Density-dependent distribution of parasitism risk among underground hosts.

Variation in parasitism risk among hosts can arise from between-patch and within-patch factors, but considerably less information is known about the latter. This study investigated how distributions of the oriental fruit fly Bactrocera dorsalis influenced its parasitism by the pupal parasitoid Dirhinus giffardii in the laboratory. Because B. dorsalis larvae pupate underground, pupation depth was considered as an important factor that affects the risk of parasitism. When the density of B. dorsalis larvae was varied (1, 10, and 100 larvae per arena), average pupation depth increased with the density. When the depth of pupae was manipulated, the rate of parasitism differed by depths. Parasitism at 0 cm differed from the random parasitoid model expectation, but parasitism at 1 cm was not different from the model expectation. Few pupae at 2 cm were parasitized. In another experiment, when pupae were simultaneously presented at 0 cm and 1 cm depths, parasitism at 1 cm was weakened by the presence of puape at 0 cm. These results imply that the density of the host influences pupation depth as well as the distribution of parasitism and plays an important role in host-parasitoid dynamics.

Verification of the influence of the arrangement of implants on the load distribution (a well-known figure by Rangert).

The purpose of this study is to evaluate the effects of the number and the placement of implants on load distribution for multiple implants with three-dimensional geometric analysis, and to verify the well-known conceptual figure by Rangert. Three teeth missing in left mandibular region was geometrically modelled in clinically simulated situation. Two implants placement as 'control', 'cantilever', 'three-implants' and 'offset placements' were analyzed with geometric analysis. The cantilever received 180-182% load of control, that is, almost same to the result by Rangert (200%). Three implants received 59-65% load of control, that is, almost same to the result by Rangert (67%). Offset arrangement received 59-65% load of control, that is, larger than the result by Rangert (40%). It was concluded that the influence of the arrangement of implants on the load distribution presented in the conceptual figure by (Rangert BR, Sullivan RM & Jemt TM, J Oral Maxillofac Implants. 1997;12:360) was verified except for the effects of the offset arrangement.

Psychotherapy for depression among incurable cancer patients.

BACKGROUND: The most common psychiatric diagnosis among cancer patients is depression; this diagnosis is even more common among patients with advanced cancer. Psychotherapy is a patient-preferred and promising strategy for treating depression among cancer patients. Several systematic reviews have investigated the effectiveness of psychological treatment for depression among cancer patients. However, the findings are conflicting, and no review has focused on depression among patients with incurable cancer. OBJECTIVES: To investigate the effects of psychotherapy for treating depression among patients with advanced cancer by conducting a systematic review of randomized controlled trials (RCTs). SEARCH STRATEGY: We searched the Cochrane Pain, Palliative and Supportive Care Group Register, The Cochrane Controlled Trials Register, MEDLINE, EMBASE, CINAHL, and PsycINFO databases in September 2005. SELECTION CRITERIA: All relevant RCTs comparing any kind of psychotherapy with conventional treatment for adult patients with advanced cancer were eligible for inclusion. Two independent review authors identified relevant studies. DATA COLLECTION AND ANALYSIS: Two review authors independently extracted data from the original reports using standardized data extraction forms. Two independent review authors also assessed the methodological quality of the selected studies according to the recommendations of a previous systematic review of psychological therapies for cancer patients that utilized ten internal validity indicators. The primary outcome was the standardized mean difference (SMD) of change between the baseline and immediate post-treatment scores. MAIN RESULTS: We identified a total of ten RCTs (total of 780 participants); data from six studies were used for meta-analyses (292 patients in the psychotherapy arm and 225 patients in the control arm). Among these six studies, four studies used supportive psychotherapy, one adopted cognitive behavioural therapy, and one adopted problem-solving therapy. When compared with treatment as usual, psychotherapy was associated with a significant decrease in depression score (SMD = -0.44, 95% confidence interval [CI] = -0.08 to -0.80). None of the studies focused on patients with clinically diagnosed depression. AUTHORS' CONCLUSIONS: Evidence from RCTs of moderate quality suggest that psychotherapy is useful for treating depressive states in advanced cancer patients. However, no evidence supports the effectiveness of psychotherapy for patients with clinically diagnosed depression.

