Genome-wide association study of degenerative bony changes of the temporomandibular joint.

OBJECTIVES: To identify susceptibility genes underlying degenerative bony changes of the temporomandibular joint (TMJ). MATERIALS AND METHODS: Bony changes of the TMJ condylar head were diagnosed by examination of panoramic radiographs and/or magnetic resonance images and/or computed tomography images. We conducted a genome-wide association study (GWAS) of 146 cases with TMJ degeneration and 374 controls from East Asian populations using an Illumina HumanOmniExpress BeadChip. After rigorous quality-control filtering, approximately 550,000 single nucleotide polymorphisms (SNPs) were used for tests of associations with disease status. RESULTS: Forty-one SNPs at 22 independent loci showed association signals at P < 1 × 10(-4). The SNP rs878962, which maps on an intron of TSPAN9 on chromosome 12, showed the strongest association (combined OR = 1.89, 95% confidence interval = 1.43-2.50, P = 8.1 × 10(-6)). According to in silico predictions of the 41 SNPs, two intronic SNPs of APOL3 (rs80575) and MRC2 (rs2460300) may fall within regulatory elements and affect DNA-protein interactions. We could not replicate SNPs located on genes that have been reported to be associated with temporomandibular disorder or temporomandibular osteoarthritis in previous studies at P < 1 × 10(-4). CONCLUSIONS: Our GWAS identified 22 independent loci showing suggestive association signals with degenerative bony changes of the TMJ. These loci provide good candidates for future follow-up studies.

Positional effects of polymorphisms in probe-target sequences on genoplot images of oligonucleotide microarrays.

Single nucleotide polymorphisms (SNPs) present in probe-target sequences (SPTS) have been shown to be associated with abnormal genoplot images. We explored the effects of SPTS positions on genoplot images using a data set from a genome-wide association study typed on an Illumina Human Hap300 platform. We screened the physical genomic positions of 308,330 autosomal probes to identify SPTS candidates deposited in dbSNP. The genoplot images across 293 individuals were inspected further in SNPs bearing an SPTS candidate. We identified 35,185 SNPs bearing a single SPTS candidate, including 264 SNPs showing abnormal genoplot images. The frequencies of SPTS at distances within 10 bases from the target SNP were significantly higher in the 264 SNPs showing abnormal genoplot images, than in the remaining 34,921 SNPs (49.62 vs 12.87%; Fisher exact test; P = 2.2 x 10(-16)). Of these 264 SNPs, we randomly selected 20 SNPs and resequenced them in 97 individuals. An SPTS within 10 bases of the target SNP was confirmed in all 20 SNPs, except for one SNP with a small deletion (7 bases) in the probe-target sequence. Taken together, these results suggest an association of a proximal SPTS with an abnormal genoplot image, which could result in spurious genotype detections, highlighting the importance of minimizing systematic errors in microarray experiments.

Gh: a GTP-binding protein with transglutaminase activity and receptor signaling function.

The alpha 1-adrenergic receptors activate a phospholipase C enzyme by coupling to members of the large molecular size (approximately 74 to 80 kilodaltons) G alpha h family of guanosine triphosphate (GTP)-binding proteins. Rat liver G alpha h is now shown to be a tissue transglutaminase type II (TGase II). The transglutaminase activity of rat liver TGase II expressed in COS-1 cells was inhibited by the nonhydrolyzable GTP analog guanosine 5'-O-(3-thiotriphosphate) or by alpha 1-adrenergic receptor activation. Rat liver TGase II also mediated alpha 1-adrenergic receptor stimulation of phospholipase C activity. Thus, G alpha h represents a new class of GTP-binding proteins that participate in receptor signaling and may be a component of a complex regulatory network in which receptor-stimulated GTP binding switches the function of G alpha h from transglutamination to receptor signaling.

Mint3-mediated L1CAM expression in fibroblasts promotes cancer cell proliferation via integrin α5ß1 and tumour growth.

