Sendai Virus and a Unified Model of Mononegavirus RNA Synthesis

Vesicular stomatitis virus (VSV), the founding member of the mononegavirus order (Mononegavirales), was found to be a negative strand RNA virus in the 1960s, and since then the number of such viruses has continually increased with no end in sight. Sendai virus (SeV) was noted soon afterwards due to an outbreak of newborn pneumonitis in Japan whose putative agent was passed in mice, and nowadays this mouse virus is mainly the bane of animal houses and immunologists. However, SeV was important in the study of this class of viruses because, like flu, it grows to high titers in embryonated chicken eggs, facilitating the biochemical characterization of its infection and that of its nucleocapsid, which is very close to that of measles virus (MeV). This review and opinion piece follow SeV as more is known about how various mononegaviruses express their genetic information and carry out their RNA synthesis, and proposes a unified model based on what all MNV have in common.     

Novel orally bioavailable EZH1/2 dual inhibitors with greater antitumor efficacy than an EZH2 selective inhibitor

Polycomb repressive complex 2 (PRC2) methylates histone H3 lysine 27 and represses gene expression to regulate cell proliferation and differentiation. Enhancer of zeste homolog 2 (EZH2) or its close homolog EZH1 functions as a catalytic subunit of PRC2, so there are two PRC2 complexes containing either EZH2 or EZH1. Tumorigenic functions of EZH2 and its synthetic lethality with some subunits of SWItch/Sucrose Non‐Fermentable (SWI/SNF) chromatin remodeling complexes have been observed. However, little is known about the function of EZH1 in tumorigenesis. Herein, we developed novel, orally bioavailable EZH1/2 dual inhibitors that strongly and selectively inhibited methyltransferase activity of both EZH2 and EZH1. EZH1/2 dual inhibitors suppressed trimethylation of histone H3 lysine 27 in cells more than EZH2 selective inhibitors. They also showed greater antitumor efficacy than EZH2 selective inhibitor in vitro and in vivo against diffuse large B‐cell lymphoma cells harboring gain‐of‐function mutation in EZH2. A hematological cancer panel assay indicated that EZH1/2 dual inhibitor has efficacy against some lymphomas, multiple myeloma, and leukemia with fusion genes such as MLL‐AF9, MLL‐AF4, and AML1‐ETO. A solid cancer panel assay demonstrated that some cancer cell lines are sensitive to EZH1/2 dual inhibitor in vitro and in vivo. No clear correlation was detected between sensitivity to EZH1/2 dual inhibitor and SWI/SNF mutations, with a few exceptions. Severe toxicity was not seen in rats treated with EZH1/2 dual inhibitor for 14 days at drug levels higher than those used in the antitumor study. Our results indicate the possibility of EZH1/2 dual inhibitors for clinical applications.     

CCR5-deficient mice develop experimental autoimmune uveoretinitis in the context of a deviant effector response

PURPOSE: Experimental autoimmune uveoretinitis (EAU) is an organ-specific, Th1-cell-mediated disease that targets the neural retina. CCR5 is a chemokine receptor expressed on Th1 cells that promotes their migration. In CCR5-deficient mice, we examined the role of CCR5 in the development of EAU induced by immunization with interphotoreceptor retinoid-binding protein (IRBP) peptide. METHODS: Wild-type or CCR5-deficient B6 mice were immunized with human IRBP peptide 1-20 (hIRBP-p), and the severity of EAU was assessed clinically and histologically. Splenocytes and cells of regional lymph nodes near the eye were collected and their proliferation and production of IL-6, IL-10, IFN-gamma, and CCL2 (MCP-1) in response to hIRBP-p stimulation were measured. Moreover, the intraocular levels of these cytokines were analyzed. RESULTS: Immunization with hIRBP-p induced EAU in CCR5-deficient mice with a severity comparable to that in wild-type mice. Histologically, T-cell infiltration of the eye was reduced, but granulocyte infiltration was augmented in CCR5-deficient mice. Although splenic T cells from CCR5-deficient mice produced IFN-gamma but not IL-10 on stimulation by hIRBP-p, T cells from the regional lymph nodes failed to produce both cytokines. IL-6 production in the eye and IL-6 and CCL2 production by splenic T cells were predominantly augmented in CCR5-deficient mice. CONCLUSIONS: The development of EAU is not prevented in CCR5-deficient mice. Although T-cell infiltration into the eye is apparently reduced in CCR5-deficient mice, the defect is compensated for by granulocyte infiltration, supposedly mediated by augmented intraocular production of IL-6.     