Reinvestigation of extremely acidic proteins in bovine brain.

Analysis of bovine brain extract by disc electrophoresis on a 20% polyacrylamide gel indicated the existence of three extremely acidic proteins. These proteins were isolated by column chromatography on DEAE-Sephadex A-50 and Sephadex G-75. The isolated proteins (PAP I-a, PAP I-b and PAP II) were homogeneous in various methods including 7.5% and 20% gel electrophoresis or gel chromatography, and share, in the extract, 85% of the total of the acidic proteins that migrate with the bromophenol blue marker in 7.5% gels. Their physicochemical properties, including molecular weight, ultraviolet absorption spectra or amino acid composition were similar, especially those between PAP I-a and PAP I-b where a part of primary structure appeared to be common in their tryptic peptide maps. These two proteins were identified to be the nervous system specific protein S 100 by immunochemical and electrophoresis methods as well as by amino acid analysis, and the other protein PAP II was revealed to be a calcium-binding protein. The existence and properties of the isolated proteins are discussed with relation to the heterogeneity problem of S 100 protein.

Specific purification of glyceraldehyde-3-phosphate dehydrogenase by hydrophobic chromatography on immobilized colchicine.

Hydrophobic column chromatography of bovine brain extracts (40-80% ammonium sulfate fraction) on immobilized colchicine resulted in the selective elution of one major protein with decreasing ionic strength of medium. This protein was identified as glyceraldehyde-3-phosphate dehydrogenase (GAPDH; EC 1.2.1.1) on the basis of its biochemical properties, N-terminal amino-acid sequence and enzymatic activity. The present method enabled GAPDH to be isolated with a high recovery (80%; 184 mg/kg brain) and could be of potential use for the purification of GAPDH from various tissues.

Combined action of paraquat and superoxide on the peroxidation of detergent-dispersed linolenic acid.

We investigated the peroxidative effect of paraquat and active oxygens on detergent-dispersed linolenic acid in phosphate buffer (pH 7.5) from the malondialdehyde (MDA) level. Our complete system and further inclusion of catalase were effective in stimulating MDA formation. On the other hand, xanthine oxidase (XOD) or paraquat omission, superoxide dismutase (SOD) inclusion or anaerobic incubation inhibited the formation of MDA. Ferrous ion was weakly associated with phosphate of the buffer, forming a complex, and the release of ferrous ion from the complex intensified the MDA levels with the complete and catalase inclusion systems. The electron paramagnetic resonance (EPR) spectra using 5,5-dimethyl-1-pyrroline N-oxide (DMPO) showed that superoxide, produced immediately after the addition of XOD, played a crucial role. We could obtain a DMPO-OOH signal at the starting stage whenever MDA stimulation was observed. The omission of paraquat, however, produced no increase in MDA level in spite of an appearance of DMPO-OOH signal, indicating that paraquat also plays an important role. On the other hand, Desferal, a ferric chelator, showed a concentration-dependent inhibition effect. There was an immediate strong intensity of DMPO-OOH and paraquat signals. We did not, however, observe MDA stimulation at 250 microM Desferal, which confirms that ferrous ion plays an essential role in the lipid peroxidation. These results indicate a combined action of paraquat (or its radical) and superoxide on the accessibility of ferrous ion, including its release from the complex with phosphate, which may be an endogenous chelator. The possibility of ternary complex participation is also discussed.

A new acidic protein in porcine brain.

An extremely acidic protein has been isolated in a purified form from porcine rain extract, by (NH4)2SO4 fractionation followed by column chromatography on DEAE-Sephadex A-50 and on Sephadex G-75. The purified protein was tentatively named as glutamic acid-rich protein because it was characterized by its remarkably high content of glutamic acid which accounted for 49% of the total amino acid composition. The protein appeared to be a single polypeptide chain with a molecular weight of 56 000-58 000, and had an isoelectric point of 4.6. The N-terminal amino acid sequence was Asp-Glu-Pro-Pro-Ser-Glu-Gly. The immunochemical analysis using rabbit antiserum prepared to the porcine protein has suggested that it is present in the brain of human, cow, cat, dog and goat as well as in various goat organs including liver, kidney, heart, small intestine and spleen.

Developmental changes in the translatable mRNA for beta subunit of S-100 protein in rat brain.

