or invasive staining. The tune-up APC microscopy system is a highly efficient platform for simultaneously observing the dynamic behaviors of diverse subcellular structures with exceptional resolution.

Deep learning-based segmentation of subcellular organelles in high-resolution phase-contrast images.

Although quantitative analysis of biological images demands precise extraction of specific organelles or cells, it remains challenging in broad-field grayscale images, where traditional thresholding methods have been hampered due to complex image features. Nevertheless, rapidly growing artificial intelligence technology is overcoming obstacles. We previously reported the fine-tuned apodized phase-contrast microscopy system to capture high-resolution, label-free images of organelle dynamics in unstained living cells (Shimasaki, K. et al. (2024). Cell Struct. Funct., 49:21-29). We here showed machine learning-based segmentation models for subcellular targeted objects in phase-contrast images using fluorescent markers as origins of ground truth masks. This method enables accurate segmentation of organelles in high-resolution phase-contrast images, providing a practical framework for studying cellular dynamics in unstained living cells.Key words: Label-free imaging, Organelle dynamics, Apodized phase contrast, Deep learning-based segmentation.

Validation of new medication use algorithms as proxies for worsening disease activity in patients with juvenile idiopathic arthritis.

PURPOSE: To facilitate claims-based research on populations with juvenile idiopathic arthritis (JIA), we sought to validate an algorithm of new medication use as a proxy for worsening JIA disease activity. METHODS: Using electronic health record data from three pediatric centers, we defined new JIA medication use as (re)initiation of disease-modifying antirheumatic drugs or glucocorticoids (oral or intra-articular). Data were collected from 201 randomly selected subjects with (101) or without (100) new medication use. We assessed the positive predictive value (PPV) and negative predictive value (NPV) based on a reference standard of documented worsening of JIA disease activity. The algorithm was refined to optimize test characteristics. RESULTS: Overall, the medication-based algorithm had suboptimal performance in representing worsening JIA disease activity (PPV 69.3%, NPV 77.1%). However, algorithm performance improved for definitions specifying longer times after JIA diagnosis (≥1-year post-diagnosis: PPV 82.9%, NPV 80.0%) or after initiation of prior JIA treatment (≥1-year post-treatment: PPV 89.7%, NPV 80.0%). CONCLUSION: An algorithm for new JIA medication use appears to be a reasonable proxy for worsening JIA disease activity, particularly when specifying new use ≥1 year since initiating a prior JIA medication. This algorithm will be valuable for conducting research on JIA populations within administrative claims databases.

Laundry detergents and surfactants-induced eosinophilic airway inflammation by increasing IL-33 expression and activating ILC2s.

INTRODUCTION: Epidemiological studies demonstrated that cleaning work and frequent use of cleaning products are risk factors for asthma. Laundry detergents have been reported to have epithelial barrier-opening effects. However, whether laundry detergents directly induce airway inflammation and its mechanisms in vivo remain to be elucidated. METHODS: Two commercial laundry detergents and two commonly used surfactants for cleaning and cosmetics (sodium lauryl sulfate and sodium dodecyl benzene sulfonate) were intranasally administered to mice. Lungs were analyzed using flow cytometry, histology, ELISA, and quantitative PCR. Human bronchial epithelial cells were stimulated with laundry detergents and analyzed using quantitative PCR and western blotting. Involvement of oxidative stress was assessed using an antioxidant. Dust samples from homes were analyzed to determine their detergent content by measuring their critical micelle concentration (CMC). RESULTS: The administered laundry detergents and surfactants-induced eosinophilic airway inflammation accompanied by increased IL-33 expression and activation of group 2 innate lymphoid cells (ILC2s). Detergent-induced eosinophilic airway inflammation was significantly attenuated in Rag2-/- Il2rg-/- , Il33-/- mice, and also in wild-type mice treated with NAC. Detergent-induced IL-33 expression in airways was attenuated by NAC treatment, both in vivo and in vitro. CMCs were found in all of the tested dust extracts, and they differed significantly among the homes. CONCLUSION: The laundry detergents and surfactants-induced eosinophilic airway inflammation in vivo through epithelial cell and ILC2 activation. They induced IL-33 expression in airway epithelial cells through oxidative stress. Furthermore, detergent residues were present in house dust and are presumably inhaled into the airway in daily life.

Direct platelet adhesion potentiates group 2 innate lymphoid cell functions.