Possibility of inducing anterior chamber-associated immune deviation by TGF-beta2 treatment of monocytes isolated from Behcet's patients

Murine macrophages treated with TGF-beta2 are capable of inducing anterior chamber-associated immune deviation (ACAID), and these macrophages are characterized by impaired IL-12 production and CD40 expression, consequently failing to promote Th1 cell differentiation. In this study, we investigated whether human monocytes can also acquire the specific functions by TGF-beta2 treatment, even when the monocytes are isolated from patients with Behcet's disease (BD). Adherent monocytes isolated from peripheral blood mononuclear cells (PBMC) of 16 BD patients and 16 healthy controls, were cultured overnight with or without 5 ng/ml of TGF-beta2. Then, TGF-beta2-treated or untreated adherent cells were co-cultured with allogeneic CD4(+) T cells obtained from healthy subjects. TGF-beta2 treatment inhibited the abilities of adherent monocytes obtained from BD patients to stimulate the proliferation and IFN-gamma production of allogeneic CD4(+) T cells. The reduced IFN-gamma production was also confirmed by IFN-gamma mRNA expression in the co-cultured T cells. IL-12 production and CD40 molecule expression by adherent monocytes obtained from BD patients were strikingly reduced by TGF-beta2 treatment. These results suggest a possibility that adherent monocytes isolated from BD patients may acquire a property to induce ACAID by treatment with TGF-beta2.     

A mouse model for Stickler's syndrome: ocular phenotype of mice carrying a targeted heterozygous inactivation of type II (pro)collagen gene (Col2a1)

The influences of targeted heterozygous inactivation of type II (pro)collagen gene (Col2a1) on eye structures in the 15-month-old C57BL/6JOlaHsd mouse was studied. The eyes were collected from C57BL mice heterozygous for a targeted inactivation of one allele of the Col2a1 gene (Col2a1(+/-) mice). The eyes of C57BL mice with normal gene alleles were used as controls (Col2a1(+/+) mice). Ocular histology was analyzed from tissue sections, stained with hematoxylin and eosin, toluidine blue and alcian blue. Type II collagen was localized by immunohistochemistry. Hyaluronan (HA) was stained utilizing the biotinylated complex of the hyaluronan-binding region of aggrecan and link protein (bHABC). The anterior segment of the eye was well-formed in both genotypes, but typical folding of ciliary processes was decreased, while increased stromal extracellular matrix vacuolization was seen in the Col2a1(+/-) mice. In the lens of these mice, subcapsular extracellular matrix changes were observed. Differences in retinal structures or the number of the eyes with retinal detachment were not detected between the genotypes. In Col2a1(+/-) mice, staining for type II collagen was weaker in cornea, ciliary body, iris, lens, vitreous, retina, choroid and sclera than in the control mice. HA staining was detected in the extraocular tissues, ciliary body, iris and the choroid of both genotypes. HA staining was observed only in the vitreous body of the control animals. Heterozygous inactivation of Col2a1 gene causes structural defects in the murine eye. The observed structural changes in the ciliary body, lens and vitreous of the Col2a1(+/-) mice may represent ocular features found in the human Stickler syndrome, where the abnormalities result from COL2A1 gene mutations which lead to functional haploinsufficiency.     

