Temporomandibular disorder is associated with a serotonin transporter gene polymorphism in the Japanese population

Background  

Selective serotonin-reuptake inhibitors (SSRIs) are commonly prescribed for the treatment of depression and can be used as nonhormonal alternatives to manage hot flashes for women with a history of breast cancer and unable to take hormone replacement therapy. There are, however, few reports on the efficacy of SSRIs for the treatment of natural postmenopausal climacteric symptoms. In this pilot study, we evaluate the SSRI, fluvoxamine, for controlling climacteric symptoms and vasomotor symptoms, in particular.     

A unique mechanism for cyclic adenosine 3',5'-monophosphate-induced increase of 32-kDa tyrosine-phosphorylated protein in boar spermatozoa

A cAMP-induced increase of tyrosine-phosphorylated proteins is involved in the expression of fertilizing ability in mammalian spermatozoa. We (Harayama, 2003: J Androl 24:831-842) reported that incubation of boar spermatozoa with a cell-permeable cAMP analog (cBiMPS) increased a 32-kDa tyrosine-phosphorylated protein (TyrP32). The purpose of this study is to characterize the signaling cascades that regulate the cAMP-induced increase of TyrP32. We examined effects of tyrosine kinase inhibitor (lavendustin A), tyrosine phosphatase inhibitor (Na3VO4), cell-permeable calcium chelator (BAPTA-AM), and cholesterol acceptor (methyl-beta-cyclodextrin: MBC) on the increase of TyrP32 and the change and loss of acrosomes in boar spermatozoa. The spermatozoa were used for detection of tyrosine-phosphorylated proteins by Western blotting and indirect immunofluorescence and for examination of acrosomal integrity by Giemsa staining. At least eight tyrosine-phosphorylated proteins including TyrP32 exhibited the cAMP-dependent increase during incubation with cBiMPS. In many proteins of them, this increase was reduced by lavendustin A but was enhanced by Na3VO4. In contrast, the cAMP-induced increase of TyrP32 was abolished by Na3VO4 but was hardly affected by lavendustin A. Giemsa staining showed that the increase of spermatozoa with weakly Giemsa-stained acrosomes (severely damaged acrosomes) or without acrosomes was correlative to the cAMP-induced increase of TyrP32. Moreover, the lack of calcium chloride in the incubation medium or pretreatment of spermatozoa with BAPTA-AM blocked the change and loss of acrosomes and the increase of TyrP32, suggesting these events are dependent on the extracellular and intracellular calcium. On the other hand, incubation of spermatozoa with MBC in the absence of cBiMPS could mimic the change and loss of acrosomes and increase of TyrP32 without increase of other tyrosine-phosphorylated proteins. Based on these results, we conclude that the cAMP-induced increase of TyrP32 is regulated by a unique mechanism that may be linked to the calcium-dependent change and loss of acrosomes.     

Disassembly of amyloid beta-protein fibril by basement membrane components

Amyloid beta-protein (A3) fibril in senile plaque may be related to the pathogenesis of Alzheimer's disease (AD). Basement membrane (BM) components are associated with the plaques in AD brain. It suggests that the BM components may play an important role in the deposition of the plaque. We investigated the potential of BM components, such as type IV collagen (collagen IV) and entactin, to induce disassembly of preformed Abeta1-42 (Abeta42) fibrils in direct comparison to laminin. Thioflavin T assays revealed that these BM components disrupted preformed Abeta42 fibrils in a dose-dependent manner. The high concentration of BM components, 100 microg/mL laminin, 50 microg/mL collagen IV and 50 microg/mL entactin, had most effect on disassembly of preformed Abeta42 fibrils (Molar ratio; Abeta42:laminin = 90:1, Abeta42:collagen IV = 34:1, Abeta42:entactin = 20:1). Circular dichroism spectroscopy data indicated that the high concentration of BM components induced structural transition in Abeta42 from beta-sheet to random structures. These results suggest that collagen IV and entactin, as well as laminin, are effective inducers of disassembly of Abeta42 fibrils. The ability of these BM components to induce random structures may be linked to the disassembly of preformed Abeta42 fibrils.     

Type IV collagen prevents amyloid beta-protein fibril formation

The potential of targeting through molecular therapeutics the underlying amyloid beta-protein (A beta) fibrillogenesis causing the initiation and progression of Alzheimer's disease (AD) offers an opportunity to improve the disease. Type IV collagen (collagen IV) is localized in senile plaques in patients with AD. By using thioflavin T fluorescence spectroscopy and electron microscopy, we found that collagen IV inhibited A beta1-40 (A beta40) fibril formation. The critical concentration of collagen IV for this inhibition was 5 microg/mL. Circular dichroism data indicate that collagen IV prevents formation of a beta-structured aggregate of A beta40. These studies demonstrated that collagen IV is apparently a potent inhibitor of A beta fibril formation.     

