Search for Two-Photon Interaction with Axionlike Particles Using High-Repetition Pulsed Magnets and Synchrotron X Rays.

We report on new results of a search for a two-photon interaction with axionlike particles (ALPs). The experiment is carried out at a synchrotron radiation facility using a "light shining through a wall (LSW)" technique. For this purpose, we develop a novel pulsed-magnet system, composed of multiple racetrack magnets and a transportable power supply. It produces fields of about 10 T over 0.8 m with a high repetition rate of 0.2 Hz and yields a new method of probing a vacuum with high intensity fields. The data obtained with a total of 27 676 pulses provide a limit on the ALP-two-photon coupling constant that is more stringent by a factor of 5.2 compared to a previous x-ray LSW limit for the ALP mass ≲0.1 eV.

Hypoxia-inducible factor-1α inhibits interleukin-6 and -8 production in gingival epithelial cells during hypoxia.

BACKGROUND AND OBJECTIVE: Hypoxia has been widely studied in inflammatory diseases as it can modulate the inflammatory response, mainly via the hypoxia-inducible factor (HIF). However, little is known about the effects of hypoxia and the role of HIF in the inflammatory responses to periodontitis. In this study, we focused on the gingival epithelium that is exposed to relatively low levels of oxygen. We investigated whether hypoxic conditions have an impact on inflammatory responses in human gingival epithelial cells (HGECs). MATERIAL AND METHODS: Pimonidazole HCl, which accumulates in hypoxic cells, was administered intraperitoneally to C57BL/6 mice with or without Porphyromonas gingivalis infection. Immunohistochemistry was then performed to detect the hypoxic cells in periodontal tissue. Immortalized HGECs were cultured under hypoxic conditions with or without interleukin (IL)-1ß, and the expression levels of IL-6 and IL-8 were measured by real-time reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. HIF-1α expression was detected by western blotting. The DNA-binding activity of HIF-1α was determined by a DNA-binding enzyme-linked immunosorbent assay. The involvement of HIF-1α in the hypoxic response was examined by transfection with HIF-1α siRNA. RESULTS: Immunohistochemistry revealed pimonidazole HCl accumulation in the gingival epithelium of both normal and P. gingivalis-infected mice, with a slightly stronger signal in the P. gingivalis-infected mice than in the normal mice. The IL-1ß-induced IL-6 and IL-8 production by HGECs was suppressed under hypoxic conditions. HIF-1α accumulated during hypoxia, and this accumulation was further enhanced by IL-1ß treatment. The hypoxia-dependent suppression of IL-6 and IL-8 expression was reversed by treating the cells with HIF-1α siRNA. CONCLUSION: Our results suggest that the gingival epithelium is exposed to low oxygen tension in periodontal tissue and that this hypoxic condition modulates the local inflammatory response of gingival epithelial cells in an HIF-1α-dependent manner.

Differential inflammatory response of dental pulp explants and fibroblasts to saliva.

AIM: To investigate the inflammatory response of dental pulp fibroblasts and the respective explants to whole saliva. METHODOLOGY: Explants from human and porcine dental pulp tissue and isolated dental pulp fibroblasts were used to investigate the inflammatory response to sterile saliva. Cytokine and chemokine expression was assessed by RT-PCR. Western blot analysis and pharmacologic inhibitors were used to determine the involvement of signalling pathways. RESULTS: Dental pulp explants of human and porcine origin exposed to human saliva exhibited no major changes of IL-6 and IL-8 mRNA expression (P > 0.05). In contrast, isolated porcine and human dental pulp fibroblasts, when stimulated with human saliva, exhibited a vastly increased expression of IL-6 and IL-8 mRNA (P < 0.05). In pulp fibroblasts, saliva also increased the expression of other cytokines and chemokines via activation of NFkappaB, ERK and p38 signalling. Notably, a significantly reduced inflammatory response was elicited when pulp fibroblasts were transiently exposed to saliva. CONCLUSIONS: Saliva has a potential impact on inflammation of dental pulp fibroblasts in vitro but not when cells are embedded in the intrinsic extracellular matrix of the explant tissue.

