Two novel mutations in the EYS gene are possible major causes of autosomal recessive retinitis pigmentosa in the Japanese population

Retinitis pigmentosa (RP) is a highly heterogeneous genetic disease including autosomal recessive (ar), autosomal dominant (ad), and X-linked inheritance. Recently, arRP has been associated with mutations in EYS (Eyes shut homolog), which is a major causative gene for this disease. This study was conducted to determine the spectrum and frequency of EYS mutations in 100 Japanese arRP patients. To determine the prevalence of EYS mutations, all EYS exons were screened for mutations by polymerase chain reaction amplification, and sequence analysis was performed. We detected 67 sequence alterations in EYS, of which 21 were novel. Of these, 7 were very likely pathogenic mutations, 6 were possible pathogenic mutations, and 54 were predicted non-pathogenic sequence alterations. The minimum observed prevalence of distinct EYS mutations in our study was 18% (18/100, comprising 9 patients with 2 very likely pathogenic mutations and the remaining 9 with only one such mutation). Among these mutations, 2 novel truncating mutations, c.4957_4958insA (p.S1653KfsX2) and c.8868C>A (p.Y2956X), were identified in 16 patients and accounted for 57.1% (20/35 alleles) of the mutated alleles. Although these 2 truncating mutations were not detected in Japanese patients with adRP or Leber's congenital amaurosis, we detected them in Korean arRP patients. Similar to Japanese arRP results, the c.4957_4958insA mutation was more frequently detected than the c.8868C>A mutation. The 18% estimated prevalence of very likely pathogenic mutations in our study suggests a major involvement of EYS in the pathogenesis of arRP in the Japanese population. Mutation spectrum of EYS in 100 Japanese patients, including 13 distinct very likely and possible pathogenic mutations, was largely different from the previously reported spectrum in patients from non-Asian populations. Screening for c.4957_4958insA and c.8868C>A mutations in the EYS gene may therefore be very effective for the genetic testing and counseling of RP patients in Japan.     

Telephone cognitive-behavioral therapy for subthreshold depression and presenteeism in workplace: a randomized controlled trial

Background

Subthreshold depression is highly prevalent in the general population and causes great loss to society especially in the form of reduced productivity while at work (presenteeism). We developed a highly-structured manualized eight-session cognitive-behavioral program with a focus on subthreshold depression in the workplace and to be administered via telephone by trained psychotherapists (tCBT).

Methods

We conducted a parallel-group, non-blinded randomized controlled trial of tCBT in addition to the pre-existing Employee Assistance Program (EAP) versus EAP alone among workers with subthreshold depression at a large manufacturing company in Japan. The primary outcomes were depression severity as measured with Beck Depression Inventory-II (BDI-II) and presenteeism as measured with World Health Organization Health and Work Productivity Questionnaire (HPQ). In the course of the trial the follow-up period was shortened in order to increase acceptability of the study.

Results

The planned sample size was 108 per arm but the trial was stopped early due to low accrual. Altogether 118 subjects were randomized to tCBT+EAP (n = 58) and to EAP alone (n = 60). The BDI-II scores fell from the mean of 17.3 at baseline to 11.0 in the intervention group and to 15.7 in the control group after 4 months (p<0.001, Effect size = 0.69, 95%CI: 0.32 to 1.05). However, there was no statistically significant decrease in absolute and relative presenteeism (p = 0.44, ES = 0.15, −0.21 to 0.52, and p = 0.50, ES = 0.02, −0.34 to 0.39, respectively).

Conclusion

Remote CBT, including tCBT, may provide easy access to quality-assured effective psychotherapy for people in the work force who present with subthreshold depression. Further studies are needed to evaluate the effectiveness of this approach in longer terms. The study was funded by Sekisui Chemicals Co. Ltd.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00885014","term_id":"NCT00885014"}}NCT00885014     

Production, characterization and applications for Toxoplasma gondii-specific polyclonal chicken egg yolk immunoglobulins

Background

Toxoplasma gondii may cause abortions, ocular and neurological disorders in warm-blood hosts. Immunized mammals are a wide source of hyperimmune sera used in different approaches, including diagnosis and the study of host-parasite interactions. Unfortunately, mammalian antibodies present limitations for its production, such as the necessity for animal bleeding, low yield, interference with rheumatoid factor, complement activation and affinity to Fc mammalian receptors. IgY antibodies avoid those limitations; therefore they could be an alternative to be applied in T. gondii model.

