Sensory neurite outgrowth on white matter astrocytes is influenced by intracellular and extracellular S100A4 protein

The central nervous system (CNS) is considered a nonpermissive environment for axonal regeneration because of the presence of myelin and associated repulsive molecules. However, neural cells transplanted to the CNS preferably migrate and extend their fibers in white matter areas. We previously showed that white matter astrocytes in vivo express the calcium-binding protein S100A4, which is strongly up-regulated in areas of white matter degeneration. To investigate the role of white matter astrocytes and their specific protein S100A4 in axonal regeneration, we developed white matter astrocyte cultures with strong S100A4 expression and grew dissociated adult dorsal root ganglion (DRG) cells on top of astrocytes for 24 hr. By using small interfering S100A4 RNA, we were able to eliminate S100A4 expression and compare growth of DRG cell neurites on S100A4-silenced and S100A4-expressing astrocytes. In addition, we studied whether extracellular S100A4 has an effect on neurite growth from adult DRG cells cultured on S100A4-expressing white matter astrocytes. Our data show that white matter astrocytes are permissive for neurite growth, although high levels of S100A4 in white matter astrocytes have a negative effect on this growth. Extracellular application of S100A4 induced extensive growth of DRG cell neurites on white matter astrocytes. These findings suggest that white matter astrocytes are able to support axonal regeneration and, furthermore, that administration of extracellular S100A4 provides strong additional support for axonal regeneration.     

Mechanism of liver tyrosine aminotransferase increase in ethanol-treated mice and its effect on serum tyrosine level

Liver tyrosine aminotransferase (TAT) activity is known to increase with ethanol treatment; however, the mechanism of this increase is unclear. Upon investigation we found that TAT activity and mRNA levels started to increase 2 h after ethanol administration and continued to increase until 6 h after ethanol administration. The increase in ethanol-induced TAT activity could not be explained by calorie loading after fasting, since ethanol loading increased TAT expression, while glucose loading decreased TAT expression. In addition, liver TAT activity was not related to serum tyrosine levels. TAT activity increased when an adenosine A2 agonist, 5'-N-ethylcarboxamide adenosine, was given. Since TAT activity is increased by cAMP, and ethanol increases cAMP production via an adenosine receptor-dependent mechanism, this increase in ethanol-induced TAT activity may occur via an adenosine receptor-dependent mechanism.     

A Murine Monoclonal Antibody that Recognizes an Extracellular Domain of the Human c-erbB-2 Protooncogene Product

A marine IgM monoclonal antibody, designated SV2–61, was generated against human c-erbB-2 gene-transfected NIH-3T3 (SV11) cells. SV2–61 defined a 185-kDa molecule present on the surface of SV11 cells, another line of c-erbB-2 gene-transfected NIH-3T3 (A4–15) cells, and MKN-7 human gastric cancer cell line carrying an amplified human c-erbB-2 gene. The SV2–61-defined antigen was found to show protein kinase activity in vitro . The SV2–61 was reactive with human c-erbB-2 gene-transfected NIH-3T3 cell lines but not with transfectants carrying c-erbB-2 gene mutants which lack a coding region for the extracellular domain. It was reactive with a portion of human epithelial cell lines hut not with native NIH-3T3, TGF-a-coding gene-, activated c-raf gene- or Ha-ras gene-transfected NIH-3T3 cells, or non-epithelial human cells. These results indicate that the SV2–61 is an antibody which recognizes an extracellular domain of the c-erbB-2 gene product, 185-kDa protein.     

Transcorneal electrical stimulation of retina to treat longstanding retinal artery occlusion

Purpose To report the outcome of transcorneal electrical stimulation (TES) of the visual system on long-standing retinal artery occlusion (RAO). Design Open labeled, case series. Patients and methods Two patients with central RAO (15 and 33 months respectively) and one with branch RAO (26 months) underwent TES therapy. Subjective and objective ophthalmological evaluations were performed before and after the TES. The ages of the patients were 38, 49, and 63 years. The TES (20 Hz biphasic pulses, 30 minutes, up to 1100 uA) was delivered by a bipolar contact lens electrode once a month for 3 months. Perimetric and/or electrophysiological examinations were performed as outcome measures. Results The visual acuity improved by more than 0.2 logMAR units in two cases, and the visual fields were improved in all three cases. The multifocal ERGs which had been reduced in the loci corresponding to the ischemic retinal area were improved after the treatment in two cases. Neither ocular nor systemic adverse effects were observed except for transient superficial keratitis. Conclusions TES of the retina can improve retinal function in eyes with long-standing RAOs. None of the authors have any financial or proprietary interest in any material or methods mentioned. This study was supported by Researches on Sensory and Communicative Disorders from the Ministry of Health, Labor, and Welfare, Japan.     

