Sumiko Nissato

Novel Germline Mutations in the SDHB and SDHD Genes in Japanese Pheochromocytomas

The SDHA, SDHB, SDHC, and SDHD genes code for subunits of succinate dehydrogenase (SDH), which forms part of the mitochondrial respiratory chain. Germline mutations in the genes encoding SDHB and SDHD have been reported in familial paragangliomas/pheochromocytomas and in apparently sporadic pheochromocytomas. SDHB and SDHD mutations are widely distributed along the genes with no apparent hot spots. SDHB mutations are often detected in malignant and extra-adrenal pheochromocytomas. SDHD mutations are also detected frequently in head and neck paragangliomas. We sequenced the entire coding regions of the SDHB and SDHD genes in 17 pheochromocytomas. Weidentified novel heterozygous G to A point mutations at the first base of intron 3 of the SDHB gene in a malignant extra-adrenal abdominal pheochromocytoma patient, and at the first base of codon 111 of the SDHD gene in an adrenal pheochromocytoma patient. Further, we confirmed the SDHD mutation by DHPLC. The prevalence of SDHB and SDHD mutations in pheochromocytomas we examined was 12% (2/17). Thus, we identified two novel SDH mutations in Japanese pheochromocytomas. Further studies will investigate the oncogenic potential of these mutations.

Expression of mRNAs for PACAP and its receptor in human neuroblastomas and their relationship to catecholamine synthesis.

PURPOSE Pituitary adenylate cyclase-activating polypeptide (PACAP), a member of the secretin/glucagon/vasoactive intestinal peptide family, induces the expression of catecholamine-synthesizing enzymes in adrenal medullary cells. In addition, PACAP and its receptor have been detected in human neuroblastoma tissues and cell lines, though it is not yet known whether PACAP enhances the expression of genes encoding catecholamine-synthesizing enzymes. To address this question, we analyzed PACAP, PACAP receptor and tyrosine hydroxylase (TH) mRNAs in neuroblastomas. METHODS The levels of mRNA for PACAP and vasoactive intestinal peptide (VIP), as well as their receptors and the mRNA for TH were measured by RT-PCR or real-time PCR analysis. RESULTS VPAC1R mRNA was detected in all of 16 tissues and 3 cell lines that were examined, while VPAC2R mRNA was detected in 5 of 16 (31%) tissue and 2 of 3 cell lines. PAC1R mRNA was detected in 6 out of 16 (38%) tissues and none of 3 cell lines. mRNA expression of PACAP and TH were detected in many tissues (10/16 and 16/16, respectively). However, neither in tissues nor cell lines did PACAP mRNA expression correlate with TH mRNA expression. CONCLUSION Our findings suggest that PACAP is not involved in the regulation of expression of TH in neuroblastomas.

Identical germline mutations in the TMEM127 gene in two unrelated Japanese patients with bilateral pheochromocytoma

Recently, TMEM127 was shown to be a new pheochromocytoma susceptibility gene; this is consistent with its function as a tumour suppressor gene (Journal of Clinical Endocrinology and Metabolism, 2009, 94, 2817). Most pheochromocytomas arise from the adrenal medulla, and in approximately half of the cases, the tumours are bilateral (Journal of Clinical Endocrinology and Metabolism, 2009, 94, 2817; Journal of the American Medical Association, 2004, 292, 943; Human Mutation, 2010, 31, 41; Science, 2009, 325, 1139). The aim of the present study was to determine whether TMEM127 mutations are involved in the pathogenesis of pheochromocytomas/paragangliomas in Japanese subjects.