The presence of mRNA coding for beta subunit of S-100 protein was demonstrated in polyadenylated RNA from the rat brain in vitro translation in a reticulocyte lysate cell-free system. The products were identified with S-100 protein beta subunit using the immunoprecipitation of the reaction products with the specific antisera, comigration of the isolated, labelled peptide with the purified S-100 protein in SDS-polyacrylamide gel electrophoresis and fluorography and the same retention time of the labelled S-100 protein beta subunit with authentic S-100 beta subunit by high performance liquid chromatography. The size determination of mRNA for S-100 protein on sucrose density gradient centrifugation gave 6-8 S. The assay gave a linear response with increasing amounts of polyadenylated RNA, allowing quantitation of mRNA level for S-100 protein in polyadenylated RNA. During the prenatal period and 10 postnatal days, only minute amounts of mRNA for beta subunit of S-100 protein could be found, however a dramatic increase of mRNA for beta subunit of S-100 was observed within the period of 10 to about 30 days and the mRNA level maintained a plateau from 40 days to adult age. These date indicate that the development changes in the amount of S-100 protein in the rat brain found by other authors is strongly correlated with the changes in the level of its translatable mRNA.

Distinct forms of the protein kinase-dependent activator of tyrosine and tryptophan hydroxylases.

Tyrosine and tryptophan hydroxylases are the key enzymes in the regulation of catecholamine and serotonin levels in neurons and other endocrine cells. Among the mechanisms proposed for the modulation of activity, phosphorylation of the enzyme is believed to be of functional significance with respect to the stimulus-response coupling, but the precise mechanism is unknown. Here, we show the existence of multiple, distinct forms of the 14-3-3 activator protein, a neuronal protein essential for activation of tyrosine and tryptophan hydroxylases by Ca2+/calmodulin-dependent protein kinase type II. Bovine brain 14-3-3 protein was resolved by reversed-phase chromatography into seven polypeptides (alpha to eta), all of which were active towards tryptophan hydroxylase when the renatured preparations were assayed in the presence of Ca2+, calmodulin and the protein kinase. Determination of the amino acid sequences of the beta and gamma chains and comparison of the sequences with the previously determined sequence of the eta chain revealed that these molecules are highly homologous, and share a common structural feature in containing an extremely acidic C-terminal region predicted as a domain for interaction with the phosphorylated hydroxylases. Northern blot analysis indicated that the beta, gamma and eta chain are expressed abundantly in the brain; however, these polypeptides appear to be expressed with different tissue specificities because gamma mRNA is found only in the brain, while lower levels of beta and eta mRNAs are detected in several other tissues. These findings suggest the involvement of a diverse family of the activator protein in the stimulus-coupled, Ca2(+)-dependent regulation of monoamine biosynthesis.

X-ray crystallographic and chromatographic characterization of the crystals of Ca2+-calmodulin complexed with bee venom melittin.

Crystals of calmodulin complexed with both Ca2+ and melittin, a peptide from bee venom, have been grown from 2-methyl-2,4-pentanediol solution by using the hanging drop method of vapour diffusion. The crystals belong to space group P2(1)2(1)2(1) with a = 97.3(9) A, b = 56.5(0) A, c = 33.4(9) A and Z = 4. Analyses of the dissolved crystals by high performance liquid chromatography show that the crystals contain a 1:1 complex of calmodulin and melittin. An asymmetric unit contains one such complex and the solvent content of the crystals is 47.5% (v/v).

Reduction of BRCA1 protein expression in Japanese sporadic breast carcinomas and its frequent loss in BRCA1-associated cases.