BACKGROUND: Platelets are thought to be involved in the pathophysiology of asthma, presumably through direct adhesion to inflammatory cells, including group 2 innate lymphoid cells (ILC2s). Here, we tried to elucidate the effects of platelet adhesion to ILC2s in vitro and in vivo, as well as the mechanisms involved. METHODS: Alternaria-induced ILC2-dependent airway inflammation models using wild-type and c-mpl-/- mice were evaluated. Both purified CD41+ and CD41- ILC2s were cultured with IL-2 and IL-33 to determine in vitro Type 2 (T2) cytokine production and cell proliferation. RNA-seq data of flow-cytometry-sorted CD41+ and CD41- ILC2s were used to isolate ILC2-specific genes. Flow cytometry was performed to determine the expression of CD41 and adhesion-related molecules on ILC2s in both mouse and human tissues. RESULTS: T2 inflammation and T2 cytokine production from ILC2s were significantly reduced in the c-mpl-/- mice compared to wild-type mice. Platelet-adherent ILC2s underwent significant proliferation and showed enhanced T2 cytokine production when exposed to IL-2 and IL-33. The functions of ILC2-specific genes were related to cell development and function. Upstream regulator analysis identified 15 molecules, that are thought to be involved in ILC2 activation. CD41 expression levels were higher in ILC2s from human PBMCs and mouse lung than in those from secondary lymphoid tissues, but they did not correlate with the P-selectin glycoprotein ligand-1 or CD24 expression level. CONCLUSION: Platelets spontaneously adhere to ILC2s, probably in the peripheral blood and airways, thereby potentiating ILC2s to enhance their responses to IL-33.

Association of home blood pressure with sleep and physical and mental activity, assessed via a wristwatch-type pulsimeter with accelerometer in adults.

This study aimed to examine the associations between home blood pressure (HBP) and sleep and activity assessed using data obtained via a wristwatch-type pulsimeter with accelerometer (Pulsense®) using original software. We recruited 28 elderlies and 40 employees aged 24-81 years who were not on hypotensive agents and sleeping drugs. Sleep, activity, and HBP were measured consecutively over a 5-7-day period. Body mass index (BMI), base heart rate (HR0), and age showed significant correlation with HBP in a simple and multiple linear regression analysis. HR0 was positively, and log deep sleep duration, negatively correlated with HBP in the adjusted multiple linear regression analysis. Physical and mental activities were negatively correlated with systolic blood pressure (SBP) in a simple linear regression, but high physical and mental activities tend to reduce deep sleep duration. Self-recorded sleep duration had no relationship with HBP. In conclusion, HR0, BMI, age, deep sleep duration, and activity showed relationships with HBP. Using this type of wristwatch and observing daily sleep and activity data with HBP measurement may have important clinical implication.

Induction of human regulatory innate lymphoid cells from group 2 innate lymphoid cells by retinoic acid.

BACKGROUND: Group 2 innate lymphoid cells (ILC2s) play critical roles in induction and exacerbation of allergic airway inflammation. Thus clarification of the mechanisms that underlie regulation of ILC2 activation has received significant attention. Although innate lymphoid cells are divided into 3 major subsets that mirror helper effector T-cell subsets, counterpart subsets of regulatory T cells have not been well characterized. OBJECTIVE: We sought to determine the factors that induce regulatory innate lymphoid cells (ILCregs). METHODS: IL-10+ ILCregs induced from ILC2s by using retinoic acid (RA) were analyzed with RNA-sequencing and flow cytometry. ILCregs were evaluated in human nasal tissue from healthy subjects and patients with chronic rhinosinusitis with nasal polyps and lung tissue from house dust mite- or saline-treated mice. RESULTS: RA induced IL-10 secretion by human ILC2s but not type 2 cytokines. IL-10+ ILCregs, which were converted from ILC2s by means of RA stimulation, expressed a regulatory T cell-like signature with expression of IL-10, cytotoxic T lymphocyte-associated protein 4, and CD25, with downregulated effector type 2-related markers, such as chemoattractant receptor-homologous molecule on TH2 cells and ST2, and suppressed activation of CD4+ T cells and ILC2s. ILCregs were rarely detected in human nasal tissue from healthy subjects or lung tissue from saline-treated mice, but numbers were increased in nasal tissue from patients with chronic rhinosinusitis with nasal polyps and in lung tissue from house dust mite-treated mice. Enzymes for RA synthesis were upregulated in airway epithelial cells during type 2 inflammation in vivo and by IL-13 in vitro. CONCLUSION: We have identified a unique immune regulatory and anti-inflammatory pathway by which RA converts ILC2s to ILCregs. Interactions between airway epithelial cells and ILC2s play an important roles in the generation of ILCregs.

