Comparing Medial Temporal Atrophy Between Early-Onset Semantic Dementia and Early-Onset Alzheimer's Disease Using Voxel-Based Morphometry: A Multicenter MRI Study.

BACKGROUND: Early-onset Semantic dementia (EOSD) and early-onset Alzheimer's disease (EOAD) are often difficult to clinically differentiate in the early stages of the diseases because of the overlaps of clinical symptoms such as language symptoms. We compared the degree of atrophy in medial temporal structures between the two types of dementia using the voxel-based specific regional analysis system for Alzheimer's disease (VSRAD). METHODS: The participants included 29 (age: 61.7±4.5 years) and 39 (age: 60.2±4.9 years) patients with EOSD and EOAD, respectively. The degree of atrophy in medial temporal structures was quantified using the VSRAD for magnetic resonance imaging data. Receiver operating characteristic (ROC) analysis was performed to distinguish patients with EOSD and EOAD using the mean Z score (Z-score) in bilateral medial temporal structures and the absolute value (laterality score) of the laterality of Z-score (| right-left |) for indicating the degree of asymmetrical atrophy in medial temporal structures. RESULTS: The EOSD group had significantly higher Z and laterality scores than the EOAD group (Zscores: mean ± standard deviation: 3.74±1.05 vs. 1.56±0.81, respectively; P<0.001; laterality score: mean ± standard deviation: 2.35±1.23 vs. 0.68±0.51, respectively; P<0.001). In ROC analysis, the sensitivity and specificity to differentiate EOSD from EOAD by a Z-score of 2.29 were 97% and 85%, respectively and by the laterality score of 1.05 were 93% and 85%, respectively. CONCLUSION: EOSD leads to more severe and asymmetrical atrophy in medial temporal structures than EOAD. The VSRAD may be useful to distinguish between these dementias that have several clinically similar symptoms.

Assessment of Amyloid Deposition in Patients With Probable REM Sleep Behavior Disorder as a Prodromal Symptom of Dementia With Lewy Bodies Using PiB-PET.

Introduction: Dementia with Lewy bodies (DLB) often exhibits REM sleep behavior disorder (RBD) at its prodromal stage. Meanwhile, DLB is often comorbid with Alzheimer's disease (AD)-type pathology. In typical AD, amyloid-ß deposition begins considerably before the onset of dementia and has already reached a plateau at the stage of mild cognitive impairment. However, it is not known when amyloid accumulation starts in DLB with AD-type pathology. In the present study, we examined amyloid deposition in patients with RBD as a prodromal symptom of DLB using [11C]-Pittsburgh compound B positron emission tomography (PiB-PET). Methods: The subjects were 12 patients with probable RBD as diagnosed by the Japanese RBD screening questionnaire. They also showed abnormality in 123I-metaiodobenzylguanidine myocardial scintigraphy, a biomarker for DLB. For comparison, 11 patients with probable DLB were included. Applying PMOD software to the PiB-PET images, the global cortical distribution volume ratio was calculated and a ratio >1.3 was regarded as PiB-positive. Results: Two of the RBD patients (16.7%) and eight of the DLB patients (72.7%) were PiB-positive. The amyloid-positive rate was significantly lower in the RBD group than in the DLB group (P = 0.012). Conclusion: The prevalence of amyloid deposition in RBD as a prodromal symptom of DLB was significantly lower than that in DLB, suggesting that amyloid accumulation does not always begin at the early stage of DLB.

Intravenously administered 2'-deoxycytidine suppresses mouse myeloma tumor growth.

We examined the in vivo effects of intravenously administered 2'-deoxycytidine (dCyd) on tumor growth and survival time in mice bearing SP2/0-Ag14 (SP2/0) myeloma tumors. Administration of dCyd tended to decrease the tumor volume and significantly decreased the tumor weight. A single intravenous administration of dCyd significantly increased survival time of the tumor-bearing mice. The effect of dCyd on tumor growth was maintained for at least 1 week after the final administration. The net amount of dCyd in the kidney, liver, and spleen of the tumor-bearing mice increased 2.5 to 5.3 fold compared with the amount in non-tumor-bearing mice. Our results suggest that the increase in dCyd in the mice inoculated SP2/0 myeloma cells plays an important role for the growth suppression of the tumor.

2'-Deoxycytidine decreases the anti-tumor effects of 5-fluorouracil on mouse myeloma cells.