Methodology/Principal Findings

In this study we immunized hens with soluble tachyzoite antigens of T. gondii (STAg) and purified egg yolk antibodies (IgY) by an inexpensive and simple method, with high yield and purity degree. IgY anti-STAg antibodies presented high avidity and were able to recognize a broad range of parasite antigens, although some marked differences were observed in reactivity profile between antibodies produced in immunized hens and mice. Interestingly, IgY antibodies against Neospora caninum and Eimeria spp. did not react to STAg. We also show that IgY antibodies were suitable to detect T. gondii forms in paraffin-embedded sections and culture cell monolayers.

Conclusions/Significance

Due to its cost-effectiveness, high production yield and varied range of possible applications, polyclonal IgY antibodies are useful tools for studies involving T. gondii.     

MUC4 and MUC1 Expression in Adenocarcinoma of the Stomach Correlates with Vessel Invasion and Lymph Node Metastasis: An Immunohistochemical Study of Early Gastric Cancer

We have previously reported that MUC4 expression is a poor prognostic factor in various carcinomas. Our previous study also showed that MUC1 expression in gastric cancers, including the early and advanced stages is a poor prognostic factor. In the present study, the expression profiles of MUC4 and MUC1 were examined by immunohistochemistry (IHC) using two anti-MUC4 monoclonal antibodies (MAbs), 8G7 and 1G8, and anti-MUC1 MAb DF3 in 104 gastrectomy specimens of early gastric adenocarcinoma with submucosal invasion (pT1b2), including 197 histological subtype lesions. Before the IHC study of the human specimens, we evaluated the specificity of the two MAbs by Western blotting and IHC of two MUC4 mRNA expressing gastric cancer cell lines. MAb 8G7 reacted clearly, whereas MAb 1G8 did not show any reactivity, in either Western blotting or IHC. In the IHC of the gastric cancers, the expression rates of MUC4/8G7 detected by MAb 8G7, MUC4/1G8 detected by MAb 1G8 and MUC1/DF3 detected by MAb DF3 in well differentiated types (70%, 38/54; 67%, 36/54; 52%, 28/54) were significantly higher than those in poorly differentiated types (18%, 10/55; 36%, 20/55; 13%, 7/55) (P<0.0001; P = 0.0021; P<0.0001), respectively. The MUC4/8G7 expression was related with lymphatic invasion (r = 0.304, P = 0.033). On the other hand, the MUC4/1G8 expression was related with lymphatic invasion (r = 0.395, P = 0.001) and lymph node metastasis (r = 0.296, P = 0.045). The MUC1/DF3 expression was related with lymphatic invasion (r = 0.357, P = 0.032) and venous invasion (r = 0.377, P = 0.024). In conclusion, the expression of MUC4 as well as MUC1 in early gastric cancers is a useful marker to predict poor prognostic factors related with vessel invasion.     

A population model for diapausing multivoltine insects under asymmetric cannibalism

Pupa-eating cannibalism occurs naturally in several insect species. Byasa alcinous is a multivoltine species of Red-bodied Swallowtail butterfly found in East Asia, which diapauses as pupa over the winter and whose larvae cannibalize eggs and pupae. We investigate the effects on population dynamics of increasing the asymmetric cannibalistic attack rate of a general insect species in different environmental conditions. We do this by theoretically formulating a generalized system of univoltine and bivoltine larvae over two generations in the spring and summer months. We predict that a lack of resources over the summer can force the population to become entirely univoltine, unless the second-generation bivoltine larvae increase their cannibalistic attack rate, and consume the diapausing pupae from the first generation. The model shows that under extreme environmental conditions, the persistence of univoltine larvae is favoured when faced with the threat of extinction. The model also predicts the conditions for the coexistence of both univoltine and bivoltine larvae, and the degree to which they can both coexist, which decreases as the resource in the second generation increases. This work provides the grounding for future theoretical and experimental consideration of the role of cannibalism in determining insect voltinism.     