Both Sphingomyelin and Cholesterol in the Host Cell Membrane Are Essential for Rubella Virus Entry.

Rubella virus (RuV) causes a systemic infection, and transplacental fetal infection causes congenital rubella syndrome. In this study, we showed that treatment of cells with sphingomyelinase inhibited RuV infection. Assays using inhibitors of serine palmitoyl transferase and ceramide transport protein demonstrated the contribution of sphingomyelin (SM) to RuV infection. Compelling evidence for direct binding of RuV to lipid membranes at neutral pH was obtained using liposome coflotation assays. The absence of either SM or cholesterol (Chol) abrogated the RuV-liposome interaction. SM and Chol (SM/Chol) were also critical for RuV binding to erythrocytes and lymphoid cells. Removal of Ca2+ from the assay buffer or mutation of RuV envelope E1 protein Ca2+-binding sites abrogated RuV binding to liposomes, erythrocytes, and lymphoid cells. However, RuV bound to various nonlymphoid adherent cell lines independently of extracellular Ca2+ or SM/Chol. Even in these adherent cell lines, both the E1 protein Ca2+-binding sites and cellular SM/Chol were essential for the early stage of RuV infection, possibly affecting envelope-membrane fusion in acidic compartments. Myelin oligodendrocyte glycoprotein (MOG) has recently been identified as a cellular receptor for RuV. However, RuV bound to MOG-negative cells in a Ca2+-independent manner. Collectively, our data demonstrate that RuV has two distinct binding mechanisms: one is Ca2+ dependent and the other is Ca2+ independent. Ca2+-dependent binding observed in lymphoid cells occurs by the direct interaction between E1 protein fusion loops and SM/Chol-enriched membranes. Clarification of the mechanism of Ca2+-independent RuV binding is an important next step in understanding the pathology of RuV infection.IMPORTANCE Rubella has a significant impact on public health as infection during early pregnancy can result in babies being born with congenital rubella syndrome. Even though effective rubella vaccines are available, rubella outbreaks still occur in many countries. We studied the entry mechanism of rubella virus (RuV) and found that RuV binds directly to the host plasma membrane in the presence of Ca2+ at neutral pH. This Ca2+-dependent binding is specifically directed to membranes enriched in sphingomyelin and cholesterol and is critical for RuV infection. Importantly, RuV also binds to many cell lines in a Ca2+-independent manner. An unidentified RuV receptor(s) is involved in this Ca2+-independent binding. We believe that the data presented here may aid the development of the first anti-RuV drug.

[Validation Study of Rapid Analytical Method for Determination of Cyclamate in Various Kinds of Foods by LC-MS/MS].

A rapid, sensitive and selective analytical method by LC-MS/MS was developed and validated for the determination of cyclamate in various kinds of foods. The Preparation of test solutions was performed by heat extraction technique in accordance with an official notification method in Japan. We aimed to reduce the matrix effects in LC-MS/MS only by diluting extracts without clean-up using solid phase column. This method was assessed for 30 kinds of foods fortifying cyclamate at the concentration level of 0.5 µg/g. As a result, trueness was 85.0 to 106.6%, repeatability (RSDr) ranged from 1.7 to 9.4%, and within-laboratory reproducibility (RSDwr) ranged from was 4.1 to 9.7%. These date supported the reliability our method. Thus, this method could be useful for a rapid determination of cyclamate in various kinds of foods.

Dual effects of the Nrf2 inhibitor for inhibition of hepatitis C virus and hepatic cancer cells.

