Spatial variation of microtubule depolymerization in large asters

Microtubule plus-end depolymerization rate is a potentially important target of physiological regulation, but it has been challenging to measure, so its role in spatial organization is poorly understood. Here we apply a method for tracking plus ends based on time difference imaging to measure depolymerization rates in large interphase asters growing in Xenopus egg extract. We observed strong spatial regulation of depolymerization rates, which were higher in the aster interior compared with the periphery, and much less regulation of polymerization or catastrophe rates. We interpret these data in terms of a limiting component model, where aster growth results in lower levels of soluble tubulin and microtubule-associated proteins (MAPs) in the interior cytosol compared with that at the periphery. The steady-state polymer fraction of tubulin was ∼30%, so tubulin is not strongly depleted in the aster interior. We propose that the limiting component for microtubule assembly is a MAP that inhibits depolymerization, and that egg asters are tuned to low microtubule density.     

Unweighted state as a sidestep preparation improve the initiation and reaching performance for basketball players

The preparatory motion of a defensive motion in contact sport such as basketball should be small and involve landing on both feet for strict time and motion constraints. We thus proposed the movement creating a unweighted state. Ten basketball players performed a choice reaction sidestepping task with and without the voluntary, continuous vertical fluctuation movement. The results indicated that the preparatory movement shortened the time of their sidestep initiation (301 vs. 314 ms, p = 0.011) and reaching performance (883 vs. 910 ms, p = 0.018) but did not increase their peak ground reaction force or movement velocity. The mechanism of the improvement was estimated to be the following: in the preparation phase, the vertical body fluctuation created the force fluctuation; after the direction signal, the unweighted state can shorten the time required to initiate the sidestepping (unweighted: 279 ms; weighted: 322 ms, p = 0.002); around the initiation phase, the dropping down of the body and weighted state can contribute to the reaching performance. We conducted additional experiment investigating muscle–tendon-complex dynamics and muscle activity using ultrasound device and electromyography. The result suggests that the building up of active state of muscle might explain the improvement of sidestepping performance.     

Synthesis of 2-Alkynylcordycepins and Evaluation of Their Vasodilating Activity

Abstract

Based on the recently developed lithiation-mediated stannyl migration of 6-chloropurine derivatives, 2-iodocordycepin was prepared from cordycepin. The reaction of this compound with terminal alkynes was carried out to synthesize a series of 2-alkynyl derivatives. The vasodilating effect of these compounds was evaluated.     

Apoptosis inhibitor of macrophage (AIM) expression in alveolar macrophages in COPD

Background

Marked accumulation of alveolar macrophages (AM) conferred by apoptosis resistance has been implicated in pathogenesis of chronic obstructive pulmonary disease (COPD). Apoptosis inhibitor of macrophage (AIM), has been shown to be produced by mature tissue macrophages and AIM demonstrates anti-apoptotic property against multiple apoptosis-inducing stimuli. Accordingly, we attempt to determine if AIM is expressed in AM and whether AIM is involved in the regulation of apoptosis in the setting of cigarette smoke extract (CSE) exposure.

Methods

Immunohistochemical evaluations of AIM were performed. Immunostaining was assessed by counting total and positively staining AM numbers in each case (n = 5 in control, n = 5 in non-COPD smoker, n = 5 in COPD). AM were isolated from bronchoalveolar lavage fluid (BALF). The changes of AIM expression levels in response to CSE exposure in AM were evaluated. Knock-down of anti-apoptotic Bcl-xL was mediated by siRNA transfection. U937 monocyte-macrophage cell line was used to explore the anti-apoptotic properties of AIM.

Results

The numbers of AM and AIM-positive AM were significantly increased in COPD lungs. AIM expression was demonstrated at both mRNA and protein levels in isolated AM, which was enhanced in response to CSE exposure. AIM significantly increased Bcl-xL expression levels in AM and Bcl-xL was involved in a part of anti-apoptotic mechanisms of AIM in U937 cells in the setting of CSE exposure.

Conclusions

These results suggest that AIM expression in association with cigarette smoking may be involved in accumulation of AM in COPD.     

