Molecular genetics of familial amyloidotic polyneuropathy

We established a diagnosis of familial amyloidotic polyneuropathy (FAP) based on DNA and demonstrated a direct link between prealbumin gene mutation and FAP. The individuals with FAP, so far examined, were heterozygous for the prealbumin gene, carrying one normal and one mutant gene. To investigate the molecular pathogenesis of FAP, we isolated a normal prealbumin gene (7 kb in length) and also a mutant prealbumin gene associated with FAP. We compared their nucleotide sequences and found that they matched except for a single-base substitution present in the second exon. In an effort to construct mouse model systems for the FAP, we developed strains of transgenic mice carrying human mutant prealbumin genes.     

Organelle specific simultaneous Raman/green fluorescence protein microspectroscopy for living cell physicochemical studies

We demonstrate a novel bio‐spectroscopic technique, “simultaneous Raman/GFP microspectroscopy”. It enables organelle specific Raman microspectroscopy of living cells. Fission yeast, Schizosaccharomyces pombe, whose mitochondria are green fluorescence protein (GFP) labeled, is used as a test model system. Raman excitation laser and GFP excitation light irradiate the sample yeast cells simultaneously. GFP signal is monitored in the anti‐Stokes region where interference from Raman scattering is negligibly small. Of note, 13 568 Raman spectra measured from different points of 19 living yeast cells are categorized according to their GFP fluorescence intensities, with the use of a two‐component multivariate curve resolution with alternate least squares (MCR‐ALS) analysis in the anti‐Stokes region. This categorization allows us to know whether or not Raman spectra are taken from mitochondria. Raman spectra specific to mitochondria are obtained by an MCR‐ALS analysis in the Stokes region of 1389 strongly GFP positive spectra. Two mitochondria specific Raman spectra have been obtained. The first one is dominated by protein Raman bands and the second by lipid Raman bands, being consistent with the known molecular composition of mitochondria. In addition, the second spectrum shows a strong band of ergosterol at 1602 cm^?1, previously reported as “Raman spectroscopic signature of life of yeast.”     

EIevated serum level of Thioredoxin in patients with Hepatocellular Carcinoma

Thioredoxin (TRX) is known to contain an active site with aredox-active disulfide and has various biological activities. The objectiveof the present study was to investigate whether circulating TRX levels areelevated in patients with chronic hepatitis (CH) or liver cirrhosis (LC) andhepatocellular carcinoma (HCC). An anti-TRX monoclonal antibody andpolyclonal antibodies that specifically recognize TRX, were generated andused for the development of an ELlSA system to measure TRX levels in humanserum. The geometric mean and its 95% confidence interval of serumlevel of TRX in healthy volunteers was 81.75 ng/ml (74.60-89.59 ng/ml). Theserum level of TRX in LC/CH patients without HCC was 80.87 ng/ml(69.66-93.88 ng/ml). The value was not statistically different from that inserum from normal volunteers (p=0.69). In contrast, the serum level of TRXin patients with HCC was 147.35 ng/ml (125.53-1 72.96 ng/ml), which wassignificantly higher when compared with the level in serum of normalvolunteers (p<0.001) and in serum of LC/CH patients without HCC(p<0.001). In four patients with HCC, the initially high level of serum TRX(>150 ng/ml) decreased below 150 ng/ml after surgical removal of the tumor.The data reported herein revealed that patients with HCC had a significantlyelevated serum level of TRX, suggesting that measurement of serum of TRXmight be a useful clinical parameter when HCC is suspected.