Malignant female adnexal tumor of Wolffian origin (FATWO) positive for CD56: a possible diagnostic role for the biomarker.

Female adnexal tumors of Wolffian origin (FATWOs) are rare tumors that arise in the broad ligament from the remnants of the mesonephric duct. Most FATWOs behave in a benign fashion, and there are only 14 case reports worldwide describing malignant FATWOs. The authors report herein the case of a 69-year-old woman with a malignant FATWO, positive for CD56. The mass was composed mainly of solid neoplastic epithelial cells, closely packed, branching, and anastomosing in slender tubules. There was an eosinophilic secretion within the lumens of some of the cysts and tubules. The number of mitoses was somewhat high in the active areas, numbering five to seven per ten high-power fields. The tumor cells were strongly positive for glutathione S-transferase π, and positive for cal- retinin, vimentin, c-Kit, CD99, and CD56; neuron-specific enolase was also partially expressed. The tumor cells were negative for inhibin α, estrogen receptors, progesterone receptors, B-cell lymphoma 2, and S100. Taken together, these immunohistochemical and pathological findings gave the diagnosis of malignant FATWO. The patient experienced a recurrence one year after her initial surgery. CD56 immunostaining was negative in two benign FATWO cases at the present institution. These findings suggest that CD56-positivity may be a diagnostic biomarker to differentiate malignant FATWOs from benign lesions.

Estradiol and raloxifene induce the proliferation of osteoblasts through G-protein-coupled receptor GPR30.

BACKGROUND: Although G-protein-coupled receptor, GPR30, has been considered as a G-protein-coupled estrogen receptor, conflicting results have been reported and the function of GPR30 in bone remains unresolved. The aim of this study was to clarify the functional role of GPR30 in osteoblasts using its derived cell line. METHODS AND RESULTS: Immunohistochemical study revealed that GPR30 is expressed in human osteoblasts. Human fetal osteoblast cell lines, hFOB cells, which express GPR30 but lack estrogen receptor, were used for the in vitro experiments. Estradiol or raloxifene induced the proliferation of hFOB cells, which was accompanied by the activation of mitogen-activated protein (MAP) kinase. Those proliferative effects were completely abrogated by the transfection of GPR30 small interfering RNA, while the transfection alone did not affect the cell viability. CONCLUSION: GPR30 is required for the proliferation of hFOB cells induced by estradiol or raloxifene. This proliferative effect was at least partly mediated via MAP kinase activation. These findings revealed a novel function of GPR30 in osteoblasts and might lead to a better understanding of how estrogen and selective estrogen receptor modulators show their osteoprotective effects.

Removal characteristics of retinoic acids and 4-oxo-retinoic acids in wastewater by activated sludge treatment.

Retinoic acid (RA) receptor (RAR) agonists are potential teratogens to various vertebrates. Their contamination has been detected in municipal wastewater in different countries. This study involved field investigations and laboratory batch treatment experiments to elucidate the removal characteristics by activated sludge treatment of RAs (all-trans RA and 13-cis RA) and 4-oxo-RAs (4-oxo-all-trans RA and 4-oxo-13-cis RA), which were identified as major RAR agonists in municipal wastewater. Results obtained in this study show that currently employed activated sludge treatments can remove RAs, 4-oxo-RAs and overall RAR agonist contamination effectively from municipal wastewater in general, although high RAR agonistic activity might sometimes remain in the effluent. Laboratory experiments revealed that RAs were removed rapidly from the aqueous phase by adsorption to the sludge, after which they were removed further by biological and/or chemical degradation. Aside from adsorption to the sludge, 4-oxo-RAs were also apparently removed by biological and chemical degradation. Biodegradation contributed greatly to the removal. Results of additional experiments indicated that novel non-identifiable RAR agonists can occur through the biodegradation of 4-oxo-RAs by activated sludge and that they can persist for a long period.

Adiponectin regulates functions of gingival fibroblasts and periodontal ligament cells.