Effects of glycyrrhetinic acid on aminonucleoside nephrosis in rats

The effects of glycyrrhetinic acid (GA), an aglycon of glycyrrhizin extracted from the roots of Glycyrrhizae radix, on puromycin aminonucleoside (PA) nephrosis were studied in rats. Urine protein excretion in female rats (130g–150g) receiving PA (50 mg/kg) alone was significantly elevated on the 2nd day after injection of PA and reached a peak on the 14th day. Urinary protein on the 14th day was reduced to 74% in animals treated with GA (20 mg/kg) starting on the 2nd day after injection of PA. The increase in serum cholesterol and the decrease in serum protein were also suppressed by GA. Observation by electron microscopy revealed that the degree of abnormality in glomerular epithelial cells, i.e. loss or fusion of foot processes, was lower in the rats treated with GA after PA injection than in the rat treated with PA alone. Moreover, pretreatment with GA did not suppress urinary protein excretion but when it was given at the same time as PA and after PA a significant decrease in urinary protein excretion was observed. Correspondence to: H. Abe at the above address     

Simple identification of Trichophyton tonsurans by chlamydospore‐like structures produced in culture media

Trichophyton tonsurans is known to be the causative agent of a worldwide epidemic of dermatophytoses among contact sports practitioners, and is spreading among the general population of Japan. Prompt and simple identification of T. tonsurans in diagnostic laboratories is crucial to control infection. The present study evaluated the availability of observation of chlamydospore‐like structures grown in culture media as a characteristic for identification of T. tonsurans. Twenty‐five strains of T. tonsurans and five strains each of Trichophyton verrucosum, Trichophyton rubrum and Trichophyton mentagrophytes were inoculated on Mycosel agar plates and inoculated Petri dishes were observed by light microscopy from the reverse side. Twenty‐three of 25 T. tonsurans strains showed chlamydospore‐like structures within 5 days, and all strains at day 8. The numbers of chlamydospore‐like structures were very abundant in most strains. The majority of strains of other species showed no chlamydospore‐like structures, or very few when present. Positive for chlamydospore‐like structures among 15 strains other than T. tonsurans was one strain at day 5 and six strains at day 8. As for the identification of T. tonsurans, presence of chlamydospore‐like structures showed 92.0% sensitivity (23/25) and 93.3% specificity (14/15) at day 5, and 100% sensitivity (25/25) and 60.0% specificity (9/15) at day 8. Electron microscopic findings suggest chlamydospore‐like structures are not true chlamydospores but are produced by inflation of actively growing hyphae by developing vacuoles in cells. In conclusion, observation of development of chlamydospore‐like structures in culture media is the simplest method for identification of T. tonsurans.     

Additive effects of orthokeratology and atropine 0.01% ophthalmic solution in slowing axial elongation in children with myopia: first year results

Purpose

To investigate the additive effects of orthokeratology (OK) and atropine 0.01% ophthalmic solution, both of which are effective procedures to slow axial elongation in children with myopia.

Study design

Prospective randomized clinical trial.

Methods

Japanese children aged 8–12 years with a spherical equivalent refractive error of ??1.00 to ??6.00 diopters were included. A total of 41 participants who had been wearing the OK lenses successfully for 3 months were randomly allocated into two groups to receive either the combination of OK and atropine 0.01% ophthalmic solution (combination group) or monotherapy with OK (monotherapy group). Subjects in the combination group started to use atropine 0.01% ophthalmic solution once nightly from 3 months after the start of OK. Axial length was measured every 3 months using non-contact laser interferometry (IOLMaster), and the axial length measurement at month 3 of OK therapy was used as the baseline value in both groups. The increase in axial length over 1 year was compared between the two groups.

Results

A total of 40 consecutive subjects (20 subjects in the combination group and 20 in the monotherapy group) were followed for 1 year. The increase in axial length over 1 year was 0.09?±?0.12 mm in the combination group and 0.19?±?0.15 mm in the monotherapy group (P?=?0.0356, unpaired t test).

Conclusion

During the 1-year follow-up, the combination of OK and atropine 0.01% ophthalmic solution was more effective in slowing axial elongation than OK monotherapy in children with myopia.