Distinct behavior of human Langerhans cells and inflammatory dendritic epidermal cells at tight junctions in patients with atopic dermatitis.

BACKGROUND: The stratum corneum and tight junctions (TJs) form physical barriers in the epidermis. Dendrites of activated Langerhans cells (LCs) extend beyond the TJs to capture external antigens in mice. LCs and inflammatory dendritic epidermal cells (IDECs) are observed in the skin of patients with atopic dermatitis (AD). OBJECTIVE: We sought to investigate the characteristics of LCs and IDECs and the distribution of their antigen capture receptors in relation to TJs in normal and AD skin. METHODS: We characterized the interactions of LCs and IDECs with TJs and the expression patterns of langerin and FcεRI by using whole-mount epidermal sheets from healthy subjects and patients with AD, ichthyosis vulgaris, and psoriasis vulgaris. RESULTS: As in mouse skin, activated LCs penetrate TJs in human skin. The number of LCs with TJ penetration increased approximately 5-fold in erythematous lesional skin of patients with AD but not in nonlesional skin of patients with AD or lesions of patients with ichthyosis vulgaris or psoriasis. In contrast, IDECs localized in the lower part of the epidermis, and their dendrites extended horizontally without penetration through TJs. Although langerin accumulated on the tips of dendrites of activated LCs, FcεRI was expressed diffusely on the cell surfaces on LCs and IDECs in lesional skin from patients with AD. CONCLUSIONS: These findings highlight interesting differences between LCs and IDECs in epidermis of patients with AD, where LCs, but not IDECs, extend dendrites through the TJs, likely to capture antigens from outside the TJ barrier with a polarized distribution of langerin but not FcεRI. These behavioral differences between skin dendritic cells might reflect an important pathophysiology of AD.

Dry eye disease is associated with deterioration of mental health in male Japanese university staff.

Dry eye disease (DED) is defined as a multifactorial disease of the tears and ocular surface that results in symptoms of discomfort, visual disturbance, and tear-film instability, with potential damage to the ocular surface. It is thought to be associated with reduced quality of life (QOL). The aim of the present study was to investigate the effects of DED on health-related QOL in Japanese university sedentary office workers who are daily users of visual display terminal. In this study, 163 university staff (99 male and 64 female), aged 23-69 years, served as study subjects. Subjects were asked to answer the following three questions. (1) How often do your eyes feel dry? (2) How often do your eyes feel irritated? (3) Have you ever been diagnosed by a clinician as having dry eye syndrome? Sixty-eight subjects who answered "constantly," "often", or "sometimes" to both questions 1 and 2 were classified as the DED Group, and the remaining 95 were defined as the Non-DED Group. QOL was assessed by the SF-36 questionnaire, which consisted of 36 items to produce three summary scores, namely, mental, physical, and role/social component summary scores. For males, the DED Group had significantly lower scores than the Non-DED Group for mental component summary (MCS) (P = 0.005). In multiple regression analysis, MCS scores were adversely related to DED in males (P = 0.015). DED was associated with worsened QOL. DED should be regarded as a factor that can lead to deterioration of mental health.

Preoperative and intraoperative assessment of myometrial invasion in endometrial cancer: comparison of magnetic resonance imaging and frozen sections.