Deficiency in Clonogenic Endometrial Mesenchymal Stem Cells in Obese Women with Reproductive Failure – a Pilot Study

Objectives

The mechanisms of obesity associated reproductive complications remain poorly understood. Endometrial mesenchymal stem-cells are critical for cyclic renewal and uterine function. Recently, W5C5⁺ cells, with high clonogenicity, capable of producing endometrial stroma in vivo, have been described. We sought to investigate the abundance and cloning efficiency of W5C5⁺ and W5C5⁻ endometrial cells in relation to Body Mass Index, age and reproductive outcome.

Design

W5C5⁺ and W5C5⁻ cells were purified from mid-luteal endometrial biopsies (n = 54) by magnetic bead separation and subjected to in vitro colony-forming assays.

Results

First trimester pregnancy losses were significantly higher in obese subjects (n = 12) compared to overweight (n = 20) and subjects with normal Body Mass Index (n = 22) (P<0.05, P<0.01, respectively). W5C5⁺ cells (%) were significantly lower in obese subjects compared to subjects with normal Body Mass Index (P<0.05). W5C5⁺ cloning efficiency was significantly lower in obese subjects compared to overweight and subjects with normal Body Mass Index (P<0.05, respectively). W5C5⁻ cloning efficiency was significantly lower in obese subjects compared to subjects with normal Body Mass Index (P<0.05). Body Mass Index was significantly negatively correlated with W5C5⁺ cloning efficiency and W5C5⁻ cloning efficiency (P<0.01, respectively), and positively correlated with first trimester loss (P<0.01). We found no significant results with age (P>0.05).

Conclusions

Our observations suggest that the regenerative capacity and plasticity of the endometrium of obese women is suboptimal, which in turn may account for the increased risk of reproductive complications associated with obesity.     

Establishment of Alternative Culture Method for Spermatogonial Stem Cells Using Knockout Serum Replacement

Since spermatogonial stem cells (SSCs) are capable of both self-renewal and differentiation to daughter cells for subsequent spermatogenesis, the development of an efficient in vitro culture system is essential for studies related to spermatogenesis. Although the currently available system is serum-free and contains only chemically-defined components, it highly relies upon bovine serum albumin (BSA), a component with batch-to-batch quality variations similar to those of fetal bovine serum. Thus, we searched for an alternative BSA-free culture system that preserved the properties of SSCs. In this study, we utilized Knockout Serum Replacement (KSR) in the SSC culture medium, as a substitute for BSA. The results demonstrated that KSR supported the continuous growth of SSCs in vitro and the SSC activity in vivo without BSA, in a feeder-cell combination with mouse embryonic fibroblasts. The addition of BSA to KSR further facilitated cell cycle progression, whereas a transplantation assay revealed that the addition of BSA did not affect the number of SSCs in vivo. The combination of KSR with BSA also allowed the elimination of GFRA1 and FGF2, and the reduction of the GDNF concentration from 20 ng/ml to 5 ng/ml, while maintaining the growth rate and the expression of SSC markers. Furthermore, KSR was also useful with SSCs from non-DBA/2 strains, such as C57BL/6 and ICR. These results suggested that KSR is an effective substitute for BSA for long-term in vitro cultures of SSCs. Therefore, this method is practical for various studies related to SSCs, including spermatogenesis and germ stem cell biology.     

Identification of Biological Properties of Intralymphatic Tumor Related to the Development of Lymph Node Metastasis in Lung Adenocarcinoma

Background

Intralymphatic tumors in the extratumoral area are considered to represent the preceding phase of lymph node metastasis. The aim of this study was to clarify the biological properties of intralymphatic tumors susceptible to the development of lymph node metastasis, with special reference to the expression of cancer initiating/stem cell (CIC/CSC) related markers in cancer cells and the number of infiltrating stromal cells.

Material and Methods

Primary lung adenocarcinomas with lymphatic permeation in the extratumoral area were retrospectively examined (n = 107). We examined the expression levels of CIC/CSC related markers including ALDH1, OCT4, NANOG, SOX2 and Caveolin-1 in the intralymphatic cancer cells to evaluate their relationship to lymph node metastasis. Moreover, the number of infiltrating stromal cells expressing CD34, α-smooth muscle actin, and CD204 were also evaluated.