BACKGROUND: We previously showed that knockdown of nuclear factor E2-related factor 2 (Nrf2) resulted in suppression of hepatitis C virus (HCV) infection. In this study, whether brusatol, an Nrf2 inhibitor, has dual anti-HCV and anticancer effects was explored. METHODS: The anti-HCV effect of brusatol was investigated by analyzing HCV RNA and proteins in a hepatic cell line persistently-infected with HCV, HPI cells, and by analyzing HCV replication in a replicon-replicating hepatic cell line, OR6 cells. Then, dual anti-HCV and anticancer effects of brusatol and enhancement of the effects by the combination of brusatol with anticancer drugs including sorafenib, which has been reported to have the dual effects, were then investigated. RESULTS: Brusatol suppressed the persistent HCV infection at both the RNA and protein levels in association with a reduction in Nrf2 protein in the HPI cells. Analysis of the OR6 cells treated with brusatol indicated that brusatol inhibited HCV persistence by inhibiting HCV replication. Combination of brusatol with an anticancer drug not only enhanced the anticancer effect but also, in the case of the combination with sorafenib, strongly suppressed HCV infection. CONCLUSIONS: Brusatol has dual anti-HCV and anticancer effects and can enhance the comparable effects of sorafenib. There is therefore the potential for combination therapy of brusatol and sorafenib for HCV-related hepatocellular carcinoma.

Targeting cellular squalene synthase, an enzyme essential for cholesterol biosynthesis, is a potential antiviral strategy against hepatitis C virus.

UNLABELLED: Hepatitis C virus (HCV) exploits host membrane cholesterol and its metabolism for progeny virus production. Here, we examined the impact of targeting cellular squalene synthase (SQS), the first committed enzyme for cholesterol biosynthesis, on HCV production. By using the HCV JFH-1 strain and human hepatoma Huh-7.5.1-derived cells, we found that the SQS inhibitors YM-53601 and zaragozic acid A decreased viral RNA, protein, and progeny production in HCV-infected cells without affecting cell viability. Similarly, small interfering RNA (siRNA)-mediated knockdown of SQS led to significantly reduced HCV production, confirming the enzyme as an antiviral target. A metabolic labeling study demonstrated that YM-53601 suppressed the biosynthesis of cholesterol and cholesteryl esters at antiviral concentrations. Unlike YM-53601, the cholesterol esterification inhibitor Sandoz 58-035 did not exhibit an antiviral effect, suggesting that biosynthesis of cholesterol is more important than that of cholesteryl esters for HCV production. YM-53601 inhibited transient replication of a JFH-1 subgenomic replicon and entry of JFH-1 pseudoparticles, suggesting that at least suppression of viral RNA replication and entry contributes to the antiviral effect of the drug. Collectively, our findings highlight the importance of the cholesterol biosynthetic pathway in HCV production and implicate SQS as a potential target for antiviral strategies against HCV. IMPORTANCE: Hepatitis C virus (HCV) is known to be closely associated with host cholesterol and its metabolism throughout the viral life cycle. However, the impact of targeting cholesterol biosynthetic enzymes on HCV production is not fully understood. We found that squalene synthase, the first committed enzyme for cholesterol biosynthesis, is important for HCV production, and we propose this enzyme as a potential anti-HCV target. We provide evidence that synthesis of free cholesterol is more important than that of esterified cholesterol for HCV production, highlighting a marked free cholesterol dependency of HCV production. Our findings also offer a new insight into a role of the intracellular cholesterol pool that is coupled to its biosynthesis in the HCV life cycle.

Association between self-reported urinary incontinence and musculoskeletal conditions in community-dwelling elderly women: a cross-sectional study.

AIMS: Urinary incontinence (UI) and musculoskeletal conditions, particularly low back pain (LBP), and osteoarthritis (OA), are common problems that have been associated with mobility limitations and future dependence in activities of daily living in the elderly. The purpose of this study was to explore the relationship between UI, UI types, and musculoskeletal conditions in elderly community-dwelling women. METHODS: A cross-sectional study was performed on 1,399 community-dwelling Japanese women aged 75-84 years. Face-to-face interviews, body composition, and physical function, including grip strength, and usual walking speed, were conducted. UI was defined as experience of urine leakage episodes more than once per week. Self-reported presence and degree of pain, LBP, and OA were assessed. Student's t-tests and chi-square tests were used to analyze continuous and categorical variables. Associations between selected variables, UI, and UI types were assessed using stepwise multiple logistic regression models. RESULTS: A total of 260 participants had UI (18.6%) and 399 had LBP (28.5%). Participants with UI were more likely to experience pain (76.0%) and LBP (36.2%) than those without UI (P < 0.001 and P = 0.002, respectively). Age, body mass index, falls, walking speed, grip strength, LBP, and pain coupled with OA were significantly associated with UI. There were significant associations between urge UI and mild (odds ratio (OR) = 1.653, 95% confidence interval (CI) = 1.031-2.650) and severe LBP (OR = 2.617, 95% CI = 1.193-5.739). CONCLUSIONS: This study showed that UI was significantly associated with musculoskeletal conditions, including LBP, and the combination of pain and OA. The risk of urge UI was greater with increasing severity of LBP.

