Affinity imaging mass spectrometry (AIMS): high-throughput screening for specific small molecule interactions with frozen tissue sections.

A novel screening system, using affinity imaging mass spectrometry (AIMS), has been developed to identify protein aggregates or organ structures in unfixed human tissue. Frozen tissue sections are positioned on small (millimetre-scale) stainless steel chips and incubated with an extensive library of small molecules. Candidate molecules showing specific affinity for the tissue section are identified by imaging mass spectrometry (IMS). As an example application, we screened over a thousand compounds against Alzheimer's disease (AD) brain tissue and identified several compounds with high affinity for AD brain sections containing tau deposits compared to age-matched controls. It should also be possible to use AIMS to isolate chemical compounds with affinity for tissue structures or components that have been extensively modified by events such as oxidation, phosphorylation, acetylation, aggregation, racemization or truncation, for example, due to aging. It may also be applicable to biomarker screening programs.

Management of angiogram-negative acute colonic hemorrhage: safety and efficacy of colonoscopy-guided superselective embolization.

BACKGROUND: We evaluated the efficacy and safety of superselective embolization with assistance of colonoscopy for acute colonic hemorrhage. METHODS: Of 92 cases of acute colonic hemorrhage requiring colonoscopic intervention, 11 (12 %) could not be successfully treated. Of these, 10 patients (9 men, mean age 65.5 years, range 39-75 years) underwent superselective embolization. Hemorrhage was caused by diverticular disease (n = 8), polypectomy (n = 1), and vascular malformation (n = 1). In all 10 cases, the radiopaque clips were placed at the bleeding point via colonoscopy. Microcatheters were used in all procedures, and embolization was performed at the level of the vasa recta leading to or near the clips with Gelfoam particles, microcoils, or both. RESULTS: Immediate hemostasis was achieved in all patients. In 6 of 10 patients (60 %), selective angiograms showed no active extravasation at the time of the procedure and the embolization was performed using clips as a landmark. In the remaining four patients, selective angiograms showed active extravasation from the vasa recta leading to the clips. The mean number of embolized vessels with no active extravasation and with active extravasation was 1.83 (range 1-3) and 1.25 (range 1-2), respectively. The mean duration of clinical follow-up was 11.6 months (range 1-29 months). One patient (10 %) bled from a different site than the treated site a month after embolization, but the bleeding ceased after endoscopic intervention. All the patients (100 %) were evaluated for objective evidence of ischemia by colonoscopy. Four of the 10 patients (40 %) were found endoscopically to have small areas of ischemia involving only the mucosa, but they remained asymptomatic. There was no bowel infarction or stricture. CONCLUSIONS: Colonoscopy-assisted superselective embolization may be a safe and useful procedure for acute colonic hemorrhage without active extravasation on angiogram.

Therapeutic efficacy of intra-arterial docetaxel and nedaplatin infusion concomitant with radiotherapy for T4 maxillary sinus squamous cell carcinoma.

The aim of this study was to evaluate the efficacy of intra-arterial chemoradiotherapy with docetaxel and nedaplatin for T4 maxillary sinus squamous cell carcinoma (MSSCC). Data were retrospectively analysed for 22 consecutive patients with T4 MSSCC who underwent intra-arterial chemoradiotherapy. Participants received intensity-modulated radiotherapy (70 Gy in 35 fractions) concomitantly with docetaxel (60 mg/m2) and nedaplatin (80 mg/m2) administered every 4 weeks for a total of three sessions. The median follow-up period was 49 months (range 12-91 months). T4a tumours were found in 16 patients (73%) and T4b tumours in six patients (27%). Cervical metastasis was found in nine patients (41%; five N2b, four N2c). The 5-year loco-regional control, disease-free survival, and overall survival rates for patients with T4a disease were 92.3%, 92.3%, and 90.3%, respectively, compared to 83.3% (P = 0.42), 66.7% (P = 0.07), and 83.3% (P = 0.46), respectively, for those with T4b disease. The 5-year loco-regional control, disease-free survival, and overall survival rates for patients with cervical lymph node metastasis were all 87.5% compared to 92.3% (P = 0.86), 84.6% (P = 0.69), and 92.3% (P = 0.93), respectively, for those without cervical metastasis. Intra-arterial chemoradiotherapy with docetaxel and nedaplatin may provide favourable loco-regional control and increased survival in T4 MSSCC.