3'-Azido-2',3'-dideoxynucleoside 5'-triphosphates inhibit telomerase activity in vitro, and the corresponding nucleosides cause telomere shortening in human HL60 cells

Telomerase adds telomeric DNA repeats to the ends of linear chromosomal DNA. 3′-Azido-3′-deoxythymidine 5′-triphosphate (AZTTP) is a known telomerase inhibitor. To obtain more selective and potent inhibitors that can be employed as tools for studying telomerase, we investigated the telomerase-inhibitory effects of purine nucleosides bearing a 3′-down azido group: 3′-azido-2′,3′-dideoxyguanosine (AZddG) 5′-triphosphate (AZddGTP), 3′-azido-2′,3′-dideoxy-6-thioguanosine (AZddSG) 5′-triphosphate (AZddSGTP), 3′-azido-2′,3′-dideoxyadenosine (AZddA) 5′-triphosphate (AZddATP) and 3′-azido-2′,3′-dideoxy-2-aminoadenosine (AZddAA) 5′-triphosphate (AZddAATP). Of these, AZddGTP showed the most potent inhibitory activity against HeLa cell telomerase. AZddGTP was significantly incorporated into the 3′-terminus of DNA by partially purified telomerase. However, AZddGTP did not exhibit significant inhibitory activity against DNA polymerases α and δ, suggesting that AZddGTP is a selective inhibitor of telomerase.

We also investigated whether long-term treatment with these nucleosides could alter telomere length and growth rates of human HL60 cells in culture. Southern hybridization analysis of genomic DNA prepared from cells cultured in the presence of AZddG and AZddAA revealed reproducible telomere shortening.

     

Spatial pattern analysis in forest dynamics: deviation from power law and direction of regeneration waves

Grid-based models have been used to understand spatial heterogeneity of the vegetation height in forests and to analyze spatio-temporal dynamics of the forest regeneration process. In this report, we present two methods of identifying lattice models when spatio-temporal data are given. The first method detects directionality of regeneration waves based on the timing of local disturbance events. The second evaluates the forest pattern by recording the fraction of high and low vegetation areas at multiple spatial scales. We illustrate these methods by applying them to patterns generated using three simple stochastic lattice models: (1) two-state model, distinguishing sites with high and low vegetation, (2) three-state model, in which sites can be in an additional disturbed state, and (3) Shimagare model, which considers a continuous range of states. The combination of the two methods provides efficient means of distinguishing the models. The first method has a more direct ecological meaning, while the second is useful when forest data are limited in time.     

Toxoplasma gondii 70 kDa Heat Shock Protein: Systemic Detection Is Associated with the Death of the Parasites by the Immune Response and Its Increased Expression in the Brain Is Associated with Parasite Replication

The heat shock protein of Toxoplasma gondii (TgHSP70) is a parasite virulence factor that is expressed during T. gondii stage conversion. To verify the effect of dexamethasone (DXM)-induced infection reactivation in the TgHSP70-specific humoral immune response and the presence of the protein in the mouse brain, we produced recombinant TgHSP70 and anti-TgHSP70 IgY antibodies to detect the protein, the specific antibody and levels of immune complexes (ICs) systemically, as well as the protein in the brain of resistant (BALB/c) and susceptible (C57BL/6) mice. It was observed higher TgHSP70-specific antibody titers in serum samples of BALB/c compared with C57BL/6 mice. However, the susceptible mice presented the highest levels of TgHSP70 systemically and no detection of specific ICs. The DXM treatment induced increased parasitism and lower inflammatory changes in the brain of C57BL/6, but did not interfere with the cerebral parasitism in BALB/c mice. Additionally, DXM treatment decreased the serological TgHSP70 concentration in both mouse lineages. C57BL/6 mice presented high expression of TgHSP70 in the brain with the progression of infection and under DXM treatment. Taken together, these data indicate that the TgHSP70 release into the bloodstream depends on the death of the parasites mediated by the host immune response, whereas the increased TgHSP70 expression in the brain depends on the multiplication rate of the parasite.