A common genetic variant of the chromogranin A-derived peptide catestatin is associated with atherogenesis and hypertension in a Japanese population

Chromogranin A (CHGA) is a major protein in the secretory granules of chromaffin cells. CHGA also gives rise to cardiovascular/metabolism regulatory peptides, such as catestatin (CST) and pancreastatin (PST). While CST is a potent inhibitor of catecholamine secretion, PST is a potent physiological inhibitor of glucose-induced insulin secretion. Recently, several SNPs were identified in the CST and PST domains of CHGA locus in different populations. Among the discovered SNPs, CST variant allele Ser-364 was associated with blood pressure alteration and PST variant allele Ser-297 was associated with significantly higher plasma glucose level. In this study, we examined whether these CST and PST variant alleles exist and influence cardiovascular and metabolic phenotypes in Japanese population. Our study comprised of 343 Japanese subjects aged 45-85 years (143 men and 200 women, mean age 66 ± 8 years). We determined the genotypes of CST and PST by PCR-direct sequencing method and carried out genotype-phenotype association analysis. In 343 participants, the minor allele frequency of CST variant Ser-364 was 6.10%. On the other hand, we did not detect the PST variant Ser-297 in this entire study population. The presence of Ser-364 allele was associated with increased in baPWV (an index of systemic arterial stiffness) that suggests an initiation and/or progression atherogenesis and hypertension. The Ser-364 allele was also associated with elevated systolic blood pressure and pulse pressure, consistent with increased baPWV. In conclusion, the CST Ser-364 allele may increase the risk for cardiovascular diseases in Japanese population.

Expression of mRNAs for succinate dehydrogenase subunits and related genes in pheochromocytoma.

Abstract:  Mutations in the genes encoding succinate dehydrogenase (SDH) have been associated with susceptibility to pheochromocytoma. However, few reports have examined the level of SDH mRNAs expression. In this study, we examined the level of expression of mRNAs encoding SDHB, SDHC, and SDHD in pheochromocytoma, pheochromocytoma subgroups, and normal adrenal gland, and compared the expression of these genes to the level of expression of related genes in the same tissues. The mean relative level of expression of SDHB, SDHC, SDHD and VHL mRNA was 28.7 ± 6.2%, 16.6 ± 4.8%, 214 ± 47.5% and 25.9 ± 8.2%, respectively, in pheochromocytoma tissues compared to normal adrenal gland. Furthermore, the mean relative level of the RET proto‐oncogene mRNA was 707 ± 149% in pheochromocytoma compared to normal adrenal gland. The level of expression of the SDH genes was highly correlated in each individual sample (P < 0.0001). The level of expression of the SDH mRNAs correlated with the level of VHL mRNA (P < 0.0001), but not with the level of RET mRNA. The level of SDH mRNAs expression also correlated with the expression of phenylethanolamine N‐methyl transferase (PNMT), an adrenaline synthesizing enzyme (P < 0.01), which may explain the correlation between SDH expression and adrenaline content (P < 0.05). The level of SDH mRNAs expression correlated strongly with the expression of VEGF mRNA (P < 0.0001). In multiple endocrine neoplasia (MEN) 2a, the expression of the SDH genes and VHL mRNA was significantly higher than that observed in adrenal or extra‐adrenal pheochromocytoma. The expression of the corticotropin‐releasing hormone (CRH) mRNA was significantly higher in extra‐adrenal pheochromocytoma than in adrenal pheochromocytoma or MEN2a. Thus, tumor‐specific gene expression exists in pheochromocytoma, which may explain the characteristics of the tumor.

A linkage study with DNA markers (D4S95, D4S115, and D4S111) in Japanese Huntington disease families

SummaryAttempts to isolate the Huntington disease (HD) gene based on its position have been frustrated by apparently contradictory recombination events in HD pedigrees that have predicted two non-over-lapping candidate regions: 100 kb at the telomere of the short arm of chromosome 4, and a 2.2 Mb region located internally at 4p16.3. The proximal location is also supported by the detection of a linkage disequilibrium between HD and some restriction fragment length polymorphisms (RF-LPs) at the D4S95, D4S98, and D4S127 loci. In the present study, a proximal marker D4S95 showed tight linkage to the disease locus in Japanese pedigrees (Zmax=3.31, θmax=0.00), while distal markers D4S115 and D4S111 did not. Particularly, a two point linkage analysis between D4S111 and HD yielded a lod score −2.01 for θ=0.015. This result leads to the exclusion, as a possible region of localization of the HD gene, of more than 3 cM of the genome around D4S111 locus. At the same time our results favor aforementioned proximal location as a candidate location for the HD gene.