Less protease-resistant PrP in a patient with sporadic CJD treated with intraventricular pentosan polysulphate.

Treatment with intraventricular pentosan polysulphate (PPS) might be beneficial in patients with Creutzfeldt-Jakob disease. We report a 68-year-old woman with sporadic Creutzfeldt-Jakob disease who received continuous intraventricular PPS infusion (1-120 microg/kg/day) for 17 months starting 10 months after the onset of clinical symptoms. Treatment with PPS was well tolerated but was associated with a minor, transient intraventricular hemorrhage and a non-progressive collection of subdural fluid. The patient's overall survival time was well above the mean time expected for the illness but still within the normal range. Post-mortem examination revealed that the level of abnormal protease-resistant prion protein in the brain was markedly decreased compared with levels in brains without PPS treatment. These findings suggest that intraventricular PPS infusion might modify the accumulation of abnormal prion proteins in the brains of patients with sporadic Creutzfeldt-Jakob disease.

Cerebrospinal fluid metabolite and nigrostriatal dopaminergic function in Parkinson's disease.

OBJECTIVES: To evaluate the association between cerebrospinal fluid (CSF) homovanillic acid (HVA) concentrations and nigrostriatal dopaminergic function assessed by positron emission tomography (PET) imaging with carbon-11-labeled 2beta-carbomethoxy-3beta-(4-fluorophenyl)-tropane ((11)C-CFT), which can measure the dopamine transporter (DAT) density, in Parkinson's disease (PD). METHODS: (11)C-CFT PET scans and CSF examinations were performed on 21 patients with PD, and six patients with non-parkinsonian syndromes (NPS) as a control group. RESULTS: In the PD group, CSF HVA concentrations were significantly correlated with the striatal uptake of (11)C-CFT (r = 0.76, P < 0.01). However, in the NPS group, two indices were within the normal range. CONCLUSIONS: In PD, CSF HVA concentrations correlate with nigrostriatal dopaminergic function. Therefore, CSF HVA concentrations may be an additional surrogate marker for estimating the remaining nigrostriatal dopaminergic function in case that DAT imaging is unavailable.

Role of NADH shuttle system in glucose-induced activation of mitochondrial metabolism and insulin secretion.

Glucose metabolism in glycolysis and in mitochondria is pivotal to glucose-induced insulin secretion from pancreatic beta cells. One or more factors derived from glycolysis other than pyruvate appear to be required for the generation of mitochondrial signals that lead to insulin secretion. The electrons of the glycolysis-derived reduced form of nicotinamide adenine dinucleotide (NADH) are transferred to mitochondria through the NADH shuttle system. By abolishing the NADH shuttle function, glucose-induced increases in NADH autofluorescence, mitochondrial membrane potential, and adenosine triphosphate content were reduced and glucose-induced insulin secretion was abrogated. The NADH shuttle evidently couples glycolysis with activation of mitochondrial energy metabolism to trigger insulin secretion.

Occlusal Support, Dysphagia, Malnutrition, and Activities of Daily Living in Aged Individuals Needing Long-Term Care: A Path Analysis.

OBJECTIVES: This study aimed to examine the interrelationships among occlusal support, dysphagia, malnutrition, and activities of daily living in aged individuals needing long-term care. DESIGN: Cross-sectional study and path analysis. SETTING: Long-term health care facilities, acute care hospitals, and the community. PARTICIPANTS: Three hundred and fifty-four individuals aged ≥ 65 years with dysphagia or potential dysphagia in need of long-term care. MEASUREMENTS: The modified Eichner Index, Dysphagia Severity Scale, Mini Nutritional Assessment Short Form, and Barthel index. RESULTS: The participants included 118 males and 236 females with a mean (standard deviation) age of 83 (8) years. A total of 216 participants had functional occlusal support with or without dentures. Of the total participants, 73 were within normal limits regarding the severity of dysphagia, 119 exhibited dysphagia without aspiration, and 162 exhibited dysphagia with aspiration. Only 34 had a normal nutritional status, while 166 participants were malnourished, and 154 were at risk of malnutrition. The median Barthel index score was 30. Path analysis indicated two important findings: occlusal support had a direct effect on dysphagia (standard coefficient = 0.33), and dysphagia was associated directly with malnutrition (standard coefficient = 0.50). Dysphagia and malnutrition were associated directly with impaired activities of daily living (standard coefficient = 0.57, 0.22). CONCLUSION: In aged individuals needing long-term care, occlusal support is associated directly with dysphagia and indirectly with malnutrition and activities of daily living via dysphagia.