Differential expression of the alternatively spliced forms of prosaposin mRNAs in rat choroid plexus.

Prosaposin has two distinct profiles. One is a precursor form that is processed into saposins thus promoting lysosomal sphingolipid hydrolase function, whereas the other is an intact form that is not processed into saposins but is abundant in certain tissues and secretory fluids, including the cerebrospinal fluid. In rats, alternative splicing in the prosaposin gene generates mRNAs with and without a 9-base insertion (Pro+9 and Pro+0 mRNAs, respectively). Pro+9 mRNA is reported to be preferentially expressed in tissues in which the intact form of prosaposin dominates, whereas Pro+0 mRNA is preferentially expressed in tissues in which the precursor dominates. The expression patterns of Pro+9 and Pro+0 mRNAs in the rat choroid plexus are examined in the present study. The specificities of 36-mer oligonucleotide probes used to detect the 9-base insertion by in situ hybridization were demonstrated by dot-blot hybridization. Next, these probes were used for in situ hybridization, which showed predominant expression of Pro+0 mRNA and weak expression of Pro+9 mRNA in the choroid plexus. These expression patterns were confirmed by reverse transcription plus the polymerase chain reaction with AlwI restriction enzyme treatment. Expression of the intact form of prosaposin in the choroid plexus was assessed by Western blotting and immunohistochemistry. Because the choroid plexus is responsible for the generation of cerebrospinal fluid containing the intact form of prosaposin, the present study raises the possibility that Pro+0 mRNA is related to the intact form in the choroid plexus and that the alternatively spliced forms of mRNAs do not simply correspond to the precursor and intact forms of prosaposin.

[From JSLH (The Japanese Society for Laboratory Hematology): An Active Team Approach to Medicine as Laboratory Technologists, through Showing Bone Marrow and Peripheral Blood Samples Directly to Patients with Hematological Malignancy].

The clinical path for the treatment of acute myeloid leukemia (AML) patients has been in practice in our hospital since 2003. In the clinical path, laboratory technologists take on the role of explaining the microscopic findings in bone marrow and peripheral blood samples to patients (with or without their families) using the view-sharing microscope in our laboratory. From July 2003 to October 2014, 56 patients were enrolled in the AML clinical path and given an explanation of their bone marrow and peripheral blood samples. The patients' median age was 62, and the median time spent for explanation was 40 minutes. We conducted a questionnaire feedback survey involving those who enrolled, and the results showed significant improvement in the recognition of the disease pathophysiology, treatment efficacy, and the importance of precautions against infectious diseases. Based on the feedback, we have made marked efforts to provide patients with an improved environment during the explanatory session. This includes installing a special display for the patients, drawing a schematic illustration that shows how the blood cells differentiate, and putting them into operation in a hematology ward to promote patient privacy and precautions against infectious diseases. Hematological laboratory technologists have played an important role in patient care in our hospital. To perform their role as effectively as possible, hematological laboratory technologists participate in the conferences of the Department of Hematology and Oncology regularly, in which medical staff members can discuss the conditions and clinical courses of patients. We aim to contribute to patient satisfaction by sophisticating specialized knowledge as hematological laboratory technologists and cooperate with other medical staff members.

Effects of a comprehensive intervention program, including hot bathing, on physical function in community-dwelling healthy older adults: a pilot randomized controlled trial.

BACKGROUND AND AIMS: To study the effects of a comprehensive intervention program comprising exercise, diet, and hot bathing in community-dwelling older adults by using a randomized controlled trial. METHODS: The program included 61 community-dwelling healthy older adults (mean [SD] age, 69.9 [5.3] years) who were using a hot bath facility. The participants were randomly assigned to four groups as follows: an exercise, diet, and hot bath intervention group (A); an exercise and diet intervention group (B); a hot bath intervention group (C); and a control group (D). Individuals in groups A and B participated in a comprehensive intervention program (including exercise and diet classes) twice a week for 3 months, and those in groups A and C took hot baths. RESULTS: After 3 months, the participants in groups A and B showed a significantly greater improvement in their timed up and go test and stepping test scores than the participants in groups C and D. However, the participants in groups A and C did not show any dependent or independent effects of hot bathing. Three months after the intervention, a follow-up assessment indicated that the group A participants maintained the effect of the intervention and showed improved lower extremity function and health-related quality of life. CONCLUSIONS: The present study suggests that a comprehensive intervention program involving hot bathing may improve lower extremity function and that its effects can be maintained even in healthy older adults. However, the dependent or independent effects of hot bathing may not be expected for healthy older adults.