BACKGROUND AND OBJECTIVE: Adiponectin is a cytokine constitutively produced by adipocytes and exhibits multiple biological functions by targeting various cell types. However, the effects of adiponectin on primary gingival fibroblasts and periodontal ligament cells are still unexplored. Therefore, we investigated the effects of adiponectin on gingival fibroblasts and periodontal ligament cells. MATERIAL AND METHODS: The expression of adiponectin receptors (AdipoR1 and AdipoR2) on human gingival fibroblasts (HGFs), mouse gingival fibroblasts (MGFs) and human periodontal ligament (HPDL) cells was examined using RT-PCR and western blotting. HGFs and MGFs were stimulated with interleukin (IL)-1ß in the presence or absence of adiponectin, and the expression of IL-6 and IL-8 at both mRNA and protein levels was measured by real-time PCR and ELISA, respectively. Furthermore, small interfering RNAs (siRNAs) in MGFs were used to knock down the expression of mouse AdipoR1 and AdipoR2. The effects of adiponectin on the expression of alkaline phosphatase (ALP) and runt-related transcription factor 2 (Runx2) genes were evaluated by real-time PCR. Mineralized nodule formation of adiponectin-treated HPDL cells was revealed by Alizarin Red staining. RESULTS: AdipoR1 and AdipoR2 were expressed constitutively in HGFs, MGFs and HPDL cells. Adiponectin decreased the expression of IL-6 and IL-8 in IL-1ß-stimulated HGFs and MGFs. AdipoR1 siRNA in MGFs revealed that the effect of adiponectin on reduction of IL-6 expression was potentially mediated via AdipoR1. Adiponectin-treated HPDL cells promoted the expression of ALP and Runx2 mRNAs and up-regulated ALP activity. Furthermore, adiponectin enhanced mineralized nodule formation of HPDL cells. CONCLUSION: Our observations demonstrate that adiponectin exerts anti-inflammatory effects on HGFs and MGFs, and promotes the activities of osteoblastogenesis of HPDL cells. We conclude that adiponectin has potent beneficial functions to maintain the homeostasis of periodontal health, improve periodontal lesions, and contribute to wound healing and tissue regeneration.

Effect of ß-carotene supply during close-up dry period on the onset of first postpartum luteal activity in dairy cows.

The aim of this study was to examine the effect of ß-carotene supply during the close-up dry period on the onset of first postpartum luteal activity in dairy cows. Twelve cows were supplied with 2000 mg of ß-carotene (20 g Rovimix(®) ß-Carotene containing 10% ß-carotene; DSM Nutrition Japan K.K., Tokyo, Japan) by oral administration daily from day 21 before expected calving date to parturition. Fourteen cows (control) did not receive ß-carotene supplementation. Blood samples were obtained on days 21, 14 and 7 before expected calving date and on days 1, 7, 14, 21 postpartum. When the plasma progesterone concentration exceeded 1 ng/ml by day 21 postpartum, luteal activity was assumed to have been initiated. The result showed that serum ß-carotene concentrations in the ß-carotene cows were higher than in the control cows during the experimental period (p < 0.01). The number of cows with the onset of luteal activity by day 21 postpartum was 9/12 in the ß-carotene cows and 4/14 in the control cows (p < 0.05). Retinol, certain metabolic parameters and metabolic hormones concentrations did not differ between ß-carotene and control cows. In addition, serum retinol concentration in ß-carotene cows without luteal activity was lower than in ß-carotene cows with luteal activity (p < 0.05), and serum gamma-glutamyl transpeptidase concentration in ß-carotene cows with luteal activity (p < 0.05) and control cows without luteal activity (p < 0.01) was higher than in control cows with luteal activity. In conclusion, ß-carotene supply during the close-up dry period may support the onset of luteal activity during early lactation in dairy cows.

Clinical outcomes of magnetic resonance-guided focused ultrasound surgery for uterine myomas: 24-month follow-up.

