Quick recovery in the generation of self-reactive CD4low natural killer (NK) T cells by an alternative intrathymic pathway when restored from acute thymic atrophy.

The thymus comprises the mainstream of T cell differentiation which produces conventional T cells and an alternative pathway which produces primordial T cells with intermediate density of T cell receptor (TCR)-CD3 complex on the surface (i.e. intermediate TCR cells or TCRint cells). We induced acute thymic atrophy in mice by an administration of hydrocortisone (10 mg) or irradiation (6.5 Gy). It was demonstrated that CD3intCD4lowNK1.1+ T cells were immediately generated by an alternative intrathymic pathway without passing through the double-positive CD4+8+ stage, when restored from thymic atrophy (days 3-14). These CD3intCD4lowNK1.1+ T cells mediated self-reactivity and appeared even in the periphery. mRNA of an invariant chain of TCR Valpha14Jalpha281 gene product was detected in these CD4low T cells, but not remaining CD4high T cells. The mainstream of T cell differentiation in the thymus was not restored up to day 14 and there was no leakage of self-reactive clones into the population generated through the mainstream. These results reveal that an alternative intrathymic pathway is associated with the generation of self-reactive T cells, in an early restoration phase after thymic atrophy.     

The Relationship Between Gait Function and Erectile Dysfunction: Results from a Community-Based Cross-Sectional Study in Japan

IntroductionAlthough physical activity is associated with a decreased risk of erectile dysfunction (ED), the association of ED with physical function remains unclear.AimTo investigate the relationship between gait function and ED in a community-dwelling population.MethodsThis cross-sectional study analyzed 324 men who participated in the Iwaki Health Promotion Project in 2015 in Hirosaki, Japan. ED was assessed with the 5-Item International Index of Erectile Function (IIEF-5). The participants were divided into 2 groups: low IIEF-5 score (≤16) and high IIEF-5 score (>16). We evaluated physical function, including gait function and grip strength. Gait function was evaluated by 10-meter gait speed and 2-step score (the ratio of the maximum length of 2 strides to height). We assessed daily physical activity, comorbidities, mental status, and laboratory data. The association between physical function and a low IIEF-5 score was analyzed by multivariate logistic regression analysis.Main Outcome MeasureThe main outcome measure was to assess whether gait function was an independent indicator for erectile dysfunction.ResultsOf 324 men, 154 (48%) had a low IIEF-5 score. Grip strength, 2-step score, and 10-meter gait speed in the low IIEF-5 group were significantly inferior to those in the high IIEF-5 group. Multivariate analysis showed that the 2-step score (odds ratio = 0.08), age, and total testosterone were independently associated with a low IIEF-5.Clinical ImplicationsThis study may motivate clinicians to investigate predictive values of physical function for ED.Strengths & LimitationsThe strength of this study was the use of simple, objective, and feasible tests for gait function to assess its association with ED. The limitations of this study were selection bias, regional bias, and nature of the cross-sectional study.ConclusionsOf the gait functional tests, not the 10-meter gait speed but 2-step score was an independent indicator for the presence of ED.Okamoto T, Hatakeyama S, Imai A, et al. The Relationship Between Gait Function and Erectile Dysfunction: Results from a Community-Based Cross-Sectional Study in Japan. J Sex Med 2019; 16:1922–1929.     

Simultaneous activation of natural killer T cells and autoantibody production in mice injected with denatured syngeneic liver tissue

Denatured syngeneic liver tissue prepared by mechanical procedures was intraperitoneally injected into adult C57BL/6 mice. In parallel with a decrease in the total number of lymphocytes in the liver, spleen, and thymus from days 1-7 after the injection, the proportion of the CD4+NK1.1+CD3(int) subset of these cells (i.e. natural killer T or NKT cells) increased in the liver. Even the absolute number of these NKT cells increased in the liver on days 14 and 21. In response to the injection of denatured liver tissue, tissue damage was induced in the liver, as shown by elevated levels of serum transaminases and hepatocyte degeneration observed by electron microscopy. Sera obtained on days 7 and 14 contained autoantibodies including anti-DNA antibodies. The proportion of CD1d(high)B cells in the liver was found to decrease on days 1-7. In other words, denatured liver tissue stimulated both NKT cells and certain B cells in the liver. These results suggest that liver lymphocytes might contain not only autoreactive T cells (e.g. CD3(int) or NKT cells) but also some B cells (e.g. B-1 cells) which produce autoantibodies and that the denatured tissue had the potential to stimulate these lymphocytes and to evoke an autoimmune-like state.     

Evaluation of DNA and RNA quality from archival formalin‐fixed paraffin‐embedded tissue for next‐generation sequencing – Retrospective study in Japanese single institution

Genetic analysis on formalin‐fixed paraffin‐embedded (FFPE) tissue specimens has become a mainstream method, from conventional direct sequencing to comprehensive analysis using next‐generation sequencing (NGS). In this study, we evaluated the quality of DNA and RNA extracted from FFPE sections, derived from surgical specimens of different tumor types. Electrophoresis was performed using a 4200 TapeStation to evaluate DNA and RNA fragmentation. DNA Ct values were higher and significantly increased over a period of 4 years compared with those from cell lines or frozen tissues. The RNA integrity number equivalent (RIN) ranged from 1 to 4.1 and DV200 ranged from 7.3 to 81%. Twelve of the 108 cases were analyzed by NGS using the AmpliSeq Cancer HotSpot Panel v2 on a Miniseq system. A sufficient number of reads and coverage were obtained in all cases. Our results revealed that NGS analysis was sufficient for FFPE‐derived DNA within 4 years of preservation. Conversely, approximately 20% of the RNA derived from FFPE within 4 years from the collection could be inappropriate for gene analysis based on RIN and DV200. It was suggested that FFPE would be adequate for genetic analysis, although it is desirable to store frozen specimens for the tumor tissues to be subjected to genetic analysis.     