Results

Among the intralymphatic tissues, low ALDH1 expression in cancer cells, high SOX2 expression in cancer cells, and a high number of CD204(+) macrophages were independent predictive factors for lymph node metastasis (P = 0.004, P = 0.008, and P = 0.028, respectively). Among these factors, only low ALDH1 expression in cancer cells was significantly correlated with the farther spreading of lymph node metastasis (mediastinal lymph node, pathological N2) (P = 0.046) and the metastatic lymph node ratio (metastatic/resected) (P = 0.028). On the other hand, in the primary tumors, ALDH1 expression in the cancer cells was not associated with lymph node metastasis. Intralymphatic cancer cells expressing low ALDH1 levels exhibited lower E-cadherin expression levels than cancer cells with high levels of ALDH1 expression (P = 0.015).

Conclusions

Intralymphatic cancer cells expressing low levels of ALDH1 and infiltrating macrophages expressing CD204 have a critical impact on lymph node metastasis. Our study also highlighted the significance of evaluating the biological properties of intralymphatic tumors for tumor metastasis.     

The Impact of Model Building on the Transmission Dynamics under Vaccination: Observable (Symptom-Based) versus Unobservable (Contagiousness-Dependent) Approaches

Background

The way we formulate a mathematical model of an infectious disease to capture symptomatic and asymptomatic transmission can greatly influence the likely effectiveness of vaccination in the presence of vaccine effect for preventing clinical illness. The present study aims to assess the impact of model building strategy on the epidemic threshold under vaccination.

Methodology/Principal Findings

We consider two different types of mathematical models, one based on observable variables including symptom onset and recovery from clinical illness (hereafter, the “observable model”) and the other based on unobservable information of infection event and infectiousness (the “unobservable model”). By imposing a number of modifying assumptions to the observable model, we let it mimic the unobservable model, identifying that the two models are fully consistent only when the incubation period is identical to the latent period and when there is no pre-symptomatic transmission. We also computed the reproduction numbers with and without vaccination, demonstrating that the data generating process of vaccine-induced reduction in symptomatic illness is consistent with the observable model only and examining how the effective reproduction number is differently calculated by two models.

Conclusions

To explicitly incorporate the vaccine effect in reducing the risk of symptomatic illness into the model, it is fruitful to employ a model that directly accounts for disease progression. More modeling studies based on observable epidemiological information are called for.     

Basal and Ischemia-Induced Transcardiac Troponin Release into the Coronary Circulation in Patients with Suspected Coronary Artery Disease

Background

Cardiac troponin is a specific biomarker for cardiomyocyte necrosis in acute coronary syndromes. Troponin release from the coronary circulation remains to be determined because of the lower sensitivity of the conventional assay. We sought to determine basal and angina-induced troponin release using a highly sensitive troponin assay.

Methods and Results

The cardiac troponin T levels in serum sampled from the peripheral vein (PV), the aortic root (AO), and the coronary sinus (CS) were measured in 105 consecutive stable patients with coronary risk factor(s) and suspected coronary artery disease (CAD) and in 33 patients without CAD who underwent an acetylcholine provocation test. At baseline, there was a significant increase in the troponin levels from AO [9.0 (6.4, 13.1) pg/mL for median (25^th, 75^th percentiles)] to CS [10.3 (7.3, 15.5) pg/mL, p<0.001] in 96 (91.4%) patients and the difference was 1.1 (0.4, 2.1) pg/mL, which reflected basal transcardiac troponin release (TTR). TTR was positively correlated with PV levels (r = 0.22, p = 0.03). Male sex, left ventricular hypertrophy determined by echocardiography, T-wave inversion, and CAD correlated with elevated TTR defined as above: median, 1.1 pg/mL. A significant increase in TTR was noted in 17 patients with coronary spasms [0.6 (0.2, 1.2) pg/mL, p<0.01] but not in 16 patients without spasms [0.0 (−0.5, 0.9) pg/mL, p = 0.73] after the acetylcholine provocation.

Conclusion

Basal TTR in the coronary circulation was observed in most of the patients with suspected CAD and risk factor(s). This sensitive assay detected myocardial ischemia-induced increases in TTR caused by coronary spasms.