Fibroblasts are some of the major cells in tumour tissues that influence tumour progression and drug resistance. However, our understanding on fibroblast-mediated tumour malignancy remains incomplete. Munc18-1-interacting protein 3 (Mint3) is known as an activator of hypoxia-inducible factor-1 (HIF-1) even during normoxia in cancer cells, macrophages and fibroblasts. Although Mint3 promotes ATP production via glycolysis by activating HIF-1 in cancer cells and macrophages, the biological role of Mint3-mediated HIF-1 activation in fibroblasts remains unclear. To address this, we examined whether Mint3 in fibroblasts contributes to tumour growth. Mint3 depletion in mouse embryonic fibroblasts (MEFs) decreased tumour growth of co-injected human breast cancer cells, MDA-MB-231 and epidermoid carcinoma A431 cells in mice. In MEFs, Mint3 also promoted cancer cell proliferation in vitro in a cell-cell contact-dependent manner. Mint3-mediated cancer cell proliferation depended on HIF-1, and further gene expression analysis revealed that the cell adhesion molecule, L1 cell adhesion molecule (L1CAM), was induced by Mint3 and HIF-1 in fibroblasts. Mint3-mediated L1CAM expression in fibroblasts stimulated the ERK signalling pathway via integrin α5ß1 in cancer cells, and promoted cancer cell proliferation in vitro and tumour growth. In cancer-associated fibroblasts (CAFs), knockdown of MT1-MMP, which promotes Mint3-mediated HIF-1 activation, or Mint3 decreased L1CAM expression. As MEFs, CAFs also promoted cancer cell proliferation in vitro, and tumour growth via Mint3 and L1CAM. In human breast cancer specimens, the number of fibroblasts expressing L1CAM, Mint3 and MT1-MMP was higher in cancer regions than in adjacent benign regions. In addition, more phospho-ERK1/2-positive cancer cells existed in the peripheral region surrounded by the stroma than in the central region of solid breast cancer nest. Thus, Mint3 in fibroblasts might be a good target for cancer therapy by regulating cancer cell-stromal cell communication.

Effect of beta-adrenergic receptor blockade on atrial natriuretic peptide in essential hypertension.

Plasma levels of atrial natriuretic peptide (ANP) were measured in 32 untreated subjects with essential hypertension and in 31 patients undergoing long-term treatment with beta-blockers. Patients receiving beta-blockers had significantly higher mean plasma ANP levels (72.0 +/- 36.0 [SD] pg/ml) than did untreated hypertensive subjects (39.8 +/- 15.8 pg/ml; p less than 0.01) and healthy normotensive controls (33.9 +/- 16.6 pg/ml; n = 61, p less than 0.01), while the mean plasma ANP concentration in untreated hypertensive subjects was not statistically different from that in control subjects. Administration of atenolol, 50 mg/day, for 4 weeks to 10 untreated subjects resulted in a significant (p less than 0.001) rise in plasma ANP levels (from 38.8 +/- 9.5 to 68.7 +/- 20.6 pg/ml). In 31 patients undergoing long-term treatment with beta-blockers, multivariate regression analysis revealed that age, pretreatment mean blood pressure, and plasma concentration of cyclic 3',5'-guanosine monophosphate (cGMP) were significant predictors of plasma ANP levels. These results suggest that beta-adrenergic receptor blockade in patients with essential hypertension elevates plasma ANP levels with a concomitant rise in cGMP concentrations, and that increased ANP in plasma may play a role in the compensatory mechanism that operates in response to beta-adrenergic receptor blockade.

Plasma levels of atrial natriuretic peptide in primary aldosteronism and essential hypertension.

Plasma levels of atrial natriuretic peptide (ANP) were measured in 9 patients with primary aldosteronism and 41 patients with essential hypertension (class I or II by WHO classification) using a specific and sensitive RIA. The mean plasma ANP concentration in patients with primary aldosteronism (mean +/- SEM, 67.1 +/- 10.8 pg/ml; n = 9) was significantly higher than that in healthy normotensive subjects (37.9 +/- 1.4 pg/ml; n = 108) or patients with essential hypertension (38.5 +/- 2.8 pg/ml; n = 41). During treatment with spironolactone, plasma levels of ANP declined in 6 of the 7 patients with primary aldosteronism, but no change occurred in the remaining patient who had cardiac enlargement of unknown etiology. The mean plasma ANP concentration in patients with essential hypertension, on the other hand, was not significantly different from that in normal subjects. These results indicate that plasma ANP levels are elevated in patients with primary aldosteronism, probably due to volume expansion, whereas no abnormality in ANP secretion exists in patients with uncomplicated essential hypertension.

Serum pseudouridine as a biochemical marker in patients with hepatocellular carcinoma.