Ro52 functionally interacts with IgG1 and regulates its quality control via the ERAD system

The 52-kDa form of SSA/Ro protein (Ro52) is one of autoantigens associated with autoimmune disorders such as systemic lupus erythematosus and Sj?gren's syndrome. Anti-SSA/Ro antibodies, the biological function of which remains unknown, are frequently found in the serum of these patients. Recent functional genomic approaches have shown that Ro52/TRIM21 is one of the TRIM family proteins with a RING-finger domain which is closely associated with E3 ubiquitin ligase activity. We found by using yeast-two hybrid screening that Ro52 has an E3 activity in vitro and interacts with human IgG1 heavy chain. We also found that IgG1 heavy chain was modified with polyubiquitination by Ro52 and degraded through the ubiquitin-proteasome system in mammalian cells. Our results also showed that Ro52 interacts with the molecular chaperone p97/VCP, which is thought to function in the endoplasmic reticulum associated degradation (ERAD) system. It is likely that Ro52 plays a role in proteasomal degradation of unfolded IgG1, which is retrogradely transferred from the endoplasmic reticulum to the cytosol. Taken together, our findings suggest that Ro52 plays a significant role in quality control of IgG1 through the ERAD system.     

Transient appearance of hepatic natural killer cells with high cytotoxicity and unique phenotype in very young mice

There were few natural killer (NK) cells in the liver in very young mice at the age of 1-2 weeks. This was because the cell yield from the liver of young mice was low. The percentage of NK cells in the liver of young mice, however, was almost comparable with that in the liver of adult mice. Lymphocytes were isolated from the liver and spleen of C57BL/6 (B6) mice, and NK cytotoxicity and phenotype were herein examined in this study. NK cytotoxicity was extremely high in the liver of very young mice. This phenomenon was seen in the liver of various normal mouse strains. In contrast, the appearance of high cytotoxicity was not seen in NK cells of the spleen, irrespective of mouse strains. The quality of NK cells in the liver of young mice was different from that in adult mice. NK cells in the liver of young mice were mainly CD69(+)Mac-1(-) Fas ligand(+), whereas those in the liver of adult mice were CD69(-)Mac-1(+) Fas ligand(-). These results revealed that the quality of hepatic NK cells changes in the process of ageing. Namely, liver NK cells in very young mice temporarily show the highest NK cytotoxicity and a unique activated phenotype. Physiological meaning of the present phenomenon was discussed.     

Five cases of listeriosis in the elderly

We reported five cases of listeriosis (sepsis and meningitis) in the elderly in our hospital during the last 4 years, where no cases of listeriosis had been found. These 5 cases had diabetes mellitus, lung cancer, chronic respiratory failure, gastric ulcer and aplastic anemia respectively as their underlying diseases. At the onset of listeriosis, 3 cases received corticosteroid and 3 cases received H2-blocker. 2 patients were cured and 3 patients died. Three autopsy cases had meningitis or meningoencephalitis and 2 cases of these autopsy cases had granulomatous changes in these spleens. In serotypes of Listeria monocytogenes (L. monocytogenes), 4 cases were 4b and 1 cases was 1b. All 5 strains were resistant to 3rd generation cephems. Wide uses of 3rd generation cephems and H2-blocker may be one of the reasons for the recent increase of listeriosis. Ingestion of contaminated food is the pathogenetic mechanism for initiating L. monocytogenes infections. And following the change of eating habits and the increase of imported foods, food-born listeriosis may increase. We suppose the increase of L. monocytogenes infections and must give attention to L. monocytogenes infections.     

Fully parametric imaging with reversible tracer <Superscript>18</Superscript>F-FLT within a reasonable time

PET enables quantitative imaging of the rate constants K ₁, k ₂, k ₃, and k ₄, with a reversible two tissue compartment model (2TCM). A new method is proposed for computing all of these rates within a reasonable time, less than 1 min. A set of differential equations for the reversible 2TCM was converted into a single formula consisting of differential and convolution terms. The validity was tested on clinical data with ¹⁸F-FLT PET for patients with glioma (n = 39). Parametric images were generated with the formula that was developed. Parametric values were extracted from regions of interest (ROIs) for glioma from the images generated, and they were compared with those obtained with the non-linear fitting method. We performed simulation studies for testing accuracy by generating simulated images, assuming clinically expected ranges of the parametric values. The computation time was about 20 s, and the quality of the images generated was acceptable. The values obtained for K ₁ for grade IV tumor were 0.24 ± 0.23 and 0.26 ± 0.25 ml^?1 min^?1 g^?1 for the image-based and ROI-based methods, respectively. The values were 0.21 ± 0.12 and 0.21 ± 0.12 min^?1 for k ₂, 0.13 ± 0.07 and 0.13 ± 0.07 min^?1 for k ₃, and 0.052 ± 0.020 and 0.054 ± 0.021 min^?1 for k ₄. The differences between the methods were not significant. Regression analysis showed correlations of r = 0.94, 0.86, 0.71, and 0.52 for these parameters. Simulation demonstrated that the accuracy was within acceptable ranges, namely, the correlations were r = 0.99, r = 0.97, r = 0.99, and r = 0.91 for K ₁, k ₂, k ₃, and k ₄, respectively, between estimated and assumed values. This results suggest that parametric images can be obtained fully within reasonable time, accuracy, and quality.