Search for a Variation of the Fine Structure Constant around the Supermassive Black Hole in Our Galactic Center.

Searching for space-time variations of the constants of Nature is a promising way to search for new physics beyond general relativity and the standard model motivated by unification theories and models of dark matter and dark energy. We propose a new way to search for a variation of the fine-structure constant using measurements of late-type evolved giant stars from the S star cluster orbiting the supermassive black hole in our Galactic Center. A measurement of the difference between distinct absorption lines (with different sensitivity to the fine structure constant) from a star leads to a direct estimate of a variation of the fine structure constant between the star's location and Earth. Using spectroscopic measurements of five stars, we obtain a constraint on the relative variation of the fine structure constant below 10^{-5}. This is the first time a varying constant of nature is searched for around a black hole and in a high gravitational potential. This analysis shows new ways the monitoring of stars in the Galactic Center can be used to probe fundamental physics.

Topical application of glycyrrhetinic acid in the gingival sulcus inhibits attachment loss in lipopolysaccharide-induced experimental periodontitis in rats.

BACKGROUND AND OBJECTIVE: Attachment loss of the junctional epithelium and alveolar bone destruction are signs of periodontitis, which is mainly caused by an inflammatory response to dental plaque. Glycyrrhetinic acid (GA), a component of the licorice herb, has been shown to have important anti-inflammatory activities; however, there are no previous reports on the ability of its inhibitory effects to prevent periodontal diseases. Hence, in this study, using our experimental periodontitis model, we attempted to evaluate whether GA had an effect on the prevention of attachment loss and alveolar bone loss. MATERIAL AND METHODS: Rats were intraperitoneally immunized with Escherichia coli lipopolysaccharide (LPS). The LPS group (n = 5) received 3 topical applications of 50 µg/µL of LPS followed by one application of the vehicle (propylene glycol:ethyl alcohol:phosphate-buffered saline [PBS] = 8:1:1) into the gingival sulcus. This protocol was repeated twice per day for 10 days. The low (n = 5) and high (n = 5) groups received topical application of LPS and 0.03% or 0.3% GA, respectively. The control group received topical application of PBS and vehicle. The rats were killed on the 10th day. Attachment loss, alveolar bone level and inflammatory cell infiltration were investigated histometrically. The formation of immune complexes and infiltration of LPS were evaluated immunohistologically. RESULTS: Attachment loss, formation of immune complexes and infiltration of inflammatory cells were increased in the LPS group compared with the control group, and were completely inhibited in the low and high groups compared with the LPS group. The LPS group showed greater alveolar bone destruction compared with the control group and GA-treated groups. In addition, invasion of LPS was detected in the LPS group, was absent in the control group and was weaker in the GA-treated groups than in the LPS group. CONCLUSION: In the present study, we showed that GA inhibits periodontal destruction in the rat experimental periodontitis model.

The histopathological comparison on the destruction of the periodontal tissue between normal junctional epithelium and long junctional epithelium.

BACKGROUND AND OBJECTIVE: The barrier function of long junctional epithelium is thought to be important after periodontal initial therapy and periodontal surgery. Although the difference between long junctional epithelium and normal junctional epithelium regarding their resistance to destruction of periodontal tissue has been investigated, the mechanism still remains unclear. Using our rat experimental periodontitis model in which loss of attachment and resorption of alveolar bone is induced by the formation of immune complexes, we investigated the resistance of periodontal tissue containing long junctional epithelium and normal junctional epithelium to destruction. MATERIAL AND METHODS: Rats were divided into four groups. In the immunized long junctional epithelium (I-LJE) group, rats were immunized with lipopolysaccharide (LPS), and curettage and root planing procedures were performed on the palatal gingiva of the maxillary first molars to obtain reattachment by long junctional epithelium. In the immunized normal junctional epithelium (I-JE) group, rats were immunized without curettage and root planing procedures. In the nonimmunized long junctional epithelium (nI-LJE) group, rats were not immunized but curettage and root-planing procedures were performed. In the control group, neither immunization nor curettage and root-planing was performed. In all rats, periodontal inflammation was induced by topical application of LPS into the palatal gingival sulcus of maxillary first molars. The rats were killed at baseline and after the third and fifth applications of LPS. Attachment loss and the number of inflammatory cells and osteoclasts in the four groups were compared histopathologically and histometrically. RESULTS: After the third application of LPS in the I-LJE group, attachment loss showed a greater increase than in control and nI-LJE groups, and inflammatory cell infiltration and osteoclasts were increased more than in the other groups. After the fifth application of LPS, attachment loss was greater and there was a higher degree of inflammatory cell infiltration in nI-LJE and I-LJE groups than in control and I-JE groups. CONCLUSION: Our findings suggest that the destruction of periodontal tissue is increased in tissue containing long junctional epithelium compared with normal junctional epithelium and that the immunized condition accelerates the destruction by forming immune complexes.