Relationship between Cognitive Function and Sway of Body in Standing Posture: A Cross-Sectional Study.

BACKGROUND: The influence of neurological or balance dysfunction on cognitive impairment has not been well studied. We compared the results of the balance test, measured by either head or foot sway to consider whole body sway, with those of the cognitive impairment test. METHODS: Individuals of either gender, aged over 60 years, underwent a 30 s balance test. We measured sway while standing on one-leg or two-legs. Sway was evaluated by the distance or area of movement of the head or foot pressure. We also evaluated the effect of visual condition: eyes-open (EO) or -closed (EC). The Mini-Mental State Examination (MMSE) was used to evaluate the degree of cognitive impairment. RESULTS: The head sway area standing on one leg was significantly correlated to MMSE score with EO (correlation r = -0.462). In standing on two legs, no sway test results showed a significant correlation to MMSE scores with EO. With EC, the magnitude of sway became greater, and was significantly correlated to MMSE scores in the head distance. CONCLUSION: Although the correlation between head sway and MMSE was not strong, head sway showed a stronger correlation than did foot pressure sway. Standing on one leg, as measured by head sway area, may thus predict cognitive impairment.

Effects of Daily Consumption of Soy Products on Basic/Instrumental Activities of Daily Living in Community-Dwelling Japanese Women Aged 75 Years and Older: A 4-Year Cohort Study.

Introduction: Since soy isoflavones compensate for age-related estrogen reduction, adequate intake of soy products may prevent the decline in activities of daily living (ADL) due to estrogen reduction in women. However, it is unclear whether regular soy product intake prevents ADL decline. This study examined the effects of soy product consumption on basic/instrumental ADL (BADL/IADL) in Japanese women 75 years or older for 4 years. Materials and Methods: The subject population consisted of 1289 women aged 75 years or older living in Tokyo who underwent private health examinations in 2008. For 1114 (or 1042) participants without baseline BADL (or IADL) disability, we examined the association between baseline soy product consumption frequency and the BADL (or IADL) disabilities 4 years later using logistic regression analyses. The models were adjusted for baseline age, or further for dietary variety for food groups other than soy products, exercise and sport participation, smoking, pre-existing disease number, and body mass index. Results: Regardless of adjustment for potential confounding factors, less frequent soy product consumption was associated with higher BADL or IADL disability incidence. In the fully adjusted models, the trend toward a higher incidence of disabilities with less frequent soy product consumption was statistically significant for both BADL (p = 0.001) and IADL (p = 0.007). Conclusions: Those who consumed soy products more frequently at baseline were less likely to develop BADL and IADL disabilities after 4 years than those who did not. The results show that daily soy product consumption may prevent functional ADL decline in older Japanese women.

PHOSPHATIDYLSERINE SYNTHASE1 is required for microspore development in Arabidopsis thaliana.

Phosphatidylserine (PS) has many important biological roles, but little is known about its role in plants, partly because of its low abundance. We show here that PS is enriched in Arabidopsis floral tissues and that genetic disruption of PS biosynthesis decreased heterozygote fertility due to inhibition of pollen maturation. At1g15110, designated PSS1, encodes a base-exchange-type PS synthase. Escherichia coli cells expressing PSS1 accumulated PS in the presence of l-serine at 23°C. Promoter-GUS assays showed PSS1 expression in developing anther pollen and tapetum. A few seeds with pss1-1 and pss1-2 knockout alleles escaped embryonic lethality but developed into sterile dwarf mutant plants. These plants contained no PS, verifying that PSS1 is essential for PS biosynthesis. Reciprocal crossing revealed reduced pss1 transmission via male gametophytes, predicting a rate of 61.6%pss1-1 pollen defects in PSS1/pss1-1 plants. Alexander's staining of inseparable qrt1-1 PSS1/pss1-1 quartets revealed a rate of 42% having three or four dead pollen grains, suggesting sporophytic pss1-1 cell death effects. Analysis with the nuclear stain 4',6-diamidino-2-phenylindole (DAPI) showed that all tetrads from PSS1/pss1-1 anthers retain their nuclei, whereas unicellular microspores were sometimes anucleate. Transgenic Arabidopsis expressing a GFP-LactC2 construct that binds PS revealed vesicular staining in tetrads and bicellular microspores and nuclear membrane staining in unicellular microspores. Hence, distribution and/or transport of PS across membranes were dynamically regulated in pollen microspores. However, among unicellular microspores from PSS1/pss1-2 GFP-LactC2 plants, all anucleate microspores showed little GFP-LactC2 fluorescence, suggesting that pss1-2 microspores are more sensitive to sporophytic defects or show partial gametophytic defects.