OBJECTIVES: To assess the volume reduction ratio, symptom improvement and reintervention rate following magnetic resonance-guided focused ultrasound surgery (MRgFUS) for uterine myomas. METHODS: A total of 91 Japanese women with symptomatic myomas underwent MRgFUS between June 2004 and June 2008 using the ExAblate 2000 system. The volume change ratio was calculated at 6, 12 and 24 months following MRgFUS based on T2-weighted magnetic resonance images. The symptom severity score (SSS) was examined before and after the treatment (at 3, 6, 12 and 24 months). Additional post-MRgFUS treatments, such as hysterectomy, myomectomy, uterine artery embolization or repeat MRgFUS, were recorded and the reinterventional treatment rates were compared according to the signal intensity of pretreatment T2-weighted magnetic resonance images of the myomas. RESULTS: The mean volume change ratios of low- and intermediate-intensity (Type 1/2) myomas were -36.5% 6 months post-procedure and -39.5% 24 months post-procedure. The mean +/- SD SSS value for patients with Type 1/2 myomas before MRgFUS was 35.1 +/- 21.0, and the values diminished significantly during the 24-month follow-up period to a mean value of around 15.0. High-intensity (Type 3) myomas were not observed to have decreased in size 6 months after MRgFUS. Of the 45 Type 1/2 myoma patients with complete follow-up, seven required reinterventional treatment within 24 months. The reintervention rates were 14.0% for Type 1/2 patients and 21.6% for Type 3 patients at 24 months post-treatment. CONCLUSIONS: Moderate volume reductions of Type 1/2 myomas were noted following MRgFUS, and the reduction in SSS values and the relatively low reintervention rates observed are encouraging. We found MRgFUS to be an appropriate treatment method for Type 1/2 uterine myomas.

Amniotic fluid 'sludge' detected in patients with subchorionic hematoma: a report of two cases.

Amniotic fluid 'sludge' is defined as the presence of dense aggregates of particulate matter in close proximity to the internal cervical os. It is of clinical significance in asymptomatic patients at high risk for spontaneous delivery, and in patients with preterm labor and intact membranes. Subchorionic hematoma is another ultrasound finding that is associated with a higher incidence of threatened miscarriage and preterm delivery. We report two cases of occurrence of amniotic fluid sludge in patients with previously detected large subchorionic hematoma. In the first case subchorionic hematoma and amniotic fluid sludge were detected by ultrasonography at 13 + 1 and 18 + 6 weeks' gestation, respectively, followed by preterm premature rupture of membranes, placental abruption and emergency Cesarean section. In the second case subchorionic hematoma and amniotic fluid sludge were detected by ultrasound at 11 + 3 and 15 + 5 weeks' gestation, respectively, followed by miscarriage with histological chorioamnionitis. The coincidence of subchorionic hematoma and amniotic fluid sludge in these cases points to a possible connection between these two significant ultrasound findings.

Clinicopathological features and prognosis in immunoglobulin light and heavy chain deposition disease.

BACKGROUND: There are three subtypes of monoclonal immunoglobulin deposition disease: light chain deposition disease (LCDD), light and heavy chain deposition disease (LHCDD), and heavy chain deposition disease (HCDD). Although it has been considered that LHCDD is a variant of LCDD, information on clinicopathological features and prognosis in LHCDD is presently limited. METHODS: We reviewed 5,443 renal biopsies, and evaluated clinicopathological features and outcomes in patients with LHCDD, in comparison with those in patients with LCDD and previously reported patients with HCDD. We also characterized paraprotein deposits in patients with LHCDD. RESULTS: We identified 6 patients with LHCDD, 6 patients with LCDD, and 1 patient with HCDD. The most common clinicopathological findings in patients with LHCDD were proteinuria, renal insufficiency, and nodular sclerosing glomerulopathy. Three patients had IgG-k deposits and 3 patients had IgG-l deposits. Heavy chain subclass analysis performed in 4 patients showed IgG3 deposits in all patients. Dual immunostaining revealed glomerular colocalization of light and heavy chains. In contrast with LCDD, glomerular C3 and C1q deposits were common findings in LHCDD and HCDD. All patients with LHCDD were treated with steroids and cytotoxic agents, but no effect on proteinuria was observed. Three patients developed end-stage renal disease requiring hemodialysis. The underlying hematological disorders in LHCDD and HCDD were milder than in LCDD. Early renal survival and overall patient survival in our patients appeared to be better in LHCDD than in LCDD. CONCLUSIONS: There are apparent differences in clinicopathological features and prognosis between LHCDD and LCDD. LHCDD is probably more similar to HCDD.

