PURPOSE: We developed a new artificial image enhancement system and evaluated its usefulness in controlling intraoperative reflection and enhancing of Brilliant Blue G (BBG) staining. METHODS: The system was composed of three kinds of filters (a polarizing filter, a blue-enhancing filter, and a sharp-cut filter Y) and attached to the inferior surface of the operating microscope. Twenty-seven post-mortem extracted porcine eyes were used for a series of examinations. We performed surgery using the 23G-vitrectomy system with a halogen light and xenon lights and compared the reduction of intraoperative reflection under air condition and visibility and BBG contrast with and without this system. The evaluation of images was calculated in CIE 1976 (L*, a*, b*) color space (CIELAB) carried out by ImageJ software. The transmission of each filter and absorbance of BBG was measured by a spectrophotometer. We measured spectral irradiance at each wavelength about each filter from each light source with a spectroradiometer. RESULTS: Under both light sources, intraoperative reflection was controlled using a polarizing (PL) filter or combination of filters under air condition. Evaluation of the value of L* within the cutter surface was changed by 37.8 % under the halogen light, and 61.6 %(averaged) under the xenon light with inserted filters versus no filter. The BBG intensity difference was obtained with sharp-cut Y filter under both light source and PL with blue enhancing filter under the halogen light using each L*, a*, b* parameter with statistically significant (p < 0.01, 0.05). However, there was a relative decrease in the observation illuminance when the filter inserted according to the attenuation total spectral irradiance. CONCLUSIONS: This system can reduce intraoperative reflections under the air condition and obtain an excellent BBG staining intensity induced by various light sources.

BACKGROUND The targeting of Ca(2+) cycling has emerged as a potential therapy for the treatment of severe heart failure. These approaches include gene therapy directed at overexpressing sarcoplasmic reticulum (SR) Ca(2+) ATPase, or ablation of phospholamban (PLN) and associated protein phosphatase 1 (PP1) protein complexes. We previously reported that PP1β, one of the PP1 catalytic subunits, predominantly suppresses Ca(2+) uptake in the SR among the three PP1 isoforms, thereby contributing to Ca(2+) downregulation in failing hearts. In the present study, we investigated whether heart-failure-inducible PP1β-inhibition by adeno-associated viral-9 (AAV9) vector mediated gene therapy is beneficial for preventing disease progression in genetic cardiomyopathic mice. METHODS We created an adeno-associated virus 9 (AAV9) vector encoding PP1β short-hairpin RNA (shRNA) or negative control (NC) shRNA. A heart failure inducible gene expression system was employed using the B-type natriuretic protein (BNP) promoter conjugated to emerald-green fluorescence protein (EmGFP) and the shRNA sequence. AAV9 vectors (AAV9-BNP-EmGFP-PP1βshRNA and AAV9-BNP-EmGFP-NCshRNA) were injected into the tail vein (2×10(11) GC/mouse) of muscle LIM protein deficient mice (MLPKO), followed by serial analysis of echocardiography, hemodynamic measurement, biochemical and histological analysis at 3 months. RESULTS In the MLPKO mice, BNP promoter activity was shown to be increased by detecting both EmGFP expression and the induced reduction of PP1β by 25% in the myocardium. Inducible PP1βshRNA delivery preferentially ameliorated left ventricular diastolic function and mitigated adverse ventricular remodeling. PLN phosphorylation was significantly augmented in the AAV9-BNP-EmGFP-PP1βshRNA injected hearts compared with the AAV9-BNP-EmGFP-NCshRNA group. Furthermore, BNP production was reduced, and cardiac interstitial fibrosis was abrogated at 3 months. CONCLUSION Heart failure-inducible molecular targeting of PP1β has potential as a novel therapeutic strategy for heart failure.

Oshiro A, Iseki S, Miyauchi M, Terashima T, Kawaguchi Y, Ikeda Y, Shinomura T. Lipopolysaccharide induces rapid loss of follicular dendritic cell-secreted protein in the junctional epithelium. J Periodont Res 2012; doi: 10.1111/j.1600-0765.2012.01482.x . © 2012 John Wiley & Sons A/S Background and Objective:  We have previously reported that mRNA encoding follicular dendritic cell-secreted protein (FDC-SP) is expressed specifically in the junctional epithelium at the gingival crevice. Other tissues, such as tonsil, prostate gland and trachea, also express high levels of FDC-SP. These tissues participate in a range of functions closely related to innate immunity. Therefore, it is hypothesized that FDC-SP plays a crucial role in close association with the host defense system within the gingival crevice. Accordingly, the main aim of this study was to investigate the expression and localization of FDC-SP in and around the junctional epithelium and to observe the dynamic changes of FDC-SP in experimental inflammation. Material and Methods:  We examined, immunohistochemically, the expression of FDC-SP in the junctional epithelium using a specific antibody raised in rabbit after immunization with a synthetic peptide derived from the hydrophilic region of FDC-SP. Experimental inflammation was induced in the upper molars of Wistar rats by applying bacterial lipopolysaccharide (LPS; 5 mg/mL in sterile saline) for 1 h. Results:  We confirmed that FDC-SP is present in the junctional epithelium in a pattern that is consistent with the expression of FDC-SP mRNA. Of special interest is that no FDC-SP was detectable in the junctional epithelium 3 h after transient topical treatment with LPS. Conclusion:  The presence of FDC-SP in the junctional epithelium and its loss after LPS treatment strongly support our hypothesis of FDC-SP playing a crucial role in close association with the host defense system within the gingival crevice.