Intracellular translocation of glutathione S-transferase pi during oligodendrocyte differentiation in adult rat cerebral cortex in vivo.

Glutathione S-transferase (GST)-pi is a cytosolic isoenzyme used as a marker for mature oligodendrocytes in the mammalian brain. However, the cellular properties of GST-pi-immunoreactive [GST-pi (+)] cells in adult brain are not completely understood. We immunohistochemically demonstrated the existence of two subtypes of GST-pi (+) cells in the cerebral cortex of adult rats: one subtype exhibited GST-pi in the cytoplasm (C-type cells), while the other did mainly in the nucleus (N-type cells). The GST-pi (+) C-type cells were also immunopositive for 2',3'-cyclic nucleotide 3'-phosphodiesterase and RIP, indicating that they were mature oligodendrocytes, while the GST-pi (+) N-type cells expressed NG2, indicating that they were oligodendrocyte progenitor cells. Furthermore, observation of the fate of newly-generated cells by 5-bromodeoxyuridine-labeling revealed that the GST-pi (+) N-type cells differentiated into C-type cells. These findings indicate translocation of GST-pi from the nucleus to the cytoplasm during oligodendrocyte maturation.

Propofol acts at the sigma-1 receptor and inhibits pentazocine-induced c-Fos expression in the mouse posterior cingulate and retrosplenial cortices.

BACKGROUND: The sigma-1 receptor is functionally linked with psychotomimetic effects of various drugs. A sigma-1 receptor agonist enhances bradykinin-induced intracellular Ca(2+) concentration ([Ca(2+)]i) increase and induces c-Fos expression in a part of the brain. The aim of this study was to investigate the effects of several intravenous anaesthetics on the sigma-1 receptor. METHODS: First, using Wistar rat brains, (+)[(3)H]SKF-10,047, a selective sigma-1 receptor agonist was displaced by propofol, dexmedetomidine, droperidol, and thiopental. Second, Fura-2 loaded NG-108 cells were incubated with (+)pentazocine, a selective sigma-1 receptor agonist, and propofol and then its fluorescence was observed after stimulation with bradykinin. Third, male ICR mice received Intrafat or propofol intraperitoneally (i.p.), followed by pentazocine i.p. Brain slices were prepared and Fos-like immunoreactivity was detected using an immunohistochemical method. results: Propofol, droperidol, and dexmedetomidine displaced (+)[(3)H]SKF-10,047 binding in a concentration-dependent manner with Ki50s of 10.2 +/- 0.6, 0.17 +/- 0.03, 5.73 +/- 1.2 microM, respectively. Thiopental sodium was practically ineffective. Propofol produced a statistically significant reduction in the maximal binding capacity (Bmax) but did not affect the dissociation constant (K(d)). (+)Pentazocine significantly enhanced bradykinin-induced [Ca(2+)]i increases, but propofol did not affect it. Pentazocine induced marked Fos-LI positive cells in the posterior cingulate and retrosplenial cortices (PC/RS), which was significantly reduced by propofol. CONCLUSIONS: These results suggest that propofol may be a sigma-1 receptor antagonist, and that various effects of propofol on the brain may be mediated, at least partly, by the sigma-1 receptor.

Expression patterns of the erbB subfamily mRNA in canine benign and malignant mammary tumors.