GATA-3 is upregulated in peripheral blood mononuclear cells from patients with minimal change nephrotic syndrome.

BACKGROUND: An imbalance of Th1 and Th2 cytokines has been reported in MCNS. Interleukin-13 (IL-13: Th2 cytokine) has been implicated in the pathogenesis of MCNS, but Th1/Th2 regulators such as T-bet (Th1-specific transcription factor) and GATA-3 (Th2-specific transcription factor) have not been examined. METHODS: We isolated PBMC from 25 patients with MCNS during nephrosis and remission phases, from 17 nephrotic patients with membranous nephropathy (MN), and from 25 healthy subjects. We measured mRNA expression levels of T-bet, GATA-3, Stat5A (regulator of Th2 priming), IFN-gamma (Th1 cytokine), IL-2 (Th1 cytokine and activator of Stat5), IL-4 (Th2 cytokine), and IL-13 in PBMC, using real-time RT-PCR. RESULTS: GATA-3, Stat5A, and IL-13 mRNA expression levels were higher in the nephrotic MCNS group compared to the others. IL-2 mRNA expression levels were higher in nephrotic patients with MCNS and MN than in MCNS patients in remission and healthy controls. There were no differences in mRNA expression levels of T-bet, IFN-gamma, and IL-4 between MCNS and MN patients and healthy controls. CONCLUSIONS: This study is the first to reveal increased mRNA expression levels of GATA-3 and Stat5A in PBMC from MCNS patients in nephrosis. This study also supports recent findings suggesting the role of IL-13 in the development of MCNS. A predominant Th2 type of T cell activation may be involved in the pathogenesis of MCNS.

Characteristics of proliferative glomerulo-nephritis with monoclonal IgG deposits associated with membranoproliferative features.

BACKGROUND: Proliferative glomerulonephritis with monoclonal IgG deposits (PGNMID) associated with membranoproliferative features is an extremely rare entity. Information on clinicopathological features and prognosis in this entity is limited. METHODS: We reviewed 5,443 renal biopsies processed at our department, and identified 4 patients with PGNMID associated with membranoproliferative features. We evaluated clinicopathological features and outcomes in these patients, and characterized paraprotein deposits by immunofluorescence studies. RESULTS: Three out of 4 patients had nephrotic syndrome with renal insufficiency at presentation. Cryoglobulin or monoclonal protein in serum and urine was not detected. Renal biopsy showed membranoproliferative features with or without nodular formation. Tubulointerstitial and vascular alterations were mild in three patients. All patients had glomerular IgG-kappa deposits. Heavy chain subclass analysis performed in 3 patients showed IgG3 deposits. Immunofluorescence studies using antibodies specific for gamma-heavy chain C(H)1, C(H)2, and C(H)3 domains and gamma3 hinge did not show any apparent deletion. Confocal microscopy revealed glomerular colocalization of light and heavy chains. On electron microscopy, granular deposits were predominantly mesangial and subendothelial. All patients were treated with steroids and cytotoxic agents, but no effect on proteinuria was observed. The renal outcome was progressive in all patients. Early death was observed in two elder patients. No patient had overt myeloma or lymphoma at presentation or over the course of follow-up (mean 43 months). CONCLUSIONS: Our study suggests a predominance of IgG3-kappa glomerular deposits of nondeleted whole immunoglobulin molecules in PGNMID associated with membranoproliferative features. The clinical outcome in patients with this entity appears to be poor.

Carrier-induced noncollinear magnetism in perovskite manganites by first-principles calculations.

We have performed noncollinear first-principles density-functional calculations of carrier-doped perovskite manganites La(1-x)Sr(x)MnO(3) (0.0≤x≤1.0). In the calculated magnetic phase diagram (T = 0) within the collinear magnetic configurations, ferromagnetic and several antiferromagnetic configurations successively appeared as a ground state with increasing x. The calculated total energies of the ferromagnetic and A-type antiferromagnetic phases are almost degenerate around the phase boundary, x = 0.5. We found that the noncollinear magnetic configurations are stable in a wide range of carrier concentrations 0.3≤x≤0.6. We discuss the effect of lattice distortions on the stability of the noncollinear magnetic phase.