More than two decades have passed since genetically modified HIV was used for gene delivery. Through continuous improvements these early marker gene-carrying HIVs have evolved into safer and more effective lentiviral vectors. Lentiviral vectors offer several attractive properties as gene-delivery vehicles, including:(i) sustained gene delivery through stable vector integration into host genome;(ii) the capability of infecting both dividing and non-dividing cells;(iii) broad tissue tropisms, including important gene- and cell-therapy-target cell types;(iv) no expression of viral proteins after vector transduction;(v) the ability to deliver complex genetic elements, such as polycistronic or intron-containing sequences;(vi) potentially safer integration site profile; and (vii) a relatively easy system for vector manipulation and production. Accordingly, lentivector technologies now have widespread use in basic biology and translational studies for stable transgene overexpression, persistent gene silencing, immunization, in vivo imaging, generating transgenic animals, induction of pluripotent cells, stem cell modification and lineage tracking, or site-directed gene editing. Moreover, in the present high-throughput '-omics' era, the commercial availability of premade lentiviral vectors, which are engineered to express or silence genome-wide genes, accelerates the rapid expansion of this vector technology. In the present review, we assess the advances in lentiviral vector technology, including basic lentivirology, vector designs for improved efficiency and biosafety, protocols for vector production and infection, targeted gene delivery, advanced lentiviral applications and issues associated with the vector system.

Androgens, the male sex hormones, exert various biological effects on many target organs through the transcriptional effects of the nuclear androgen receptor (AR). ARs are expressed not only in classical target organs, such as the brain, genital organs, bone, and skeletal muscles, but also in the cardiovascular system. Because the female sex hormones estrogens are well known to protect against cardiovascular disease, sex has been considered to have a significant clinical impact on cardiovascular mortality. However, the influence of androgens on the cardiovascular system has not been fully elucidated. To clarify this issue, we analyzed the effects of administration of angiotensin II and doxorubicin, an anticancer agent, in a loading model in male wild-type and AR-deficient mice. In this review, we focus on the actions of androgens as potential targets for the prevention of cardiovascular diseases in males.

Most of inherited retinal diseases such as retinitis pigmentosa (RP) cause photoreceptor cell death resulting in blindness. RP is a large family of diseases in which the photoreceptor cell death can be caused by a number of pathways. Among them, light exposure has been reported to induce photoreceptor cell death. However, the detailed mechanism by which photoreceptor cell death is caused by light exposure is unclear. In this study, we have shown that even a mild light exposure can induce ectopic phototransduction and result in the acceleration of rod photoreceptor cell death in some vertebrate models. In ovl, a zebrafish model of outer segment deficiency, photoreceptor cell death is associated with light exposure. The ovl larvae show ectopic accumulation of rhodopsin and knockdown of ectopic rhodopsin and transducin rescue rod photoreceptor cell death. However, knockdown of phosphodiesterase, the enzyme that mediates the next step of phototransduction, does not. So, ectopic phototransduction activated by light exposure, which leads to rod photoreceptor cell death, is through the action of transducin. Furthermore, we have demonstrated that forced activation of adenylyl cyclase in the inner segment leads to rod photoreceptor cell death. For further confirmation, we have also generated a transgenic fish which possesses a human rhodopsin mutation, Q344X. This fish and rd10 model mice show photoreceptor cell death caused by adenylyl cyclase. In short, our study indicates that in some RP, adenylyl cyclase is involved in photoreceptor cell death pathway; its inhibition is potentially a logical approach for a novel RP therapy.

Our aim was to explore important inflammatory mediators for synovial chondromatosis in the temporomandibular joints (TMJs) by analysing synovial fluid. Samples were collected from 10 patients with unilateral synovial chondromatosis of the TMJ. Control samples were obtained from 11 subjects with no symptoms in the TMJ. Concentrations of aggrecan, interleukin (IL)-2, IL-4, IL-5, IL-6, IL-8 (CXCL8), IL-10, interferon (IFN)-γ, tumour necrosis factor (TNF)-α and vascular endothelial growth factor (VEGF)-A were measured in the samples of synovial fluid, and the results in the two groups compared. The tissues from the affected TMJ were examined histologically and immunohistochemically. Of the proteins evaluated, the concentrations of aggrecan, IL-6, and VEGF-A were significantly higher in the group with synovial chondromatosis. The immunohistochemical analysis showed that the synovial cells around the osteocartilaginous nodules were vigorously expressing VEGF-A. IL-6 and VEGF-A are thought to have important roles in the pathology of synovial chondromatosis of the TMJ.