OBJECTIVE: To compare the diagnostic characteristics of the evaluation of myometrial invasion (MI) retrospectively between preoperative magnetic resonance imaging (MRI) and intraoperative frozen sections. DESIGN: A retrospective study. SETTING: University hospital. SAMPLE: 201 women diagnosed with endometrial carcinoma. METHODS: All women underwent preoperative MRI and 111 of them also underwent intraoperative frozen section assessment. The final pathological evaluation was used as the definitive diagnosis. MAIN OUTCOME MEASURES: In women who underwent MRI and frozen sections (n = 111), the accuracies of detection of MI and of deep invasion (defined as ≥50% invasion) were compared. RESULTS: The accuracy, sensitivity, and specificity of MRI for detection of MI were 65.8, 58.8, and 88.5%, and those in frozen sections were 90.1, 90.6, and 88.5%, respectively. The accuracy and sensitivity of frozen sections were significantly higher (p < 0.001, p < 0.001), whereas the specificity of the two methods did not differ (p = 1.000). The accuracy, sensitivity, and specificity of MRI for detection of deep invasion were 83.8, 69.2, and 88.2%, and those of frozen sections were 93.7, 73.1, and 100.0%, respectively. The accuracy and specificity of frozen sections were significantly higher (p = 0.007 and p < 0.001, respectively), whereas sensitivity did not show a significant difference (p = 0.999). CONCLUSION: In assessment of MI, the accuracy of frozen sections was significantly higher than that of MRI. Since the diagnostic characteristics differ between two methods, additional intraoperative frozen sections are recommended for more accurate assessment of MI when MRI is negative for the presence of any MI or positive for the presence of deep invasion.

Rituximab therapy for refractory autoimmune bullous diseases: A multicenter, open-label, single-arm, phase 1/2 study on 10 Japanese patients.

This was a multicenter study of rituximab, a chimeric monoclonal immunoglobulin G antibody directed against CD20, for the treatment of refractory autoimmune bullous diseases (pemphigus and pemphigoid). Ten patients (three with pemphigus vulgaris, six with pemphigus foliaceus and one with bullous pemphigoid) were treated with a single cycle of rituximab (four weekly infusions at a dose of 375 mg/m2 of body surface area). The primary end-points were the number of serious adverse events and rate of complete remission at 40 weeks. Five patients (50%) achieved complete remission with minimal therapy (defined as no active lesions with lower doses of systemic corticosteroids compared to that with prednisolone 10 mg/day). Improvements in clinical scores (Pemphigus Disease Area Index) and decreases in autoantibody titers in the sera were observed in the four pemphigus patients who failed to achieve complete remission. This suggests that rituximab was effective in nine of 10 cases. Two serious adverse events (Pneumocystis carinii pneumonia and septic shock due to infectious arthritis) were observed and adequately treated with hospitalization. CD19-positive B lymphocytes in the peripheral blood decreased on day 29 following rituximab treatment, and remained at low levels throughout the observation period (280 days). Our results confirmed the efficacy of rituximab therapy for refractory autoimmune bullous diseases in Japan.

Characterization of centriole duplication in human epidermis, Bowen's disease, and squamous cell carcinoma.

BACKGROUND: Centrosomes contain two centrioles: a pre-existing mature centriole and a newly formed immature centriole. Each centriole is duplicated once within a cell cycle, which is crucial for proper centrosome duplication and cell division. OBJECTIVE: To describe the centrosome duplication cycle in human epidermis, Bowen's disease (BD), and squamous cell carcinoma (SCC). METHODS: Immunofluorescent staining of centriolar proteins and Ki-67 was used to evaluate cell cycles and the number of centrioles. Centrobin and Outer dense fiber of sperm tails 2 (ODF2) were used as markers for immature and mature centrioles, respectively. RESULTS: Normal human primary epidermal keratinocytes in a monolayered culture have one centrobin+ centriole (CTRB1+ cells) supposed in G0/G1 phases or have two centrobin+ centrioles (CTRB2+ cells) supposed in S-G2 phase. In a three-dimensional culture and in vivo human epidermis, the majority of suprabasal cells were CTRB2+ cells, in spite of their non-proliferative Ki-67- nature. The tumor mass of BD and SCC contained CTRB1+ cells and Ki-67+ proliferating and Ki-67- non-proliferative CTRB2+ cells. Clumping cells in BD had increased numbers of centrioles, with an approximate 1:1 to 2:1 ratio of centrobin+ to ODF2+ centrioles. CONCLUSIONS: The cell cycle arrest of suprabasal cells is distinct from the G0 arrest of monolayered epithelial cells. Tumor mass of BD and SCC contained non-proliferative cells with the characteristics of the suprabasal cells of normal epidermis. A constant ratio of the number of centrobin+ to ODF2+ centrioles indicates that multiple centrioles were induced by cell division failure rather than centriole overduplication in clumping cells.