The concentration of serum pseudouridine, a degradation product of transfer ribonucleic acid, was determined by high-performance liquid chromatography in patients with hepatocellular carcinoma, liver cirrhosis, other benign hepatobiliary diseases, and healthy controls. The serum pseudouridine concentration in patients with hepatocellular carcinoma was significantly higher than that in patients with cirrhosis or the controls. Twenty-seven (51.9%) of 52 patients with hepatocellular carcinoma had serum pseudouridine concentrations higher than the mean value for healthy controls plus 2 SD. Fourteen of the 36 patients who had serum alpha-fetoprotein levels below 400 ng/ml, had elevated serum pseudouridine concentration. In total, 36 of the 52 patients (69.2%) could be detected by combination of these two markers. Two patients who had developed hepatocellular carcinoma during the course of cirrhosis and were continuously negative for alpha-fetoprotein, had higher levels of the pseudouridine concentration when hepatocellular carcinoma occurred. Furthermore, 4 of the 7 patients who had a very small cancer and were negative for alpha-fetoprotein, had elevated serum pseudouridine concentration. These results indicate that serum pseudouridine is a useful biochemical marker and that serum pseudouridine and alpha-fetoprotein in combination are considered to serve as complementary markers, for the diagnosis of hepatocellular carcinoma.

Studies on a beta-migrating high density lipoprotein.

An unusual serum lipoprotein (Lp) profile was detected in a Japanese family. A double beta-Lp was observed when serum was subjected to polyacrylamide gel electrophoresis. The slower migrating beta-Lp was identified as a subfraction of high density lipoprotein (HDL). It was present in the d 1.063--1.21 fraction, migrated to the position designated as the midband L.1 (sinking pre-beta-lipoprotein) by Mead, M.G. and Dangerfield, W.G. (1974) (Clin. Chim. Acta 51, 173--182) [1], and reacted against human anti-beta-Lp antiserum. This lipoprotein contained greater amounts of triglyceride than the usual beta-lipoprotein and could not be clearly detected by paper electrophoresis. Individuals exhibiting this high density midband lipoprotein appeared to be heterozygous for an autosomal dominant gene. Although other reports have indicated the possibility of a positive association between the occurrence of serum lipoproteins with unusual eletrophoretic mobility and premature ischemic heart disease, no such correlation was demonstrable in these subjects.

Bilateral vermilion flaps for lower lip repair.

A more natural reconstructive procedure of the lower lip using bilateral vermilion flaps was applied in five patients with excellent results. The vermilion defects were about two-fifths to three-fifths. In three patients, the vermilion defect was repaired using bilateral vermilion flaps alone. In the remaining two patients, a narrow horizontal lip defect was repaired by bilateral vermilion flaps and a subcutaneous V-Y advancement flap of the lower lip. A single vermilion flap or bilateral vermilion flaps are considered to be of great value for vermilion reconstruction because of the inherent elasticity and common anatomic unit. The postoperative scars are not remarkable at all. A long and narrow horizontal lip defect (perhaps within 1.5 cm downward from the vermilion border) may be effectively repaired by the combination of vermilion flap(s) and a V-Y advancement flap without sacrificing any additional healthy tissue.

Development of a real-time hand dose monitor for personnel in interventional radiology.

Medical procedures denoted as interventional radiology require operation near an X ray beam, which brings high dose exposures to the operators' hands. For the effectual control of their extremity doses, a prototype of a real-time wrist dosemeter has been developed, hand dose monitor (HDM), based on a single silicon detector. Experiments were performed to test its response to diagnostic X rays. The HDM was highly sensitive and showed a linear response down to doses of a few tens of microsieverts. Though dose rate, energy and angular dependence of the response were observed in some extreme conditions, the HDM was proved to be of practical use if it was appropriately calibrated. Since an HDM enables personnel to check their hand doses on a real-time basis, it would enable medical staff to control the exposure themselves.

[Behavior of Trp-P-1 and its metabolites in rat excreta].

To study of the behavior of Trp-P-1 and its metabolites in rat feces and urine, rats were orally administered with Trp-P-1 (750, 1,500 and 2,500 micrograms/rat), and excreted Trp-P-1 was analyzed using HPLC assay and bacterial mutagenicity assay. The extraction of Trp-P-1 from urine was performed by using the chloroform extraction method, and blue rayon was used for the extraction from feces. When Trp-P-1 was added to rat feces and urine, the recoveries of Trp-P-1 were 85.9 +/- 3.9% and 91.3 +/- 3.7%, respectively. The extracts of feces and urine from rats administered with Trp-P-1 were individually fractionated by thin layer chromatography on C18 gel. The major mutagenic zone corresponding to Trp-P-1 was found at Rf 0.09 in both extracts, while the feces extract gave two additional mutagenic zones at Rf 0.15 and 0.20. More than 97% of the fecal mutagenic activity was due to unchanged Trp-P-1. In rats administered with 750 micrograms of Trp-P-1, the amount of extracted Trp-P-1 and the number of His+ colonies induced by whole excreta were 81.6 +/- 7.1 micrograms (n = 6) and (432 +/- 77) x 10(4) for feces, and 28.7 +/- 4.9 micrograms and (171 +/- 28) x 10(4) for urine. The recoveries of Trp-P-1 in the feces and urine were 10.8 +/- 0.9% and 3.8 +/- 0.7% by HPLC analysis, and 11.1 +/- 2.0% and 4.4 +/- 0.7% by mutagenicity assay respectively. The results of the two assays seemed to show similar patterns of recovery.