2'-Deoxycytidine (dCyd), a pyrimidine nucleoside found at high concentrations in the plasma of cancer patients with a poor prognosis after chemotherapy, is considered to be a biomarker for breast cancer. 5-Fluorouracil (5FU) is a nucleoside analog and is used as an anti-tumor agent in patients whose plasma dCyd concentrations are increased. Because both dCyd and 5FU are pyrimidine analogues, it is possible that they have pharmacokinetic/ pharmacodynamic interaction, by which the anti-cancer efficacy of 5FU would be reduced. Here, we examined the effects of dCyd on the cytotoxicity of 5FU on mouse myeloma SP2/0-Ag14 (SP2/0) cells lacking hypoxanthine-guanine-phosphoribosyl transferase (HGPRT) and RH4 hybridomas with HGPRT under asynchronized conditions. The reduced cell viability by 5FU was restored by co-, but not pre-, treatment of dCyd in both SP2/0 and RH4 cells, but this effect in the former tended to be greater than that in the latter, suggesting a possible involvement of HGPRT in the interaction, although this might not be a major mechanism. Moreover, dCyd administration to SP2/0 myeloma-bearing mice tended to shorten their 5FU-induced prolonged survival in vivo. Collectively, these results indicate that dCyd decreases the anti-tumor efficacy of 5FU and that a metabolic pathway via HGPRT is involved partially in this interaction. The evaluation of dCyd as a biomarker is believed to provide valuable information for effective and safe chemotherapy with 5FU.

S-sulfonation of transthyretin is an important trigger step in the formation of transthyretin-related amyloid fibril.

Senile systemic amyloidosis and familial amyloid polyneuropathy are caused by oxidative deposition of conformationally altered transthyretin (TTR). We identified oxidative modification of the 10th cysteine of TTR through S-sulfonation in vitro. Based on mass spectrometric analysis, we determined the spectrophotometric, western blotting, and fluorescent microscopic properties of TTR incubated with and without cysteine-S-sulfonate in acidic (pH 4) and alkaline (pH 8) conditions at 37 degrees. The absorption of the aggregated TTR molecules increased more with incubation time and the concentration of cysteine-S-sulfonate at pH 4 than at pH 8. The Congo red binding to the S-sulfonated TTR at pH 4 was saturated with an apparent Bmax of 2.01 mol per mole of the S-sulfonated TTR and apparent KD of 7.75x10(-6) M. On the other hand, the Bmax of cysteinyl TTR was 1.38, and its KD was 3.52x10(-6) M while the Bmax of reduced TTR was 0.86, and its KD was 2.86x10(-6) M. Moreover, we detected positive amyloid fibril staining using Thioflavin T and Congo red with the S-sulfonated TTR but not with untreated or reduced TTR by microscopic fluorescent analysis. After modification of TTR in vitro, oligomers resisted reduction and denaturation was irreversibly induced, and which contributed differences in the Western blotting patterns obtained with four anti-TTR antibodies. In conclusion, this study showed that the formation of S-sulfonation of TTR through oxidative modifications of the thiol residue on the 10th cysteine of TTR is an important trigger step in the formation of transthyretin-related amyloid fibril.

Changes in 2'-deoxycytidine levels in various tissues of tumor-bearing mice.

The nucleoside 2'-deoxycytidine (dCyd) increases in the plasma of cancer patients with poor prognoses. 5-fluorouracil (5FU) is one of the anti-cancer agents used in chemotherapy for patients whose plasma dCyd is elevated. We examined the free dCyd level in various tissues of mice, with and without tumors, and in mice with and without the administration of 5FU or of dCyd, and investigated the effects of dCyd in tumor-bearing animals. SP2/0-Ag14 mouse myeloma cells were transplanted subcutaneously into mice and 5FU or dCyd was administered intraperitoneally. Free dCyd was measured in blood and tissues by HPLC at two time points, once when mouse body weight was maximally decreased (1 day after the last administration of 5FU, day 16) and again when it returned to control level at 1 week after the last 5FU treatment (day 22). Results showed that in tumor-bearing mice, the level of dCyd (per g wet weight) increased in the spleen. The change in liver weight caused by the administration of 5FU correlated with the level of dCyd in the liver. Notably, the relative tumor volume and tumor weight was decreased in dCyd-treated animals in comparison with controls. In conclusion, the levels of dCyd were markedly altered in the tissues of the reticuloendothelial and lymphatic systems, such as liver and spleen, and dCyd apparently had the ability to inhibit tumor growth in the body.

[Oxidative precipitation of transthyretin--a clue to elucidate amyloidosis].