BRCA1 is a tumor suppressor gene that is responsible for hereditary breast and ovarian cancer syndrome. To clarify the possible involvement of the BRCA1 protein in mammary carcinogenesis in sporadic and hereditary forms, we have analyzed the BRCA1 protein expression pattern in five breast epithelial cell lines, including a BRCA1-deficient cell line, and 162 breast cancer tissue samples [including 108 sporadic, 35 hereditary (BRCA1 status unknown), and 19 BRCA1-associated cases] from Japanese women. Twelve anti-BRCA1 antibodies were tested by fixation conditions, in which nuclear localization of BRCA1 protein was preserved, and by specificity of the antibodies, which was evaluated in BRCA1-deficient cancer cells. Using monoclonal antibodies applicable to immunohistochemical analysis of paraffin-embedded tissue sections, we found high-level expression of BRCA1 protein in normal mammary epithelium and various degrees of reduced expression in breast cancer cells. Of the 19 BRCA1-associated breast cancer tissues, 15 (79%) showed reduction (8 cases) or complete loss (7 cases) of nuclear expression. Thirty (28%) of 108 sporadic and 6 (17%) of 35 hereditary carcinomas showed reduced BRCA1 protein expression. Reduction of BRCA1 protein expression in sporadic carcinomas was associated with solid-tubular phenotype, with poor tubular differentiation, and with an overexpression of c-erbB-2 protein, which is one of the prognostic factors in breast cancer. Our data suggest that reduced expression of BRCA1 protein may play an important role in mammary carcinogenesis, not only in BRCA1-associated breast carcinomas, but also in sporadic carcinomas, and also suggest that mechanisms other than mutation may be involved in its reduced expression.

Neonatal intramuscular injection with recombinant adeno-associated virus results in prolonged beta-glucuronidase expression in situ and correction of liver pathology in mucopolysaccharidosis type VII mice.

For many metabolic diseases, early correction of the inherited deficiency is required to prevent long-term sequelae. We examined the ability of adeno-associated virus (AAV) to mediate efficient gene transfer during the neonatal period in mice with the lysosomal storage disease mucopolysaccharidosis type VII (MPS VII). Quadriceps of newborn MPS VII mice were injected with an AAV vector containing human beta-glucuronidase (GUSB) cDNA. High-level intramuscular GUSB expression was seen as early as 2 weeks of age, and persisted for at least 16 weeks with no reduction in activity. In addition, GUSB activity was detected in both liver and spleen at later time points. The level of GUSB activity resulted in a significant reduction in lysosomal storage in the liver and a minimal reduction in the spleen at 16 weeks. However, the temporal and spatial pattern of hepatic GUSB activity, coupled with the presence of GUSB cDNA in liver sections, suggests that hematogenous dissemination of virus at the time of injection led to gene transfer to hepatic cells. These results demonstrate that AAV vectors can successfully infect neonatal muscle and persist through the rapid growth phase following birth. However, GUSB secretion from an intramuscular source is inefficient, limiting the therapeutic efficacy of this approach.

Liver-directed gene therapy: a retroviral vector with a complete LTR and the ApoE enhancer-alpha 1-antitrypsin promoter dramatically increases expression of human alpha 1-antitrypsin in vivo.

Hepatic gene therapy could improve the treatment of many inherited disorders. Although retroviral vectors result in long-term expression in hepatocytes in vivo, their low level of expression currently precludes most clinical applications. Four copies of the liver-specific apolipoprotein E (ApoE) enhancer were placed upstream of the human alpha 1-antitrypsin (hAAT) promoter in either orientation into a retroviral vector with a complete long terminal repeat (LTR) and the hAAT cDNA to generate ApoE(+)hAAT-LTR and ApoE(-)hAAT-LTR. In addition, the ApoAI promoter was placed upstream of the hAAT cDNA in a similar retroviral vector backbone. Amphotropic retroviral vectors were transferred into regenerating rat liver cells in vivo by intraportal injection. ApoE(-)hAAT-LTR and ApoE(+)hAAT-LTR led to average hAAT levels of 5 micrograms/ml (0.5% of normal levels of a very abundant protein), and 2.5 micrograms/ml, respectively, which was stable for at least 10 months after transduction. This level of serum hAAT was > 25-fold higher than what was observed from the ApoAI promoter used in this study. Serum levels of hAAT were > 15-fold higher than what was observed from retroviral vectors containing the hAAT cDNA that were analyzed previously by this lab. In some cases, improved expression was due to the promoter chosen. In other cases, the increase in expression was primarily due to the higher titers obtained by using a retroviral backbone with an intact LTR as opposed to a vector with a deletion in the LTR. The increased expression levels observed from this enhancer/promoter combination in an intact retroviral backbone may enable one to achieve therapeutic levels of clinically important genes from a retroviral vector in liver cells of animals.