Effects of laser in situ keratomileusis on mental health-related quality of life.

PURPOSE: The aims of our study were to investigate whether laser in situ keratomileusis (LASIK) improves health-related quality of life (HRQoL) and to identify factors that affect postoperative HRQoL. MATERIALS AND METHODS: A total of 213 Japanese patients who underwent primary LASIK were analyzed in this study. The average age of patients was 35.0±9.4 years. The subjects were asked to answer questions regarding subjective quality of vision, satisfaction, and quality of life (using the Japanese version of 36-Item Short Form Health Survey Version 2) at three time points: before LASIK, 1 month after LASIK, and 6 months after LASIK. Longitudinal changes over 6 months in the outputs of mental component summary (MCS) score and the physical component summary (PCS) score from the 36-Item Short Form Health Survey Version 2 questionnaire were compared between time points using a linear mixed-effects model. Delta MCS and PCS were calculated by subtracting the postoperative score (1 month after LASIK) from the preoperative score. Preoperative and postoperative factors associated with a change in the MCS score or PCS score were evaluated via a linear regression model. RESULTS: The preoperative MCS score was 51.0±9.4 and increased to 52.0±9.8 and 51.5±9.6 at 1 month and 6 months after LASIK, respectively, and the trend for the change from baseline in MCS through 6 months was significant (P=0.03). PCS score did not change following LASIK. Delta MCS was significantly negatively associated with preoperative spherical equivalent, axial length, and postoperative quality of vision, after adjusting for potential confounding factors. CONCLUSION: Mental HRQoL is not lost with LASIK, and LASIK may improve mental HRQoL. Preoperative axial length may predict postoperative mental HRQoL.

Analysis of neurons created from wild-type and Alzheimer's mutation knock-in embryonic stem cells by a highly efficient differentiation protocol.

It is impossible to obtain and amplify live neurons from Alzheimer's disease (AD) patients. To establish the neurons harboring AD abnormality, we constructed mouse embryonic stem (ES) cells, in which the AD-causative V642I mutation was introduced to the endogenous amyloid precursor protein (APP) gene, in combination with a protocol to efficiently differentiate ES cells into postmitotic neurons without using a cell sorter. By this protocol, ES cells differentiated into >90% of the central type of adult postmitotic neurons. Neurons derived from V642I-APP knock-in ES cells were indistinguishable from wild-type ES-derived neurons, as determined by the expression of various markers for neuronal differentiation. Notably, V642I-APP knock-in ES cell-derived neurons exhibited significantly increased secretion of Abeta42 without AD-related hyperphosphorylation of tau, indicating that the direct output of the AD-causative mutation is increased Abeta42 secretion. In this study, we analyze created neurons with wild-type and AD genotypes and propose a new strategy for generating neurons for any dominantly inherited neurodegenerative diseases. The strategy can be applied to create human neurons with AD or any other neurodegenerative disease by using human ES cells.

Glaucomatous visual field defect severity and the prevalence of motor vehicle collisions in Japanese: a hospital/clinic-based cross-sectional study.

Purpose. This study examined the association between the severity of visual field defects and the prevalence of motor vehicle collisions (MVCs) in subjects with primary open-angle glaucoma (POAG). Methods. This is a cross-sectional study. Japanese patients who have had driver's licence between 40 and 85 years of age were screened for eligibility. Participants answered a questionnaire about MVCs experienced during the previous 5 years. Subjects with POAG were classified as having mild, moderate, or severe visual field defect. We evaluated associations between the severity of POAG and the prevalence of MVCs by logistic regression models. Results. The prevalence of MVCs was significantly associated with the severity of POAG categorized by worse eye MD (control: 30/187 = 16.0%; mild POAG: 17/92 = 18.5%; moderate POAG: 14/60 = 23.3%; severe POAG: 14/47 = 29.8%; P = 0.025, Cochran-Armitage trend test). Compared to the control group, the adjusted OR for MVC prevalence in subjects with mild, moderate, or severe POAG in the worse eye was 1.07 (95% CI: 0.55 to 2.10), 1.44 (95% CI: 0.68 to 3.08), and 2.28 (95% CI: 1.07 to 4.88). Conclusions. There is a significant association between the severity of glaucoma in the worse eye MD and the prevalence of MVCs.