Common precipitation of amyloid proteins occurs in amyloidosis such as dementia and prion diseases. The precipitates are characteristically stained with congo red, although it is not yet known why this occurs. We have found a kind of amyloid protein, transthyretin(TTR), that is S-sulfonated at the residue of cysteine in blood from patients with deficiency of molybdenum cofactor, which is essential to sulfite oxidase. The S-sulfonated TTR was easily stained with congo red, whereas TTR itself was not. Further, the S-sulfonated TTR slowly made fibrils from precipitates, which is well known as a common characteristic in amyloidosis. It seemed an important clue that S-sulfonation is as the starting reaction to induce the precipitation of the protein. We have studied reaction products from TTR and its synthetic octapeptide around the cysteine residue under moderate conditions by mass-spectrometry and estimated the reaction mechanism. Another clue is chinoform(or, clioquinol), which is effective for patients with dementia. The mechanism was assumed that metal ions such as copper and iron involved in oxidative precipitation of the proteins were masked with chinoform. In 1970, Yoshioka, one of the authors, found that in green urine from a patient with SMON disease was identified a chelate compound of chinoform with ferric ion. This finding led to the resolution of SMON disease caused by excess dose of chinoform. In this review, we discuss amyloidosis related with SMON disease.

Modification of cysteine residue in transthyretin and a synthetic peptide: analyses by electrospray ionization mass spectrometry.

Cysteine and cystine in protein are modified to various derivatives in vitro and in vivo. By electrospray ionization mass spectrometry (ESI-MS), we previously found derivatives of serum transthyretin (TTR) in which cysteine residue at position 10 was changed to glycine residue and sulfocysteine residue. The change, cysteine to glycine, was unique and the origin of the sulfonic acid group was controversial. In the present paper, we show the molecular masses of various TTR derivatives including these two, and the modification process was studied using a synthetic peptide with the same sequence as cysteine containing part of TTR, i.e., SKCPLMVK. After incubation of the peptide at pH 8.3, various derivatives were generated, which showed changes of cysteine residue to glycine, dehydroalanine, S-thiocysteine, and S-sulfocysteine residues, which were confirmed by molecular mass and collision-induced dissociation spectra. Dehydroalanine may react with other amino acids and contribute to form cross-linking fibril, causing amyloidosis.

A screening system of prodrugs selective for MAO-A or MAO-B.

We synthesized several prodrugs of glycine and gamma-aminobutyric acid. In order to establish a screening system from the prodrugs of selective activity to MAO-A or MAO-B, we examined purification conditions such as solubilization with Triton X-100, precipitation with ammonium sulfate, gel filtration and anion exchange chromatography. MAO-B was purified from various tissues such as guinea pig brain, kidney and spleen. MAO-A from human placenta without MAO-B was unstable in above purifications and used as crude. At each purification step, we checked sensitivity of the enzyme to specific inhibitors by developing a convenient fluorescence assay, in which hydrogen peroxide produced by the enzyme was reacted with p-hydroxyphenylpropionic acid. A fluorescence microplate reader measured a fluorescence of the fluorescent product from p-hydroxyphenylpropionic acid with horseradish peroxidase. In comparison with milacemide, N,N-bis(carbamoylmethyl)-N-pentylamine was the best and exclusive substrate for MAO-B. 2-N-(phenylethylamino)-acetoamide was the good substrate for MAO-A and MAO-B same as milacemide. 4-N-(n-pentylamino)-butyric acid and 4-(N-phenylethylamino)-butyric acid were the moderate substrates for both enzymes, which should release gamma-aminobutyric acid. These drugs will be new leading compounds.

The peak height ratio of S-sulfonated transthyretin and other oxidized isoforms as a marker for molybdenum cofactor deficiency, measured by electrospray ionization mass spectrometry.

Molybdenum cofactor deficiency is a fatal neurological disorder, which follows an autosomal-recessive trait and is characterized by combined deficiency of the enzyme, sulfite oxidase, xanthine dehydrogenase and aldehyde oxidase. Early detection of molybdenum cofactor-deficient patients is essential for their proper care and genetic counseling of families at risk. We demonstrate the use of S-sulfonated transthyretin (TTR) as a marker for molybdenum cofactor deficiency. Plasma or sera obtained from 4 patients with molybdenum cofactor deficiency and 57 controls were studied by electrospray ionization mass spectrometry (ESIMS) following selective enrichment of TTR by immunoprecipitation using protein G/A agarose. The data obtained from molybdenum cofactor deficiency samples indicated a strong increase in the peak height of S-sulfonated TTR. A more significant difference was revealed if the peak height ratio of S-sulfonated TTR and the sum of the other oxidized TTR were determined. By accurate determination of the ratio, the samples of molybdenum cofactor deficiency patients could clearly be distinguished from controls without molybdenum cofactor deficiency.