Involvement of Sl00–related Calcium–binding Protein pEL98 (or mts1) in Cell Motility and Tumor Cell Invasion

We examined the relationship between cell motility and the expressions of pEL9S ( mtsl ) mRNA and protein in various imirine normal and transformed cells. The expression of pEL98 ( mtsl ) in v–Ha– ras –transformed NIH3T3 cells and in normal rat kidney cells transformed by either v–Ha– ras or v–src was increased over that in the corresponding parental cells at both mRNA and protein levels. The expression in normal rat fibroblasts (3Y1) transformed by v–Ha– ras was also increased compared with that in 3Y1 cells. However, the expression of pEL98 (mtsl) in 3Y1 cells transformed by v –src was increased in one clone (src 3Y1–K), but decreased in another clone (src 3Y1–H). The expression level of pEL98 (mtsl) correlated well with cell motility, which was examined by measuring cell tracks by phagokinesis. In order to test direct involvement of the pEL98 ( mtsl ) protein in cell motility, src 3Y1–H cells that showed low cell motility were transfected with pEL98 cDNA. The transfectants expressing large amounts of the pEL98 protein showed significantly higher cell motility than src 3Y1–H cells. The expression of pEL98 ( mtsl ) was also found to be correlated with motile and invasive abilities in various clones derived from Lewis lung carcinoma. These results suggest that the pEL98 ( mtsl ) protein plays a role in regulating cell motility and tumor cell invasiveness.     

Presence in Pieris rapae of Cytotoxic Activity against Human Carcinoma Cells

Cytotoxic activity in extracts of pupae and adults of various kinds of butterflies and moths was tested in vitro against the human gastric carcinoma cell line, TMK-1, which was chosen as an example of human carcinoma cells. Among the species examined, cytotoxicity was limited to Pieris rapae, Pieris napi and Pieris brassicae . Activity was found down to a dilution of 1/10⁴, while with the other butterflies and moths no activity was observed, even at 1/10². When the cytotoxicity of the three developmental stages, larvae, pupae and adults, of Pieris rapae was compared, the pupae showed the strongest activity, the IC₅₀ against TMK-1 cells being at the 1/10⁶ dilution. For larvae and adults, the respective IC₅₀ values were at the 1/10⁵ and 5/10⁵ dilutions. The active principle in the pupae of Pieris rapae was found to be heat-labile and not extractable with organic solvents, but precipitated with ammonium sulfate and digested by proteases, suggesting that it is a protein. This cytotoxic factor was named pierisin.     

Treatment or prevention of herpes simplex virus retinitis with intravitreally injectable crystalline 1-O-hexadecylpropanediol-3-phospho-ganciclovir

PURPOSE: To evaluate an intraocular drug delivery system consisting of the crystalline ammonium salt of 1-O-hexadecylpropanediol-3-phospho-ganciclovir (HDP-P-GCV) as a slow-release form of the drug. METHODS: A dosage of 0.885, 1.57, 2.8, 4.486, or 8.85 micromol of ammonium salt HDP-P-GCV in 0.1 mL was intravitreally injected into rabbit vitreous. The toxicity and safety were evaluated with ophthalmoscopy, electroretinography, and pathology. Drug vitreous levels were determined at various time intervals by means of HPLC. The treatment efficacy and duration of efficacy were tested in a herpes simplex virus (HSV)-1 retinitis rabbit model. RESULTS: Intravitreal injections of the compound revealed clear vitreous of optic axis, a desirable drug depot in the inferior vitreous cavity, and no clinical toxicity, except for variable mild local posterior subcapsular cataract and local retinal toxicity with high doses. HPLC analysis showed free ammonium salt of HDP-P-GCV in the upper vitreous at a level of 0.2 microM 12 weeks after the 2.8-micromol initial intravitreal dose. Drug concentration was still 1.95 microM 20 weeks after the 8.85-micromol initial intravitreal dose. These concentrations (0.2 and 1.95 microM) were 10 and 100 times higher, respectively, than the median inhibitory concentration (IC(50)) of HSV-1 (0.023 microM). Treatment with the highest nontoxic dose (2.8 micromol) and the highest dose (8.85 micromol) showed significant protection from HSV-1 infection (P < 0.05) and provided sustained antiviral effect after a single intravitreal drug injection. CONCLUSIONS: The crystalline ammonium salt of HDP-P-GCV may be a very useful local therapy for herpes family viral retinitis.     

Predictors of drusen reduction after subthreshold infrared (810 nm) diode laser macular grid photocoagulation for nonexudative age-related macular degeneration