Risk factors for decreased bone mineral density in inflammatory bowel disease: A cross-sectional study.

BACKGROUND & AIM: Although inflammatory bowel disease (IBD) patients are at risk for metabolic bone disease, studies analyzing this correlation have identified various risk factors, including disease phenotype, age, sex and steroid therapy. Furthermore, few studies have assessed risk factors for bone loss in Japanese IBD patients. This study analyzed risk factors for metabolic bone disease in Japanese IBD patients. METHODS: This cross-sectional study assessed 388 patients with IBD aged 20-50 years, including 232 with ulcerative colitis (UC) and 156 with Crohn's disease (CD). Bone mineral density of the femoral neck, total femur and lumbar spine was quantified by dual-energy X-ray absorptiometry. The blood concentrations of bone metabolism markers were measured. History of smoking and bone fracture, and nutritional intake were assessed using questionnaires. RESULTS: Of the 388 patients with IBD, 78 (20.1%; UC, 17.2%; CD, 24.4%) had osteopenia and 17 (4.4%; UC, 3.4%; CD, 5.8%) had osteoporosis, as assessed by T-score. Bone mineral density of the lumbar vertebrae was lower in males than in females. Multivariate regression analysis showed that risk factors for bone loss in UC patients were male sex, low body mass index (BMI), high steroid dose and disease location. Risk factors for bone loss in CD patients were male sex and low BMI. CONCLUSION: Among Japanese patients with IBD, male sex and low BMI were associated with increased risk for metabolic bone disease. In addition, Steroid therapy shouldn't be indiscriminate in UC patients. These findings may help identify patients at particularly high risk of metabolic bone disease and may help implement appropriate therapies in a timely manner and improve long-term quality of life.

Postoperative flat anterior chamber: incidence, risk factors, and effect on the long-term success of trabeculectomy.

PURPOSE: To investigate the long-term effects of postoperative flat anterior chamber (FAC) development on outcomes following trabeculectomy with mitomycin C. METHODS: This was a retrospective cohort study. Data on 383 consecutive patients (383 eyes) who underwent trabeculectomy at our institution between 1999 and 2009 were followed up. Patients who developed FAC after trabeculectomy and patients with maintained anterior chamber were evaluated. The primary outcome variable was the success of the initial trabeculectomy, which was defined at 3 different levels by the achievement of the following intraocular pressure (IOP)-related criteria without secondary IOP-lowering surgery: (a) IOP ≤ 12 mmHg and ≤30 % reduction in IOP from the preoperative level; (b) IOP ≤ 16 mmHg and ≤20 % reduction in IOP; and (c) IOP ≤ 21 mmHg. The hazard ratios (HRs) for the failure of trabeculectomy caused by FAC within 5 years of surgery were examined in conjunction with the Cox proportional hazards regression model. RESULTS: FAC was observed in 90 of the 383 eyes examined (23.4 %). Postoperative mild FAC was associated with the long-term success of trabeculectomy when evaluated according to our strictest success criterion, Criterion-A [HR = 0.72 (95 % CI 0.53-0.98); P = 0.04]. In contrast, severe FAC was inversely associated with the long-term success of the surgery when evaluated according to our most lenient criterion, Criterion-C [HR = 1.93 (95 % CI 1.16-3.22); P = 0.01]. CONCLUSION: Mild postoperative FAC after trabeculectomy is associated with a favorable long-term outcome, whereas severe postoperative FAC leads to an unfavorable prognosis.