Mechanical regulation of bone homeostasis through p130Cas-mediated alleviation of NF-κB activity.

Mechanical loading plays an important role in bone homeostasis. However, molecular mechanisms behind the mechanical regulation of bone homeostasis are poorly understood. We previously reported p130Cas (Cas) as a key molecule in cellular mechanosensing at focal adhesions. Here, we demonstrate that Cas is distributed in the nucleus and supports mechanical loading-mediated bone homeostasis by alleviating NF-κB activity, which would otherwise prompt inflammatory processes. Mechanical unloading modulates Cas distribution and NF-κB activity in osteocytes, the mechanosensory cells in bones. Cas deficiency in osteocytes increases osteoclastic bone resorption associated with NF-κB-mediated RANKL expression, leading to osteopenia. Upon shear stress application on cultured osteocytes, Cas translocates into the nucleus and down-regulates NF-κB activity. Collectively, fluid shear stress-dependent Cas-mediated alleviation of NF-κB activity supports bone homeostasis. Given the ubiquitous expression of Cas and NF-κB together with systemic distribution of interstitial fluid, the Cas-NF-κB interplay may also underpin regulatory mechanisms in other tissues and organs.

Up-regulated expression of Toll-like receptors mRNAs in peripheral blood mononuclear cells from patients with systemic lupus erythematosus.

Recent studies in animal models for systemic lupus erythematosus (SLE) have shown that Toll-like receptors (TLR-7 and TLR-9) and interferon (IFN)-alpha are involved in the pathogenesis of murine lupus. Recent studies using flow cytometry have also shown increased expression of TLR-9 in peripheral blood mononuclear cells (PBMCs) from SLE patients. In this study, we performed quantitative real-time reverse transcription-polymerase chain reaction analyses of PBMCs from 21 SLE patients and 21 healthy subjects, to estimate TLR2, TLR3, TLR4, TLR5, TLR7, TLR8, TLR9, IFN-alpha and LY6E (a type I IFN-inducible gene) mRNA expression levels. Expression levels of TLR2, TLR7, TLR9, IFN-alpha and LY6E mRNAs in SLE patients were significantly higher than those in healthy controls. Expression levels of TLR7 and TLR9 mRNAs correlated with that of IFN-alpha mRNA in SLE patients. These results suggest that up-regulated expression of TLR7 and TLR9 mRNAs together with increased expression of IFN-alpha mRNA in PBMCs may also contribute to the pathogenesis of human lupus.

Differential alterations in expressions of ryanodine receptor subtypes in cerebellar cortical neurons of an ataxic mutant, rolling mouse Nagoya.

This study aimed to clarify changes in the spatial expressions of types 1, 2 and 3 ryanodine receptors (RyR1, RyR2 and RyR3) in the cerebellum of a Ca(2+) channel alpha(1A) subunit mutant, rolling mouse Nagoya. Semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) revealed that the mRNA signal levels of RyR1 and RyR3 were altered in the rolling cerebellum, which exhibited lower densities of RyR1 bands and higher densities of RyR3 bands than in the control cerebellum. Quite consistent with the RT-PCR results, the staining intensity of RyR1 and RyR3 was altered in the rolling cerebellum. RyR1 immunostaining appeared in somata and the proximal dendrites of Purkinje cells, and the staining intensity of both subcellular regions was equally lower in all cerebellar lobules of rolling mice than in those of controls. Although RyR3 immunostaining appeared in the dendrites of granule cells, more intense RyR3 staining in rolling mice than in controls was uniformly observed throughout all cerebellar lobules. The present study further examined co-localizations of ryanodine receptor subtypes and voltage-gated Ca(2+) channel alpha(1) subunits in the rolling cerebellum. Somatodendritic RyR1 immunostaining in Purkinje cells overlapped with either a mutated Ca(2+) channel alpha(1A) subunit (P/Q-type), or a Ca(2+) channel alpha(1C) subunit (L-type; dihydropyridine receptor) immunostaining. Immunostaining of these alpha(1) subunits also emerged in granule cells. Those results suggest non-region-related alterations in RyR1 and RyR3 expressions in the rolling mouse cerebellum. Such expressional changes in ryanodine receptor subtypes may be involved in Ca(2+) channel alpha(1A) subunit gene mutation, and may alter regulation of intracellular Ca(2+) concentrations in cerebellar cortical neurons.