ErbB subfamily genes, known as proto-oncogenes, encode receptor tyrosine kinases, and are expressed in relation to tumorigenesis of the mammary gland in humans. In this study, we examined the expression of erbB subfamily mRNAs in two canine normal mammary glands and 12 mammary tumor samples by reverse transcriptase-coupled polymerase chain reaction (RT-PCR). Each primer set was designed from the nucleotide sequence of the region conserved in erbB subfamily cDNA among other species. No erbB subfamily mRNAs were expressed in the normal mammary gland. In contrast, all of the subfamily mRNAs were expressed in a benign mammary tumor, and more than one type of the subfamily mRNA were observed in 11 malignant mammary tumors. The length of RT-PCR products were 380 bp for erbB1, 500 bp for erbB2, 644 bp for erbB3, and 416 bp for erbB4. These sequences were highly homologous to the cDNA sequences of other species. Therefore, these results suggest that the expression of erbB subfamily mRNAs in canine mammary tumors plays an important role in tumorigenesis of the mammary gland.

Effect of bile acids on biliary excretion of cyclosporin A in the rat.

Cyclosporin A, a substrate of P-glycoprotein (P-gp), is known to cause cholestasis in humans and in rat experimental models. Tauroursodeoxycholate is reported to be effective in CyA-induced cholestasis in rats. In the present study, to investigate the mechanism of the inhibition of CyA induced cholestasis, effect of bile acids on biliary cyclosporin A excretion was studied in rats. Infusion of both taurocholate and tauroursodeoxycholate at the rate of 0.8 mmol/min per 100 g bodyweight increased bile flow and biliary cyclosporin A excretion, and the extent was more prominent with tauroursodeoxycholate. It was suggested that these findings were caused by the enhanced vesicular targeting of P-gp to the canalicular membrane by bile acids, thus increasing the numbers of P-gp in the canalicular membrane.

Prostaglandin D2 selectively induces chemotaxis in T helper type 2 cells, eosinophils, and basophils via seven-transmembrane receptor CRTH2.

Prostaglandin (PG)D2, which has long been implicated in allergic diseases, is currently considered to elicit its biological actions through the DP receptor (DP). Involvement of DP in the formation of allergic asthma was recently demonstrated with DP-deficient mice. However, proinflammatory functions of PGD2 cannot be explained by DP alone. We show here that a seven-transmembrane receptor, CRTH2, which is preferentially expressed in T helper type 2 (Th2) cells, eosinophils, and basophils in humans, serves as the novel receptor for PGD2. In response to PGD2, CRTH2 induces intracellular Ca2+mobilization and chemotaxis in Th2 cells in a Galphai-dependent manner. In addition, CRTH2, but not DP, mediates PGD2-dependent cell migration of blood eosinophils and basophils. Thus, PGD2 is likely involved in multiple aspects of allergic inflammation through its dual receptor systems, DP and CRTH2.

Presence of B220 within thymocytes and its expression on the cell surface during apoptosis.

B220 is the full-length splicing isoform of a tyrosine phosphatase CD45 and is predominantly expressed as a transmembrane protein on B cells. Other splicing isoforms of CD45 are yielded by alternative splicing of exons 4, 5 and 6. Recently, the expression of B220 on peripheral T cells during activation-induced cell death has been reported. To investigate whether B220 is implicated in apoptosis of immature T cells, we analysed (by flow cytometry using the anti-B220 monoclonal antibody, RA3-6B2) the expression of B220 on mouse thymocytes undergoing X-irradiation- and dexamethasone (DEX)-induced apoptosis. The expression of B220 on thymocytes positive for Thy-1 was induced by X-irradiation or DEX treatment and increased with length of incubation. The expression of B220 was pronounced on the apoptotic hypodiploid cells in the fraction showing lower forward scattering values. Reverse transcription-polymerase chain reaction detected mRNA containing exons 4, 5 and 6 of CD45 in normal thymocytes as well as those exposed to X-rays or DEX. Surprisingly, cytoplasmic B220 antigens were detected in a considerable fraction of normal thymocytes. Moreover, the expression level of the 220 000-MW protein in normal thymocytes was similar to that in the thymocytes undergoing apoptosis. During apoptosis, the expression level of B220 antigen was reduced in the cytoplasm but, conversely, up-regulated on the surface of thymocytes. These results suggest that B220 is constitutively expressed as a cytoplasmic form within thymocytes and possibly translocated to the cell membrane during apoptosis.