The association between visual field defect severity and fear of falling in primary open-angle glaucoma.

PURPOSE: To determine if glaucomatous visual field defect severity is associated with fear of falling. METHODS: This is a cross-sectional study. A total of 387 consecutive subjects with POAG were enrolled in this study along with 293-ocular disease-free control subjects, who were screened at the same institutions. We defined mild POAG as MD of -6 dB or better, moderate POAG as MD of -6 to -12 dB, and severe POAG as MD of -12 dB or worse in the better eye. All participants were requested to answer a questionnaire on fear of falling. Associations between POAG severity and the prevalence of fear of falling were evaluated with the Cochran-Armitage trend test. Multivariable factors including age-adjusted odds ratios (ORs) for the prevalence of fear of falling and 95% confidence intervals (CIs) were evaluated with logistic regression models. RESULTS: The prevalence of fear of falling was 35/293 (11.9%) in the control group, 38/313 (12.1%) in the mild POAG group, 12/48 (25.0%) in the moderate POAG group, and 6/26 (23.1%) in the severe POAG group, and the trend was statistically significant (P = 0.028 Cochran-Armitage trend test). The adjusted ORs for prevalence in the mild, moderate, and severe POAG groups compared with that in the control group were 1.44 (95% CI: 0.83-2.51), 2.33 (95% CI: 1.00-5.44), and 4.06 (95% CI: 1.39-11.90), respectively. CONCLUSIONS: Among patients with POAG, the severity of visual field defects is associated with fear of falling. (http://www.umin.ac.jp/ctr/index.htm number, UMIN000005574.).

Indocyanine green fluorescence imaging for evaluation of uterine blood flow in cynomolgus macaque.

BACKGROUND: Uterine blood flow is an important factor in uterine viability, but the number of blood vessels required to maintain viability is uncertain. In this study, indocyanine green (ICG) fluorescence imaging was used to examine uterine hemodynamics and vessels associated with uterine blood flow in cynomolgus macaque. METHODS: The uterus of a female cynomolgus macaque was cut from the vaginal canal to mimic a situation during trachelectomy or uterine transplantation surgery in which uterine perfusion is maintained only with uterine and ovarian vessels. Intraoperative uterine hemodynamics was observed using ICG fluorescence imaging under conditions in which various nutrient vessels were selected by clamping of blood vessels. A time-intensity curve was plotted using imaging analysis software to measure the T(max) of uterine perfusion for selected blood vessel patterns. Open surgery was performed with the uterus receiving nutritional support only from uterine vessels on one side. The size of the uterus after surgery was monitored using transabdominal ultrasonography. RESULTS: The resulting time-intensity curves displayed the average intensity in the regions of the uterine corpus and uterine cervix, and in the entire uterus. Analyses of the uterine hemodynamics in the cynomolgus macaque showed that uterine vessels were significantly related to uterine perfusion (P=0.008), whereas ovarian vessels did not have a significant relationship (P=0.588). When uterine vessels were clamped, ovarian vessels prolonged the time needed to reach perfusion maximum. Postoperative transabdominal ultrasonography showed that the size of the uterus was not changed 2 months after surgery, with recovery of periodic menstruation. The cynomolgus macaque has got pregnant with favorable fetus well-being. CONCLUSION: Uterine vessels may be responsible for uterine blood flow, and even one uterine vessel may be sufficient to maintain uterine viability in cynomolgus macaque. Our results show that ICG fluorescence imaging is useful for evaluation of uterine blood flow since this method allows real-time observation of uterine hemodynamics.

The association between primary open-angle glaucoma and motor vehicle collisions.