Study of neuroprotection of donepezil, a therapy for Alzheimer's disease.

Donepezil is a potent acetylcholinesterase inhibitor used for the treatment of Alzheimer's disease. Although acetylcholinesterase inhibitors are thought to be symptomatic treatment of Alzheimer's disease, it is not clear whether they are effective against progressive degeneration of neuronal cells. In this study, we investigated the neuroprotective effects of donepezil against ischemic damage, N-methyl-d-aspartate (NMDA) excitotoxicity, and amyloid-beta (Abeta) toxicity using rat brain primary cultured neurons. Lactate dehydrogenase (LDH) released into the culture medium was measured as a marker of neuronal cell damage. As an ischemic damage model, we used oxygen-glucose deprivation in rat cerebral cortex primary cultured neurons. Pretreatment with donepezil (0.1, 1 and 10 microM) significantly decreased LDH release in a concentration-dependent manner. However, other acetylcholinesterase inhibitors (galantamine, tacrine and rivastigmine) did not significantly decrease LDH release. In a NMDA excitotoxicity model, pretreatment with donepezil (0.1, 1 and 10 microM) decreased the LDH release in a concentration-dependent manner. In binding assay for glutamate receptors, donepezil at 100 microM only slightly inhibited binding to the glycine and polyamine sites on NMDA receptor complex. We further examined the effect of donepezil on Abeta (1-40)- and Abeta (1-42)-induced toxicity in primary cultures of rat septal neurons. Pretreatment with donepezil (0.1, 1 and 10 microM) significantly decreased LDH release induced by Abetas in a concentration-dependent manner. However, other acetylcholinesterase inhibitors (galantamine and tacrine) and NMDA receptor antagonists (memantine and dizocilpine (MK801)) did not significantly decrease LDH release. These results demonstrate that donepezil has protective effects against ischemic damage, glutamate excitotoxicity and Abeta toxicity to rat primary cultured neurons and these effects are not dependent on acetylcholinesterase inhibition and antagonism of NMDA receptors. Thus, donepezil is expected to have a protective effect against progressive degeneration of brain neuronal cells in ischemic cerebrovascular disease and Alzheimer's disease.

Predominant tubulointerstitial nephritis in a patient with systemic lupus erythematosus: phenotype of infiltrating cells.

A 63-year-old man with systemic lupus erythematosus developed tubular proteinuria. All subclasses of serum IgG increased, and the largest IgG subclass increase was IgG4. A renal biopsy showed lupus nephritis (Class II) with severe tubulointerstitial nephritis (so-called predominant tubulointerstitial lupus nephritis, an unusual form of lupus nephritis). Immunofluorescence microscopy revealed positive granular staining for IgG, C3 and C1q in the mesangium and peritubular interstitium, and along the tubular basement membranes (TBM). Electron microscopy also showed electron-dense deposits in the mesangium and TBM. Immunophenotyping of interstitial infiltrating cells disclosed a predominance of T cells. CD8-positive cytotoxic T cells infiltrated the peritubular interstitium, and some of these cells infiltrated the tubules. B cell-rich lymphoid follicles were also observed. IgG subclass analyses showed glomerular IgG1, IgG2 and IgG4 deposition, positive staining of IgG4 in the peritubular interstitium and along the TBM, and abundant IgG1-, IgG3- and IgG4-positive plasma cells in the interstitium. The patient responded well to moderate-dose steroid therapy. This is the first report of immunophenotyping of interstitial infiltrates in predominant tubulointerstitial lupus nephritis. The results suggest CD8-positive cytotoxic T cell-mediated tubular injury. Furthermore, immune complexes containing IgG4 might be one of etiologic factors.