[Assessment of hydroxylated metabolites of polychlorinated biphenyls and polychlorinated dibenzofurans as potential estrogens by yeast two-hybrid system].

The estrogenic activities of several hydroxylated metabolites of PCBs and PCDFs were investigated by yeast two-hybrid assay based on the ligand-dependent interaction of estrogen receptor with coactivator. For the hydroxylated PCBs, the order of estrogenic potency was 4-OH-2',4',6'-triCB > 4-OH-4'-monoCB, 4-OH-biphenyl. These compounds were evaluated as 10(3) to 10(4) less potent than 17 beta-estradiol based on the concentrations of test compounds showing 10% activity of 10(-7) M 17 beta-estradiol. 2-OH-3',4,4'-triCB, 4-OH-2',3,4'-triCB and 3-OH-/4-OH-2,2',5,5'-tetraCB, the metabolites of 2,2',5,5'-tetraCB were inactive as estrogens at the highest concentrations used in this study (10(-5) M). Also 4-OH-3,3',4',5-tetraCB, the metabolite of 3,3',4,4'-tetraCB was inactive as estrogen, indicating that this hydroxylated metabolite did not take part in the estrogenic activity of 3,3',4,4'-tetraCB. OH group at 4-position of biphenyl was necessary for the expression of estrogenicity, but one or two chloro-substitution adjacent to OH group inhibited the activity. For the hydroxylated PCDFs, 8-OH-2-monoCDF, 7-OH-3,4-diCDF, 8-OH-3,4-diCDF, 8-OH-3,4,6-triCDF and 3,8-(OH)2-2-monoCDF exhibited estrogenic activity. The estrogenic activity of 3,8-(OH)2-2-monoCDF was comparable to those of 4-OH-2',4',6'-triCB and 4-nonylphenol (mixture of compounds with branched sidechain). The order of activity was 3,8-(OH)2-monoCDF > 8-OH-3,4-diCDF, 7-OH-3,4-diCDF > 8-OH-2-monoCDF, 8-OH-3,4,6-triCDF. These compounds were evaluated as 2.5 x 10(3) to 3 x 10(4) less potent than 17 beta-estradiol. On the other hand, no estrogenic activity was observed for 2-OH-dibenzofuran, 3-OH-2,8-diCDF, 6-OH-3,4-diCDF and 9-OH-3,4-diCDF at concentrations as high as 10(-4) M. Substitution of OH group at 2(8)- or 3(7)-position of dibenzofuran and no chloro-substitution adjacent to OH group was required for the estrogenic activity.

The use of link provider data to improve national genetic evaluation across weakly connected subpopulations.

Data from 3 prefectures and a nationwide farming corporation were used to assess the usefulness of the "link provider data" in providing indirect genetic links for the national genetic evaluation for carcass weight across weakly connected subpopulations of the Japanese Black cattle. The data from the farming corporation provided genetic links to those of all prefectures and was therefore used as the link provider data. Two national genetic evaluation strategies under an animal model were compared, based on the generalized coefficient of determination (CD) of contrasts between mean EBV of sires or maternal grandsires (MGS) from different prefectures: strategy PA-1 was a pooled analysis of the data sets of the 3 prefectures, and strategy PA-2 was a pooled analysis of the data sets of the 3 prefectures and the farming corporation. The CD of the contrasts were greater for PA-2 than for PA-1. Under PA-2, the CD of the contrasts between mean EBV of sires or MGS ranged from 0.67 to 0.78 or from 0.61 to 0.70, respectively. Pooling the data from the 3 prefectures and the farming corporation increased the degree of connectedness through the link provider data rather than the amount of information by adding more data, thus improving the accuracy of prediction. The differences between mean EBV of sires or MGS from different prefectures were smaller for PA-1 than for PA-2. This finding suggests that genetic differences in carcass weight among prefectures are present, but that they would be confused with the environmental differences under PA-1 because of the lack of genetic connectedness among the prefectures. On the other hand, the genetic differences among the prefectures would be predicted precisely under PA-2 because the genetic connectedness among the prefectures was improved by using the link provider data. The results demonstrate that the link provider data could be used to unify within-prefecture evaluation to form a Japanese national genetic evaluation across weakly connected subpopulations.