Sex-determining gene(s) on distal 9p: clinical and molecular studies in six cases.

We report on clinical and molecular findings in five karyotypic males (cases 1-5) and one karyotypic female (case 6) with distal 9p monosomy. Cases 1-3 and 6 had female external genitalia, case 4 showed ambiguous external genitalia, and case 5 exhibited male external genitalia with left cryptorchidism and right intrascrotal testis. Gonadal explorations at gonadectomy in cases 3 and 4 revealed that case 3 had left streak gonad and right agonadism, and case 4 had bilateral hypoplastic testes. Endocrine studies in cases 1-4 and 6 showed that cases 1, 3, and 6 had definite primary hypogonadism, with basal FSH levels of 54, 39, and 41 IU/L, respectively, whereas case 2 with severe malnutrition was unremarkable for the baseline values, and case 4 had fairly good testicular function. Fluorescence in situ hybridization and microsatellite analyses demonstrated that all cases had hemizygosity of the 9p sex-determining region distal to D9S1779, with loss of the candidate sex-determining genes DMRT1 and DMRT2 from the abnormal chromosome 9. Sequence analysis in cases 1-4 and 6 showed that they had normal sequences of each exon of DMRT1 and the DM domain of DMRT2 on the normal chromosome 9, and that cases 1-4 had normal SRY sequence. The results provide further support for the presence of a sex-determining gene(s) on distal 9p and favor the possibility of DMRT1 and/or DMRT2 being the sex-determining gene(s). Furthermore, as hemizygosity of the 9p sex-determining region was associated with a wide spectrum of gonadogenesis from agonadism to testis formation in karyotypic males and with primary hypogonadism regardless of karyotypic sex, it is inferred that haploinsufficiency of the 9p sex-determining gene(s) primarily hinders the formation of indifferent gonad, leading to various degrees of defective testis formation in karyotypic males and impaired ovary formation in karyotypic females.

Brain micro glutamic acid-rich protein is the C-terminal endpiece of the neurofilament 68-kDa protein as determined by the primary sequence.

The amino acid sequence of bovine brain micro glutamic acid-rich protein was determined by analysis of tryptic and Trimeresurus flavoviridis protease peptides of the molecule. The protein comprised 82 amino acid residues and has an Mr of 8992. The established sequence was highly homologous (90% identity) to the sequence of C-terminal 82 residues of the neurofilament 68-kDa protein from porcine spinal cord; there are differences of 8 residues which could be species-specific amino acid substitutions. This indicates that the micro glutamic acid-rich protein may arise by a restricted proteolysis of the neurofilament 68-kDa protein, with the break occuring toward the C-terminus.

S-100 proteins and microtubules: analysis of the effects of rat brain S-100 (S-100b) and ox brain S-100a0, S-100a and S-100b on microtubule assembly-disassembly.

Rat brain S-100 (S-100b) and ox brain S-100a0, S-100a and S-100b have been tested for their ability to control the assembly and disassembly of brain microtubule proteins in the presence of either Ca2+ or Zn2+, in vitro. In the presence of Ca2+, single S-100 isoforms have similar, if not identical, effects, i.e. they inhibit assembly and promote disassembly. In the presence of Zn2+ from 0.1 to 1 mM (free concentration), rat S-100 and ox S-100a and S-100b inhibit assembly, while S-100a0 is without effect. These data are briefly discussed in relation to the cellular localization of single S-100 isoforms in the brain.

Brain 14-3-3 protein is an activator protein that activates tryptophan 5-monooxygenase and tyrosine 3-monooxygenase in the presence of Ca2+,calmodulin-dependent protein kinase II.

We have found that the 14-3-3 protein, an acidic neuronal protein, is substantially identical to the 'activator' protein [(1981) J. Biol. Chem. 256, 5404-5409] that activates tryptophan 5-monooxygenase and tyrosine 3-monooxygenase in the presence of Ca2+, calmodulin dependent protein kinase II. This finding is based on the remarkable similarity of both these proteins in physicochemical, biochemical and immunochemical properties, as well as on detection for the 14-3-3 protein of an activator activity towards tryptophan 5-monooxygenase. The result suggests that the 14-3-3 protein plays a role in the regulation of serotonin and noradrenaline biosynthesis in brain.