Susceptibility gene for non-obstructive azoospermia located near HLA-DR and -DQ loci in the HLA class II region

The technical developments and expanded indications for testicular sperm extraction (TESE) with intracytoplasmic sperm injection (ICSI) provide great advantages for patients with non-obstructive azoospermia. Such success, however, also means that genetic abnormalities in non-obstructive azoospermia can be transmitted to the next generation, demonstrating the importance of being able to understand the genetic background of non-obstructive azoospermia. We have previously reported that human leukocyte antigens (HLA)-A33 and -B44 in the HLA class I region and the HLA-DRB1*1302 allele in the HLA class II region are linked to susceptibility to non-obstructive azoospermia in Japanese men. However, strong linkage of HLA-DRB1*1302 with HLA-A33 and -B44 is also evident in the Japanese population. Thus, uncertainty prevails as to whether the HLA class I or class II molecule is more directly associated with non-obstructive azoospermia. In the present study, we performed association analysis with 21 polymorphic microsatellite markers identified near the HLA genes to map the gene involved in the development of non-obstructive azoospermia more precisely. Microsatellite markers located in the HLA class I region or the class III region showed no statistically significant association with this disorder, although once again the HLA-A33 and -B44 alleles showed a significant association. In contrast, some of the microsatellite markers in the HLA class II region and at the HLA-DRB1 and -DQB1 loci displayed strong associations with non-obstructive azoospermia. Taken together, our previous and present data suggest that the critical region for development of non-obstructive azoospermia is near the HLA-DRB1 and -DQB1 segments in the HLA class II region.     

Target site specificity of the Tos17 retrotransposon shows a preference for insertion within genes and against insertion in retrotransposon-rich regions of the genome

Because retrotransposons are the major component of plant genomes, analysis of the target site selection of retrotransposons is important for understanding the structure and evolution of plant genomes. Here, we examined the target site specificity of the rice retrotransposon Tos17, which can be activated by tissue culture. We have produced 47,196 Tos17-induced insertion mutants of rice. This mutant population carries approximately 500,000 insertions. We analyzed >42,000 flanking sequences of newly transposed Tos17 copies from 4316 mutant lines. More than 20,000 unique loci were assigned on the rice genomic sequence. Analysis of these sequences showed that insertion events are three times more frequent in genic regions than in intergenic regions. Consistent with this result, Tos17 was shown to prefer gene-dense regions over centromeric heterochromatin regions. Analysis of insertion target sequences revealed a palindromic consensus sequence, ANGTT-TSD-AACNT, flanking the 5-bp target site duplication. Although insertion targets are distributed throughout the chromosomes, they tend to cluster, and 76% of the clusters are located in genic regions. The mechanisms of target site selection by Tos17, the utility of the mutant lines, and the knockout gene database are discussed. --The nucleotide sequence data were uploaded to the DDBJ, EMBL, and GenBank nucleotide sequence databases under accession numbers AG020727 to AG025611 and AG205093 to AG215049.     

Properties of Chitosanase from Bacillus cereus S1

Chitosanase from Bacillus cereus S1 was purified, and the enzymatic properties were investigated. The molecular weight was estimated to 45,000 on SDS-PAGE. Optimum pH was about 6, and stable pH in the incubation at 40°C for 60 min was 6–11. This chitosanase was stable in alkaline side. Optimum temperature was around 60°C, and enzyme activity was relatively stable below 60°C. The degradations of colloidal chitosan and carboxymethyl cellulose (CMC) were about 30 and 20% relative to the value of soluble chitosan, respectively, but colloidal chitin and crystalline cellulose were not almost hydrolyzed. On the other hand, S1 chitosanase adsorbed on colloidal chitin completely and by about 50% also on crystalline cellulose, in contrast to colloidal chitosan, which it did not adsorb. S1 chitosanase finally hydrolyzed 100% N-deacetylated chitosan (soluble state) to chitobiose (27.2%), chitotriose (40.6%), and chitotetraose (32.2%). In the hydrolysis of various chitooligosaccharides, chitobiose and chitotriose were not hydrolyzed, and chitotetraose was hydrolyzed to chitobiose. Chitobiose and chitotriose were released from chitopentaose and chitohexaose. From this specificity, it was hypothesized that the active site of S1 chitosanase recognized more than two glucosamine residues posited in both sides against splitting point for glucosamine polymer. Received: 8 June 1999 / Accepted: 20 July 1999     

Effects of TGF-β1 on the proliferation and differentiation of human periodontal ligament cells and a human periodontal ligament stem/progenitor cell line

Periodontal ligament (PDL) is a specialized connective tissue that influences the lifespan of the tooth. Transforming growth factor-β1 (TGF-β1) is a multifunctional cytokine, but little is known about the effects of TGF-β1 on PDL cells. Our aim has been to demonstrate the expression of TGF-β1 in rat PDL tissues and to evaluate its effects on the proliferation and gene expression in human PDL cells (HPLCs) and a human PDL stem/progenitor cell line, line 1-11, that we have recently developed. The expression of TGF-β1 in the entire PDL tissue was confirmed immunohistochemically, and both HPLCs and cell line 1-11 expressed mRNA from the TGF-β1, TGF-β type I receptor, and TGF-β type II receptor genes. Although exogenous TGF-β1 stimulated the proliferation of HPLCs, it did not upregulate the expression of alpha-smooth muscle actin (α-SMA), type I collagen (Col I), or fibrillin-1 (FBN1) mRNA or of α-SMA protein in HPLCs, whereas expression for these genes was attenuated by an anti-TGF-β1 neutralizing antibody. In contrast, exogenous TGF-β1 reduced the proliferation of cell line 1-11, although it upregulated the expression of α-SMA, Col I, and FBN1 mRNA and of α-SMA protein in this cell line. In addition, interleukin-1 beta stimulation significantly reduced the expression of TGF-β1 mRNA and protein in HPLCs. Thus, TGF-β1 seems to play an important role in inducing fibroblastic differentiation of PDL stem/progenitor cells and in maintaining the PDL apparatus under physiological conditions.