PURPOSE: To investigate and compare the prevalence of motor vehicle collisions (MVCs) in individuals with or without primary open-angle glaucoma (POAG). METHODS: A total of 265 subjects were consecutively enrolled: 121 (79 men, 42 women; age, 62.1 ± 8.0 years) with POAG; and 144 (95 men, 49 women; age, 61.2 ± 7.9 years) who were free of ocular disease. Participants answered a questionnaire on MVC experience during the previous 10 years, past driving experience, and daily driving habits. The POAG group was subdivided into three groups according to disease severity (mild, moderate, or severe), to assess the relationship between POAG severity and MVC. RESULTS: A statistically significant association between POAG severity and MVC frequency was observed; 3.5% of the controls, 0.0% of the mild POAG group, 3.9% of the moderate POAG group, and 25.0% of the severe POAG group had experienced MVCs (P = 0.007, Cochran-Armitage trend test). The severe POAG group had experienced a much higher frequency of MVCs during the surveyed period than had the control group (P < 0.010; Fisher's exact test). Logistic regression analyses to account for confounding factors (age, presence of diabetes mellitus, driving history, time spent driving per day, and best corrected visual acuity in the better or worse eye) produced consistent results. CONCLUSIONS: Advanced POAG with marked visual field defects may be a risk factor for MVCs.

Narrow band imaging hysteroscopy: a comparative study using randomized video images.

Narrow band imaging (NBI) has been used in the gastrointestinal endoscopy field as a novel endoscopic imaging technique and has contributed to improved qualitative diagnosis and detection of lesions. However, there are only a few studies of use of NBI in the gynecology field. We applied NBI in flexible hysteroscopy at our outpatient clinic and evaluated the utility of NBI hysteroscopy for diagnosis of malignant endometrial lesions by comparison of the sensitivity and specificity between white light alone and white light+NBI using hysteroscopic video images. The subjects were 65 patients with a suspected endometrial lesion in the uterine cavity. These patients underwent flexible hysteroscopy using NBI in addition to conventional white light. Video images from 65 patients were edited into two groups, white light alone (WL group) and white light+NBI (NBI group) (130 images in total). Computerized block randomization of the order was then performed. Four raters independently diagnosed the images without use of other clinical information. Using the pathological diagnosis as the gold standard, we evaluated the sensitivity and specificity of diagnosis of atypical endometrial hyperplasia (AEH) or endometrial carcinoma compared between the WL and NBI groups. The sensitivity of diagnosis of AEH or endometrial carcinoma was numerically higher in the NBI group for all raters, and the average sensitivity was significantly higher in the NBI group compared to the WL group (78.6% vs. 63.7%, P<0.001). The specificity for each rater and the average specificity were comparable between the two groups. Compared to white light hysteroscopy, NBI hysteroscopy had a higher sensitivity for diagnosis of AEH or endometrial carcinoma without loss of specificity. This suggests that NBI hysteroscopy may be more useful than white light hysteroscopy for endoscopic diagnosis of malignant endometrial lesions.

Flexible hysteroscopy with narrow band imaging (NBI) for endoscopic diagnosis of malignant endometrial lesions.

Narrow band imaging (NBI) for detection of blood vessels and microstructures on the mucosal surface is used in gastrointestinal endoscopy since it can improve qualitative diagnosis and detection of lesion. However, there are no studies on flexible hysteroscopy using NBI. We performed flexible hysteroscopy with NBI for outpatients to investigate the sensitivity and specificity of endoscopic diagnosis of malignant endometrial lesions. Of patients who attended our hospital for suspected lesions in the uterine cavity between April 2009 and May 2010, 104 subjects underwent hysteroscopy with NBI, in addition to white light. Using the pathological diagnosis as the gold-standard, we evaluated the sensitivity and specificity of NBI hysteroscopy for detecting atypical endometrial hyperplasia (AEH) or carcinoma. The results were also compared with historical data (n=209) for conventional hysteroscopy using white light only in 2008. The sensitivities were 97.2% [95% confidence interval (95% CI): 90.3-99.7%] and 82.6% (95% CI: 74.4-89.0%) for NBI hysteroscopy and conventional hysteroscopy, respectively. The 95% CIs for the two methods did not overlap and the sensitivity of lesion detection was higher with NBI hysteroscopy. Specificities were comparable, 90.6% (95% CI: 75.0-98.0%) and 85.1% (95% CI: 76.3-91.6%) between the methods. NBI hysteroscopy has increased sensitivity for detection of atypical endometrial hyperplasia (AEH) or carcinoma. A comparison with historical data suggested that NBI may be useful for diagnosis of malignant endometrial lesions. As far as we are aware, this is the first evaluation of flexible hysteroscopy with NBI for diagnosis of malignant endometrial lesions.