Effect of phenobarbital on intralobular expression of CYP2B1/2 in livers of rats: difference in the expression between single and repetitive administrations.

Phenobarbital (PB) was shown to induce the major PB-inducible cytochrome P450 (CYP) isoforms, CYP2B1/2, in perivenular hepatocytes by a single injection, and in midzonal and periportal hepatocytes in addition to perivenular hepatocytes by injections of the same dosage once a day for 3 days in rat livers. The present study was undertaken to determine whether the spread of enzyme induction to midzonal and periportal hepatocytes is caused by the increase in total dose of the drug by repetitive injections or by the repetitive injections of the drug themselves. Male adult rats were administered PB by a single injection (80 mg/kg) or repetitive injections (20 mg/kg once a day for 4 days; a total dose of 80 mg/kg), and the molar content of CYP2B1/2 was measured by quantitative immunohistochemistry in the cytoplasm of perivenular, midzonal, and periportal hepatocytes. In addition, the molar content of total CYP in the cytoplasm was measured by microphotometry, and the expression of CYP2B2 mRNA was examined by in situ hybridization. When animals received the single injection, the isoforms and CYP2B2 mRNA increased markedly in perivenular hepatocytes, increased somewhat in midzonal hepatocytes, and remained unchanged in periportal hepatocytes. If animals received the repetitive injections, however, although the isoforms and the mRNA increased markedly in perivenular hepatocytes, they also increased markedly in midzonal hepatocytes and somewhat in periportal hepatocytes. These findings demonstrated that the enlargement of the sublobular area in which induction of the isoforms occurred was caused by the repetitive injections of PB themselves.

Substrate specificity of human methylpurine DNA N-glycosylase.

The activity of human methylpurine DNA N-glycosylase (hMPG) for major substrates was directly compared using two types of substrates, i.e., natural DNA and synthetic oligonucleotides. By the use of ARP assay detecting abasic sites in DNA, we first investigated the activity on the natural DNA substrates containing methylpurines, ethenopurines, or hypoxanthine (Hx) prepared by the conventional methods. After the treatment with hMPG, the amount of AP sites in methylated DNA was much higher than that in DNA containing ethenopurines or Hx. The oligodeoxynucleotide having a single 7-methylguanine (7-mG) was newly synthesized in addition to 1, N(6)-ethenoadenine (epsilonA)-, Hx-, and 8-oxoguanine-containing oligonucleotides. 7-mG was effectively excised by hMPG, though it might be less toxic than the other methylated bases with respect to mutagenesis and cell killing. The kinetic study demonstrated that k(cat)/K(m) ratios of the enzyme for epsilonA, Hx, and 7-mG were 2.5 x 10(-3), 1.4 x 10(-3), and 4 x 10(-4) min(-1) nM(-1), respectively. The oligonucleotides containing epsilonA effectively competed against 7-mG, while Hx substrates showed unexpectedly low competition. Concerning the effect of the base opposite damage, hMPG much preferred Hx.T to other Hx pairs, and epsilonA.C and epsilonA.A pairs were better substrates than epsilonA.T.

Flow cytometric analysis of thymocyte subpopulations in mice after whole-body X-irradiation.

To determined the cellular kinetics of thymocyte subpopulations in DBA1 mice after whole-body 6.8 Gy X-irradiation, they were analyzed for the expression of several cell surface antigens using flow cytometry. The results show that i) The majority of thymocytes rapidly depleted by irradiation was CD4+8+ cells. ii) radioresistant CD4+8- and CD4-8+ survived 18-48 hr after X-irradiation were considered to be relatively mature type, since they expressed high levels of CD3 and LECAM-1. iii) CD3-positive cells were detected in CD4-8- cells at 72 hr after irradiation.

B220 is expressed on apoptotic thymocytes induced by X-irradiation.