The orphan nuclear receptor steroidogenic factor 1 (SF-1) is expressed in both Sertoli and Leydig cells in testes. The present study investigates postnatal development of the testes of a gonad-specific Sf-1 knockout (KO) mouse, in which Sf-1 was specifically inactivated. The KO testes appeared histologically normal from postnatal day (P) 0 until P7. However, disorganized germ cells, vacuoles, and giant cells appeared by P14 in the seminiferous tubules of KO, but not control mice. Expression of SF-1 and various factors was examined by immunohistochemistry (IHC). The number of SF-1-positive Sertoli cells in the KO testes was lower compared to controls at all the developmental stages, and decreased to nearly undetectable levels by P21. IHC for AMH and p27, immature and mature Sertoli cell markers respectively, indicated a delay of Sertoli cell maturation in the KO testes. The number of Sertoli cells expressing factors involved in Sertoli cell differentiation including WT1, SOX9, GATA4, and androgen receptor, were lower in the KO testes compared to controls. Furthermore, fewer PCNA-positive proliferative germ cells were observed, and the number of TUNEL-labeled cells was significantly higher in the KO testes compared to controls at P14 and P21, indicating impaired spermatogenesis. IHC for SCC indicated a presence of steroidogenic Leydig cells in the interstitium of the KO testes at all stages examined. These results suggest that SF-1 is essential for Sertoli cell maturation and therefore spermatogenesis, during postnatal testis development.

Background: Eltrombopag is an oral, nonpeptide thrombopoietin receptor agonist that has shown efficacy and safety in chronic immune thrombocytopenia (ITP). However, ethnic differences in eltrombopag exposure have been reported: area under the curve exposure to eltrombopag was 87% greater among ITP patients of East Asian descent compared to non-East Asian ITP patients. Objectives: In this study a lower starting (12.5 mg) and maximum (50 mg) dose of eltrombopag was used for Japanese ITP patients compared with the standard starting (50 mg) and maximum (75 mg) dose approved in the United States and Europe to evaluate its efficacy and safety. Patients: We examined 23 Japanese patients with previously treated chronic ITP with a platelet count <30,000/μL, in a multicenter study comprising a randomized, double-blind, placebo-controlled phase for 6-week evaluation (15 eltrombopag, 8 placebo) and an open-label phase for 6-month evaluation (23 eltrombopag). Results and Conclusions: The response rate (platelet count ≥50,000/μL) at week 6 of the 6-week double-blind phase was 60% in eltrombopag-treated patients and 0% in placebo-treated patients. Ten of 23 patients (43.5%) responded for ≥75% of predefined assessment visits during the 6-month open-label phase. Notably, 22%(5/23) of patients responded to 12.5 mg of eltrombopag, which was administered within the first 3 weeks of eltrombopag treatment. Bleeding decreased with eltrombopag treatment compared to baseline. Eltrombopag was generally well-tolerated; 1 patient experienced a transient ischemic attack on day 9. Eltrombopag (12.5 mg to 50 mg) is effective for the management of Japanese patients with chronic ITP (NCT00540423). © 2012 International Society on Thrombosis and Haemostasis.

Indoxyl sulfate (IS) is an organic anion uremic toxin which accumulates in chronic kidney disease (CKD) patients. The aims of this study were to examine the kinetic profiles of IS in humans at a steady state after multiple doses of L-tryptophan (Trp), a precursor of IS, and the in vivo interaction of IS with the angiotensin-converting enzyme inhibitor (ACEI) quinapril, whose active metabolite is a substrate of organic anion transporter 3 (OAT3), in rats. First, 12-h kinetics after the single doses of Trp (2, 4 and 8 g) were examined in two healthy volunteers. Second, 24-h kinetics after the single dose of 2 g Trp was studied in 6 volunteers. Third, 35-h kinetics after the single and multiple doses of 2 g Trp were examined in 5 volunteers. In anesthetized rats, quinapril or probenecid, an inhibitor of OATs, following IS was intravenously given and blood and urine were taken until 90 min. Trp and IS concentrations were determined by high performance liquid chromatography (HPLC). Ultrafiltration was used to measure serum unbound IS concentrations. Renal tubular secretion of IS accounted for more than 90% of its renal clearance in the steady state of serum IS levels after multiple doses in humans. In animals, the serum area under the curve (AUC) of IS increased in conjunction with a decrease in renal clearances after co-administration of IS with quinapril or probenecid. It is concluded that quinapril may inhibit the urine excretion of IS via OAT3-mediated renal tubular transport in CKD patients.