Molecular epidemiologic analysis of a Pneumocystis pneumonia outbreak among renal transplant patients.

Between 18 November and 3 December 2011, five renal transplant patients at the Department of Nephrology, Toho University Omori Medical Centre, Tokyo, were diagnosed with Pneumocystis pneumonia (PCP). We used molecular epidemiologic methods to determine whether the patients were infected with the same strain of Pneumocystis jirovecii. DNA extracted from the residual bronchoalveolar lavage fluid from the five outbreak cases and from another 20 cases of PCP between 2007 and 2014 were used for multilocus sequence typing to compare the genetic similarity of the P. jirovecii. DNA base sequencing by the Sanger method showed some regions where two bases overlapped and could not be defined. A next-generation sequencer was used to analyse the types and ratios of these overlapping bases. DNA base sequences of P. jirovecii in the bronchoalveolar lavage fluid from four of the five PCP patients in the 2011 outbreak and from another two renal transplant patients who developed PCP in 2013 were highly homologous. The Sanger method revealed 14 genomic regions where two differing DNA bases overlapped and could not be identified. Analyses of the overlapping bases by a next-generation sequencer revealed that the differing types of base were present in almost identical ratios. There is a strong possibility that the PCP outbreak at the Toho University Omori Medical Centre was caused by the same strain of P. jirovecii. Two different types of base present in some regions may be due to P. jirovecii's being a diploid species.

Two types of chicken 2',5'-oligoadenylate synthetase mRNA derived from alleles at a single locus.

We have isolated two types of chicken 2',5'-oligoadenylate synthetase cDNAs, A and B, which encode predicted proteins of 508 amino acids (58316 Da) and 476 amino acids (54336 Da), respectively. The region of A-protein comprising 33 amino acid residues from 385Ala to 417Cys is substituted by a single amino acid 385Tyr in B-protein. The homology between chicken and mammalian 2',5'-oligoadenylate synthetases is 49.5% over the amino-terminal 337 residues. Proteins expressed from A- and B-cDNAs in E. coli cells were both active in synthesizing 2',5'-oligoadenylate. However, the activity of B-protein was 10-15% of that of A-protein. Southern blotting hybridization indicated that the chicken synthetases are encoded by a single gene. RT-PCR and PCR analyses of RNA and DNA of chicken erythrocytes together with the sequence data of the PCR products showed that A- and B-mRNAs are derived from alleles at a single locus encoding chicken 2',5'-oligoadenylate synthetase, designated as OAS * A and OAS * B. Chickens carrying OAS * A/B produce two types of synthetase with molecular masses of 58 and 54 kDa, and those carrying OAS * A/A produce only a single type of 58 kDa.

A multidisciplinary treatment strategy that includes high-dose chemotherapy for metastatic retinoblastoma without CNS involvement.

The prognosis of patients with metastatic retinoblastoma is poor with conventional chemotherapy and radiation. Since retinoblastoma is highly chemosensitive, dose-escalation of chemotherapeutic agents with stem cell support should be promising. We report our experience with high-dose chemotherapy (HDC) and autologous stem cell transplantation (SCT) in patients with metastatic retinoblastoma. Five patients with metastatic retinoblastoma underwent HDC with autologous SCT following conventional chemotherapy and local radiation therapy. Stem cells (bone marrow in four and peripheral blood stem cells in one) were collected after marrow involvement was cleared. Melphalan was a key drug in all patients, and was administered in combination with other agents such as cisplatin, cyclophosphamide, carboplatin or thiotepa. Three patients are currently alive disease-free at 113, 107 and 38 months, respectively, from the time of SCT. They had no central nervous system (CNS) involvement. The two patients who died of disease had CNS involvement. No long-term sequelae of HDC have been noted. Our treatment strategy using HDC appears to be effective for treating metastatic retinoblastoma without CNS involvement.