A real-time three-dimensional echocardiographic quantitative analysis of left atrial function in left ventricular diastolic dysfunction.

The evaluation of left ventricular diastolic function provides important information about hemodynamics and has prognostic implications for various cardiac diseases. In particular, left atrial (LA) volume is an increasingly significant prognostic biomarker for diastolic dysfunction. The aim of this study was to assess left ventricular diastolic function by measuring changes in LA volume using real-time 3-dimensional echocardiography. The 106 subjects were divided into 4 groups (normal, impaired relaxation, pseudonormal, and restrictive) on the basis of diastolic function, as assessed by transmitral flow patterns. LA volume was measured during a heart cycle using real-time 3-dimensional echocardiography. LA stroke volume (maximum LA volume - minimum LA volume) and the LA ejection fraction (LA stroke volume/maximum LA volume x 100) were calculated using Doppler imaging to assess their correlation with other parameters used to evaluate left ventricular diastolic function, including transmitral flow pattern and early diastolic mitral annular velocity (E'). LA volume indexed to body surface area was dilated in subjects with left ventricular diastolic dysfunction, whereas the LA ejection fraction was lower. The maximum LA volume, minimum LA volume, and LA ejection fraction were significantly different between each group, and each was significantly correlated with the ratio of early diastolic transmitral flow velocity (E) to E' (E/E'). The LA ejection fraction correlated best with E/E' (r = -0.68, p <0.0001). In conclusion, cyclic changes in LA volume could be measured using real-time 3-dimensional echocardiography, and measuring LA function with this method may be a viable alternative for the accurate assessment of left ventricular diastolic function.

A humanin derivative, S14G-HN, prevents amyloid-beta-induced memory impairment in mice.

Humanin (HN) is a 24-amino acid peptide that protects neuronal cells from death caused by Alzheimer's disease (AD)-related genes and amyloid-beta (Abeta). Multiple studies have revealed its biochemical and neuroprotective characteristics in vitro; however, little has been known regarding whether HN is effective in vivo in AD model systems. We examined the effect of S14G-HN, a 1,000-fold more potent derivative of HN in vitro, on amnesia induced by Abeta25-35 in mice. The Y-maze test revealed that at least 50 pmol of S14G-HN by intracerebroventricular injection prevented Abeta-induced impairment of short-term/spatial working memory; however, 5 nmol of S14A-HN, a neuroprotection-defective mutant in vitro, did not prevent Abeta-induced amnesia. These results are in agreement with the structure-function correlation shown previously in vitro. In the water-finding task, S14G-HN prevented prolongation of finding latency (the time to find water) observed in Abeta-amnesic mice, indicating that S14G-HN also blocked Abeta-induced impairment of latent learning. In accordance with these observations, immunohistochemical analysis showed that S14G-HN sustained the number of cholinergic neurons in the basal forebrain and the striata nearly to the normal level. Furthermore, genistein, a specific inhibitor of tyrosine kinases, blocked recovery from scopolamine-induced amnesia by S14G-HN, suggesting that certain tyrosine kinase(s) are involved in the inhibitory function of S14G-HN in vivo. Taking these findings together, we conclude that S14G-HN has rescue activity against memory impairment caused by AD-related insults in vivo by activating the same intracellular neuroprotective machinery as elucidated previously in vitro.

p53-independent apoptosis is induced by the p19ARF tumor suppressor.