[The influences of exercise fulfillment on mental and physical functions of targeted older adults and the effect of a physical exercise intervention].

OBJECTIVES: To investigate the influence of the differences in exercise fulfillment on mental and physical functions and the effects of exercise intervention on community-dwelling older adults. METHODS: Participants in this study included 260 community-dwelling older adults (mean age +/- SD, 70.4 +/- 6.0 years) who participated in the exercise intervention study (intervention and control groups). Exercise fulfillment levels (low or high), physical activity levels (low or high), mental health (WHO-5 scores), health-related QOL (SF-8 score), and physical abilities of these adults were measured during a baseline health checkup. Based on the status of the 3 exercise fulfillment groups, multivariate analysis of variance (MANOVA), which was adjusted for age, sex, and physical activity levels, was performed to compare the results of the outcome measures among the 3 groups. The intervention group (n = 88, aged 70.3 +/- 6.2 years) was divided into 2 subgroups: the deterioration subgroup (participants with low-exercise fulfillment after the intervention) and the improvement subgroup (participants with high-exercise fulfillment after the intervention). Subsequently, the intervention effects were assessed by repeated measurements of the analysis of variance (ANOVA) between the 2 subgroups. RESULTS: MANOVA analysis revealed that body mass index, grip strength, maximum walking speed, the WHO-5 score, and the SF-8 subscale (8 items) score differed significantly amongst the groups. The high-exercise fulfillment group demonstrated better results for these variables than the low-exercise fulfillment group. Similar results were obtained for each group with respect to the physical activity levels. The repeated-measures ANOVA revealed that time had an important effect on lower physical functions and the SF-8 subscale (1 item) score; it also revealed the important effects of body mass index, the WHO-5 score, the SF-8 subscale (6 items) score, and psychological independence on the group. CONCLUSION: Older adults with higher exercise fulfillment demonstrated better mental and psychological health, regardless of their physical activity levels. Older adults with low-exercise fulfillment could potentially improve their physical abilities; however, their mental and psychological health significantly differed from that of older adults with medium- or high-exercise fulfillment after exercise intervention. These findings provide preliminary evidence, which indicates that exercise can provide sufficient fulfillment and contribute to the promotion and improvement of health in older adults. Moreover, performing adequate tests on exercise fulfillment may aid in assessing the effects of intervention programs in regional healthcare systems.

[The association between foot problems and falls in older adults -a covariance structure analysis-].

AIM: To investigate the manner in which community-dwelling older adults' foot problems affect their history of falls. METHODS: This study included 112 community-dwelling older adults. Foot problems (e.g., inflammation, ingrown nails, and pain while walking), self-rated physical ability (e.g., gait, tripping over, and balance), history of falls within a year, and physical ability (e.g., walking speed, Timed Up & Go test, and one leg balance test) were measured during a routine health checkup. Of these, five subjects were excluded due to incomplete all the measurement. Thus, the subjects eligible for analysis were 107 older adults (mean age±standard deviation=73.0±5.5 years). Covariance structure analysis was used to identify the inter-relationships among all measurements. RESULTS: The covariance structure analysis showed that foot problems negatively influenced participants' self-rated physical ability, and this relationship was also linked to history of falls. The overall fit of this model was judged to be statistically satisfactory (GFI=0.959, AGFI=0.912, CFI=0.981, RMSEA=0.043). CONCLUSIONS: Our model indicated that the association between foot problems and history of falls was affected by self-rated physical ability. Furthermore, in order to prevent falls, the current results suggest that foot care could be an important intervention in older adults to prevent decline in their overall physical ability.