CD45 is cell surface glycoprotein and expressed on all haematopoietic cells except mature erythrocytes and platelets. Eight isoforms of CD45 are generated by alternative splicing of exons 4-6. B220 including all three exons is expressed specifically on pan-B cell lineage. Recently, it was reported that B220 was expressed on apoptotic T cells induced by staphylococcal enterotoxin B (SEB). In the present study, we investigated the expression of B220 on murine thymocytes after whole-body X-irradiation. We used the forward light scattering of flow cytometry as a parameter of cell size, and defined two populations; FSChigh (normal cell size) and FSClow (correspond to apoptotic cell in size) fraction. B220+ cells in FSChigh fraction reached a maximum value (35%) at 18 hr after irradiation. In FSClow fraction, 40-60% cells were positive for B220 at any time points. These results suggest that B220 is expressed on thymocytes in the pre-apoptotic stage, because B220 was expressed on not only FSClow cells but also FSChigh cells.

Increased LH pulse frequency and estrogen secretion associated with termination of anestrus followed by enhancement of uterine estrogen receptor gene expression in the beagle bitch.

The relationships among pulsatile LH secretion pattern, estrogen secretion, and expression of the uterine estrogen receptor gene were examined throughout the estrous cycle in beagle bitches. In Experiment 1, blood samples were collected from 30 bitches every 10 min for 8 h from a cephalic vein during different phases of the estrous cycle. An increase in the mean plasma levels of LH occurred from mid to late anestrus (P < 0.01). The LH pulse frequency increased (P < 0.01) from late anestrus to proestrus, and was strongly correlated (r = 0.96, P < 0.001) with the mean plasma level of estradiol-17 beta (E2). In Experiment 2, middle uterine samples, including the myometrium and endometrium, from 18 bitches were taken at 6 stages of the estrous cycle. The total number of estrogen receptors and nuclear estrogen receptor and its mRNA levels in the uterus also increased (P < 0.01) from late anestrus to proestrus. Mean plasma E2 level and the number of uterine estrogen receptor were positively correlated (r = 0.81, P < 0.05). In Experiment 3, nine bitches were ovariectomized in mid anestrus. Two weeks later they received a single injection of 10 or 50 micrograms/kg, i.m., estradiol benzoate. The number of uterine estrogen receptor and their mRNA levels for ovariectomized bitches were low, but increased (P < 0.05) after treatment with a low dose of estradiol benzoate. These results suggest that increases in LH pulse frequency and estrogen secretion are associated with termination of anestrus and that subsequent enhancement of uterine estrogen receptor expression may be up-regulated by estradiol.

Putative tumor suppressor gene region within 0.85 cM on chromosome 12 in radiation-induced murine lymphomas.

Analyses of genetic alterations in tumors from F1 hybrid mice produced by inter-subspecific crosses between genetically well-characterized inbred strains provide precise and comprehensive evidence for genetic abnormalities such as allelic loss. We performed loss of heterozygosity (LOH) analyses of 125 radiation-induced lymphomas of (BALB/cHeA x MSM/Ms)F1 hybrid mice by polymerase chain reaction (PCR) analysis of microsatellite DNA polymorphic markers. Very frequent LOH was found at a distal region on chromosome 12. To precisely define the most common region of LOH, we first determined the order of and distances between the available microsatellite loci around the region by using 586 (CXSD x MSM/Ms)F2 hybrid mice (1172 meiosis). The locus order and distances were [centromere]-D12Mit132-(0.34 cM)-D12Mit5O-(2.05 cM)-[D12Mit122, D12Mit53]-(0.85 cM)-D12Mit233-(0.43 cM)-D12Mit279-(O.17 cM)-D12Mit181-[telomere]. We then investigated the features of LOH at these loci. The highest frequency of LOH (83 of 125, 66%) was found at D12Mit233. The LOH patterns of individual lymphomas indicated that the most common region of LOH was within the 0.85 cM between D12Mit53 and D12Mit233, a region homologous to human chromosome 14q32.1. These results suggest that a putative novel tumor suppressor gene exists within this region.

Repair kinetics of abasic sites in mammalian cells selectively monitored by the aldehyde reactive probe (ARP).

Human methylpurine N-glycosylase (MPG) activity was investigated by monitoring abasic (AP) sites resulting from removal of alkylated bases. The amount of AP sites in MMS-treated HeLa cells transiently increased at 3 h, then gradually decreased to 40% at 24 h. The presence of adenine, an inhibitor of AP endonucleases, in the repair incubation of MMS-treated cells induced moderate accumulation of AP sites, suggesting inhibition of the activities of MPG as well as AP endonucleases by adenine metabolites.