p19(ARF) is a potent tumor suppressor. By inactivating Mdm2, p19(ARF) upregulates p53 activities to induce cell cycle arrest and sensitize cells to apoptosis in the presence of collateral signals. It has also been demonstrated that cell cycle arrest is induced by overexpressed p19(ARF) in p53-deficient mouse embryonic fibroblasts, only in the absence of the Mdm2 gene. Here, we show that apoptosis can be induced without additional apoptosis signals by expression of p19(ARF) using an adenovirus-mediated expression system in p53-intact cell lines as well as p53-deficient cell lines. Also, in primary mouse embryonic fibroblasts (MEFs) lacking p53/ARF, p53-independent apoptosis is induced irrespective of Mdm2 status by expression of p19(ARF). In agreement, p19(ARF)-mediated apoptosis in U2OS cells, but not in Saos2 cells, was attenuated by coexpression of Mdm2. We thus conclude that there is a p53-independent pathway for p19(ARF)-induced apoptosis that is insensitive to inhibition by Mdm2.

Molecular characterization of neurohybrid cell death induced by Alzheimer's amyloid-beta peptides via p75NTR/PLAIDD.

One of the most important pathological features of Alzheimer's disease (AD) is extracellular senile plaques, whose major component is amyloid-beta peptides (Abeta). Abeta binds to the extracellular domain of p75NTR (p75 neurotrophin receptor) and induces neuronal cell death. We investigated the molecular mechanism of Abeta-induced neurotoxicity in detail from the standpoint of interaction between p75NTR and its recently identified relative, PLAIDD (p75-like apoptosis-inducing death domain). Using F11 neuronal hybrid cells, we demonstrate that there are two distinct pathways for Abeta-induced toxicity mediated by p75NTR. One pathway that has been previously elucidated, is mediated by p75NTR, Go, JNK, NADPH oxidase and caspase3-related caspases. We found that PLAIDD and Gi proteins, heterotrimeric G proteins, are involved in the alternative Abeta-induced neurotoxicity mediated by p75NTR. The alternative pathway triggered by Abeta is thus mediated by p75NTR, PLAIDD, Gi, JNK, NADPH oxidase and caspase3-related caspases. In addition, we found that HN, ADNF, IGF-I, or bFGF inhibits both pathways of Abeta-induced neurotoxicity mediated by p75NTR.

Targeted introduction of V642I mutation in amyloid precursor protein gene causes functional abnormality resembling early stage of Alzheimer's disease in aged mice.

While the exact aetiology of Alzheimer's disease (AD) is unknown, distinct genetic mutations have been identified for the rare cases of familial AD (FAD). V642I mutation in amyloid precursor protein (APP) co-segregates with FAD with perfect penetration, and the clinicopathological characteristics of patients with this mutation resemble that of sporadic AD. To examine the pathogenic process of this FAD-linked trait in vivo, we produced a mouse with the corresponding point mutation in the APP gene using homologous recombination and Cre-loxP site-specific recombination ('knock-in' technique). Mice with the heterozygous V642I-APP allele most precisely reflected the genotype of humans bearing this mutation. For the observation period of 2.5 years the mutants stayed apparently indistinguishable from the wild-type littermates. However, behavioural analysis revealed significantly deteriorated long-term memory in mutants when examined for the retention of spatial attention. Interestingly, acquisition of spatial memory was slightly affected but short-term working memory was not deteriorated at all. Histological examination was negative for formation of neuritic plaques or neurofibrillary tangles, whereas the relative amount of longer form of beta-amyloid species A beta 42(43) was significantly increased against that of the shorter form (A beta 40) in the mutant brain homogenates. We conclude that a V642I-APP mutant allele in aged mice confers functional components, but not organic components, of the AD-related phenotype that are observed in the early stage of AD. This V642I-APP knock-in mutant line may serve as a model to study the early pathogenic processes of AD in vivo and to develop therapeutics for this stage.