PURPOSE: To determine the predictors of drusen reduction in eyes with nonexudative age-related macular degeneration (ARMD) treated with subthreshold infrared (810 nm) diode laser macular grid photocoagulation. Additionally, to determine the relationship of laser-induced drusen reduction and best-corrected visual acuity (BCVA) 18 months after laser treatment.DESIGN: Randomized controlled clinical trial.METHODS: Fifty patients (100 eyes) with bilateral nonexudative ARMD were enrolled at two centers. One eye of each patient was randomized to the observation; the other eye was treated with 48 subthreshold (invisible end point) applications of infrared (810 nm) diode laser in a macular grid pattern. The eyes that received subthreshold laser treatment were compared with the eyes that received no treatment. The baseline fundus characteristics (number, size, and distribution of drusen, as well as focal hyperpigmentation) from two macula areas (central 1500 micro diameter, pericentral 1500 micro ring area) on stereo color photographs, the number of laser-induced lesions, and the area of laser induced retinal pigment epithelial (RPE) lesions on fluorescein angiography 3 months after treatment were studied as predictors of major drusen reduction (> or = 50% drusen reduction from baseline) 18 months after laser treatment. BCVA at baseline and 18 months later was compared in observation eyes and in laser-treated eyes.RESULTS: Eighteen months after randomization, 24 (48%) of 50 eyes treated with subthreshold laser had major drusen reduction compared with three (6%) of 50 observation eyes (P =.00001). At 3 months post-treatment in laser-treated eyes with major drusen reduction, the mean number of laser-induced lesions on fluorescein angiography was 30.7 and the mean area of RPE change was 0.81 mm(2) compared with 14.8 laser-induced lesions and 0.35 mm(2) area of RPE change in eyes without major drusen reduction (P =.0001 and P =.0003, respectively). At baseline, fundus characteristics were not significantly different between observation eyes and laser-treated eyes or between the major drusen reduction group and the nonmajor drusen reduction group. At 18 months after treatment, BCVA was not significantly different in laser-treated eyes and in observation eyes.CONCLUSIONS: Subthreshold infrared (810 nm) diode laser macular grid photocoagulation in eyes with nonexudative ARMD significantly reduced drusen 18 months after laser treatment. Both the number of subthreshold laser lesions and the area of RPE changes visible on fluorescein angiography 3 months after treatment appeared to be predictors for major drusen reduction 18 months after treatment. However, it remains to be determined whether laser-induced drusen reduction is beneficial for visual acuity or reduces the incidence of choroidal neovascularization (CNV) in eyes with nonexudative ARMD.     

Anti-osteopenic effect of taurine: possible involvement of activated MEK-ERK-Cbfa1 signaling

Previously we first noted that taurine (TR) has anti-osteopenic effect on low Ca diet-induced osteopenia in rats (1). Employing osteoblastic MC3T3-E1 cells, the mechanism of the anti-osteopenic effect was explored in vitro. TR (1 mM) was found to promote mineralization of extracellular matrices, without affecting alkaline phosphataase activity. Gel shift assay using 32P-labeled OSE2 (osteoblast-specific cis-element 2: the consensus sequence for Cbfa1, refer to 2) indicated that TR (1 mM) increased the nuclear localization of Cbfa1, just as TPH (1-34) (3,4) and bisphosphonates did (5). In addition, TR was found to stimulate ERK phosphorylation. PD98059, a MEK inhibitor, suppressed effects of TR on both Cbfa1 transactivation and ERK activation. The results strongly suggest that TR first activates intracellular MEK-ERK-Cbfa1 signaling system thereby promoting mineralization and finally leading to its bone anabolic action.     

General practitioner's recognition of service cooperation with community pharmacists related to home care in a suburban area

OBJECTIVES: To clarify the factors affecting a general practitioner's demands and the recognition of service cooperation with community pharmacists related to home care in a suburban area. METHODS: A questionnaire on pharmacy services and functions was administered to 215 general practitioners in a suburban area. Gender, age, specialty, length of practice, status of home visiting, requests for community pharmacists, awareness of a home visiting service by community pharmacists, status of issues about home visiting orders to community pharmacists and the criteria for deciding to issue an order, recognition of the necessity of a home visiting service by pharmacists, expectation value to the community of the pharmacists' participation in the home care service and requests, recognition of cooperation with other home care related professions, ease of cooperation with community pharmacists, disincentives for cooperation with community pharmacists, factors necessary to promote cooperation between practitioners and community pharmacists, and factors necessary to promote the home care system were surveyed. Using the results, chi 2 test and principal component analysis were performed. RESULTS: It was found that the general practitioners' main demands were support and management of pharmaceutical therapy. Meanwhile, the practitioners' low cognition of pharmacists' home visiting seemed to be one disincentive to cooperation with pharmacists. Every practitioner who had issued visiting orders to pharmacists practiced home visiting and issued the visiting orders based on patients' condition at home. Practitioners who practiced home visiting were more active in promoting the home care system and had a better cognition and more extensive demands for pharmacists' home visiting compared to practitioners with no home visiting. Practitioners with good recognition of cooperation with community pharmacists had better cognition, realized the necessity of, and expected pharmacist's home visiting compared to practitioners with low awareness. However, practitioners' overall recognition of cooperation with pharmacists was relatively low. As disincentives, practitioners pointed out lack of acquaintance, the unclear function of pharmacists, and no opportunity for cooperation. So, enhancing practitioners' awareness of home care, clarifying the pharmacist's role, establishing a relationship of mutual trust through information feedback and/or exchange based on practice would be effective in promoting service cooperation with community pharmacists.