Novel putative fragile sites observed in feline fibroblasts treated with aphidicolin and fluorodeoxyuridine.

Fragile sites are non-randomly distributed chromosomal breaks and gaps observed in the cells cultivated under certain conditions. Feline fragile sites were analyzed using skin fibroblast strains after the treatments with aphidicolin and fluorodeoxyuridine in combination with caffeine. Three aphidicolin-induced fragile sites (A1q21, C2q13 and E1p21) as well as a folate-sensitive site (B1q14) were observed in all the 3 fibroblast strains tested for each treatment group. The loci in A1q21 and B1q14 are very close to that reported previously for peripheral blood lymphocytes and lung cells. Two chromosomal break points in C2q13 and E1p21 seem to be new fragile sites. Fifteen candidates for feline fragile sites were also assigned their locations in feline chromosomes. Both the incidence and distribution of feline fragile sites in skin fibroblasts seem to be different at least in part from those in lymphocytes.

Induction of fragile sites by fluorodeoxyuridine and caffeine accompanies with misincorpolation of endogenous uridine nucleotide into DNA of feline fibroblasts.

Fluorodeoxyuridine, an inhibitor of thymidylate synthetase, is known to induce chromosomal fragile sites. The drug treatment may cause deprivation of intracellular thymidine nucleotide pool followed by a serious imbalance of deoxynucleotide pool. Though the stress is probably related to the induction of folate-sensitive fragile sites, the exact mechanism is still to be investigated. The present study has been carried out to test the possibility that the fragile sites are originated, at least in part, from incorpolated uracil residues. The incorpolated uracil residue can be detected by a novel assay for abasic sites after treatment with uracil-DNA N-glycosylase (UDG). About 2.7 abasic sites per 10(4) nucleotides were detected in the DNA extracted from feline fibroblasts after the treatment with FUdR and caffeine. By digesting the DNA with UDG prior to the assay, significant increase in the number of abasic sites were observed. These results indicate that the large amount of uracil residues are present in the feline fibroblast DNA under the condition which induces chromosomal fragile sites.

Frequent expression of the c-kit proto-oncogene in canine malignant mammary tumor.

The mammary tumor is one of the popular neoplastic diseases in female dogs. In the present study, the expression of canine c-kit proto-oncogene in mammary tumor specimens was investigated to evaluate its potential usefulness as a tumor marker. By comparing the homology among the nucleotide sequences reported for human mouse, rat and feline c-kit c-DNA, a pair of primers was synthesized for the reverse transcriptase-polymerase chain reaction (RT-PCR) method. The RT-PCR product of canine spleen total RNA was shown to have 756 bp in size and to be highly homologous to the corresponding sequences reported for the mammalian species. The expression of c-kit transcript was detected in 11 mammary tumors of different histopathology including adenocarcinomas, benign and malignant mixed tumors. The level of the transcription in adenocarcinomas was significantly higher than those in malignant mixed tumors. Fifteen canine tumor specimens originated from various tissues were also tested for their c-kit transcript. In all of the mastocytoma samples examined, high expression of the mRNA was detected. Of other 12 tumors, only low level of RT-PCR products were detected in 5 samples, whereas no apparent amplification was observed in 7 tumors. These results indicate that the high expression of c-kit transcript is helpful for the diagnosis of canine mammary tumors.

Changes in biliary excretory mechanisms in rats with ethinyloestradiol-induced cholestasis.

Several excretory pathways for cholephilic compounds have been known. To examine the changes in excretory pathways in cholestasis induced by ethinyloestradiol, various bile acids, organic anions and organic cations were intravenously administered to ethinyloestradiol-treated rats and their biliary excretion was studied. Biliary excretion of taurocholate was slightly delayed, but its excretory maximum was markedly decreased. Biliary excretion of lithocholate-3-O-glucuronide, leukotriene C4, sulphobromophthalein and pravastatin was markedly impaired to a similar extent. Biliary excretion of vinblastine, a P-glycoprotein substrate, was increased, suggesting increased expression of P-glycoprotein. In contrast, biliary excretion of erythromycin, a cationic antibiotic, was markedly impaired. In conclusion, ethinyloestradiol treatment altered the biliary excretion of organic compounds, which may partly be related to changes of the canalicular transporters.