Effect of lysophosphatidic acid on the preimplantation development of mouse embryos.

The effect of lysophosphatidic acid (LPA) on the preimplantation development of mouse embryos was examined. The blastocyst rates from 2-cell and 4-cell stage embryos were significantly higher in the presence of LPA at the pronuclear stage. However, the blastocyst rate from 4-cell stage embryos was significantly lower when LPA was added to the medium at the 4-cell stage than when it was added at the pronuclear stage. Furthermore, pretreatment of pronuclear stage embryos with pertussis toxin suppressed LPA-induced embryo development, suggesting that it is probably mediated by a Gi-protein-linked receptor.

Calpain-mediated degradation of p35 to p25 in postmortem human and rat brains.

Tau in Alzheimer neurofibrillary tangles has been shown to be hyperphosphorylated and CDK5, GSK3, MAP kinase and SAP kinases are the candidate kinases for the phosphorylation of tau. Recently, it was reported that the conversion of p35, the activator of CDK5, to p25 was upregulated in Alzheimer's disease (AD) brains, and that p35 is cleaved to yield p25 by calpain. Here we show that p35 is rapidly cleaved to p25 in rat and human brains within a short postmortem delay and that the conversion of p35 to p25 is partially dependent on calpain activity. Immunoblot analysis of brains prepared from patients with AD or age-matched control individuals with a short postmortem delay revealed no specific increase in the levels of p25 in AD brains, whereas the levels of active form of calpain were increased in AD brains compared to the those in controls. These observations suggest that the conversion of p35 to p25 is a postmortem degradation event and may not be upregulated in AD brains.

Accumulation of neurofilaments and SOD1-immunoreactive products in a patient with familial amyotrophic lateral sclerosis with I113T SOD1 mutation.

OBJECTIVE: To report neuropathologic features of argyrophilic inclusions in the anterior horn cells, motor cortex Betz cells, and neurons of the medullary reticular formation, spinal posterior horn, and Clarke column in a Japanese case of familial amyotrophic lateral sclerosis with I113T substitution in exon 4 of the copper-zinc superoxide dismutase (SOD1) gene. METHODS AND RESULTS: These inclusions were stained pale pink on the hematoxylin-eosin stain and dark on the Bielschowsky stain. They were positive for antibodies to phosphorylated neurofilaments, ubiquitin, and SOD1. On electron microscopy, they consisted of abundant intermediate filaments of 10 to 20 nm in diameter with disordered array indicating neurofilaments. CONCLUSION: These findings suggest that the I113T mutation induces accumulation of neurofilaments and SOD1 in the central nervous system neurons.

Expression of tau immunoreactivity in the spinal motor neurons of Alzheimer's disease.

Tau-related pathology was investigated in the spinal cord of 11 patients with AD. Ten ALS and 10 age-matched neurologically intact patients served as controls. Tau immunoreactivity was detected in neurons of the anterior horn in all AD cases and to a lesser extent in the intermediate zone and in the posterior horn. Tau immunoreactivity was rare in controls. Neurofibrillary tangles were identified in seven AD cases, but none was observed in the controls.

Lewy body in neurodegeneration with brain iron accumulation type 1 is immunoreactive for alpha-synuclein.

In familial PD, a mutation of the alpha-synuclein gene has been identified. Alpha-synuclein also was revealed in Lewy bodies in idiopathic PD. Lewy bodies in neurodegeneration with brain iron accumulation type 1 (NBIA 1; Hallervorden-Spatz syndrome) were found to show immunostaining for alpha-synuclein/precursor of non-A beta component of Alzheimer's disease amyloid, indicating that alpha-synuclein is commonly associated with the formation of Lewy bodies in other sporadic and familial neurodegenerative diseases apart from PD.

Spatial and temporal disease progression of adult-onset subacute sclerosing panencephalitis.

An adult-onset case of subacute sclerosing panencephalitis with occipitofrontal spread of the infection documented clinically and by MRI is reported. Autopsy revealed numerous intranuclear viral inclusions and widespread demyelination in both frontal lobes. In the occipital lobes where the disease started 5 years previously, inclusions were rare, but degenerative tissue changes were prominent. This case underlines the importance of measles virus migration for the progression of this fatal disorder.

Unusual MRI and pathologic findings of progressive multifocal leukoencephalopathy complicating adult Wiskott-Aldrich syndrome.

We present a long-surviving patient with Wiskott-Aldrich syndrome complicated by atypical progressive multifocal leukoencephalopathy (PML). MRI showed multiple tiny spots of Gd-DTPA-enhanced lesions on the T1-weighted image. Pathologic findings for brain biopsy were patchy demyelinated vascularized lesions infiltrated by a surprising number of eosinophils. The presence of polyomavirus JC was confirmed by in situ hybridization and polymerase chain reaction. PML should be included in the differential diagnosis when Gd-DTPA-enhanced spotty lesions are present in the white matter, especially in patients who have a mild immunologic defect.

Distal sensory axonopathy after sarin intoxication.

A 51-year-old man inhaled sarin during a terrorist attack on the Tokyo subway system and died 15 months later. Neuropathologic examination revealed marked nerve fiber decrease in the sural nerve, moderate nerve fiber loss in the sciatic nerve, and unremarkable dorsal root ganglia, dorsal roots, and posterior column of the spinal cord. This pathology is consistent with dying-back degeneration of the peripheral nervous system and could represent a late sequela of sarin intoxication.

Early-onset, rapidly progressive familial tauopathy with R406W mutation.

An early-onset and rapidly progressive familial tauopathy with R406W mutation is described. The patient was a 47-year-old man who first presented with psychiatric symptoms followed by overt dementia at age 52 and died 1 year later. Postmortem study revealed tangle-associated neuronal degeneration, accentuated in the medial temporal lobe. R406W mutation was determined by sequence analysis and immunocytochemically with anti-mutant tau antibody.

The external radiotherapy with three-dimensional conformal boost after the neoadjuvant androgen suppression for patients with locally advanced prostatic carcinoma.

PURPOSE: To analyze the results in patients with locally advanced prostatic carcinoma treated by hormonal therapy followed by external radiotherapy using three-dimensional conformal radiation therapy (3D-CRT) boost. METHODS AND MATERIALS: From 1987 to 1995, 46 patients with histologically proven locally advanced adenocarcinoma of the prostate were treated with 3D-CRT at the National Cancer Center Hospital, Tokyo. The neoadjuvant androgen suppression started immediately after the diagnosis followed by radical radiation therapy, according to the prospective protocol. They were treated with photons of 6-14 MV for wide fields and the boost, of which a multiple-leaf collimator of 2-cm width was available. The boosted dose was delivered with the rotational 3D-CRT, after the delivery of whole pelvis 4-field box from a dose of 40-46 Gy up to 66 Gy. The planning target volume encompassed 1 cm outside throughout the clinical target volume, and the prostate and the seminal vesicles were included in the boost field. RESULTS: The 3D-CRT boost treatment completed as planned in all 46 patients. The median follow-up for all the patients was 60 months (range, 5-120 months). Nineteen of 46 patients died. Of these, 11 patients died of the intercurrent diseases. For all 46 patients, the 5- and 8-year overall survival rates were 61.3% and 42.4%, and the 5- and 8-year cause-specific survival rates were 82.4% and 64.4%, respectively. The prostate-specific antigen (PSA) relapse-free rates for 5- and 8-year were 64.6% and 52.5%, and the clinical local control rates for 5 and 8 years were 75.3% and 69.9%, respectively. The preradiation therapy PSA and the Gleason score were the factors that significantly associated with PSA relapse-free survival. Sixteen of 46 patients (35%) showed at least one form of late toxicities. Of these, 3 patients experienced late complications of Grade 3 (urinary, 2, proctitis, 1). CONCLUSION: The treatment results were fairly good and were consistent with those in Western countries, indicating that this study shows the preliminary status of 3D-CRT for the locally advanced prostate cancer in Japan. Preradiation therapy PSA seems to be a significant predictor of PSA relapse-free survival (p = 0.004) after neoadjuvant androgen suppression.