Dietary restriction augments erythropoiesis in mice

Previously, we have reported that a 40% dietary restriction caused microcytic and short-lived erythrocytes without anemia in mice (Internat. J. Vitr. Nutr. Res. (1989) 59, 406-412). To elucidate the mechanism of these phenomena, the effects of dietary restriction on the proliferation of erythroid precursor cells were investigated. In the diet restricted mice, a striking increase of erythropoietin (EP - responsive erythroid precursor cells without increment of EP was observed. Remarkable increases in the numbers of femoral nucleated cells and erythroid stem cells (CFU-S) were also observed in the diet restricted mice. Furthermore, the percentage of femoral CFU-S in DNA synthetic stage (S phase) revealed remarkably higher levels in the diet restricted mice than in the control mice. Our results strongly suggest that the microcytic change of erythrocytes in the diet restricted mice may be attributed to the increased number of erythroid precursor cells without an increment of hemoglobin synthesis and that the avoidance of becoming anemic from the short-lived erythrocytes in the diet restricted mice may be caused by a vigorous erythropoiesis.     

Monoclonal antibodies reveal cell-type-specific antigens in the sexually dimorphic olfactory system of Manduca sexta. I. Generation of monoclonal antibodies and partial characterization of the antigens

The olfactory system of the moth Manduca sexta is sexually dimorphic. Male moths possess a male-specific olfactory "subsystem," comprising olfactory receptor cells (ORCs) and CNS neurons and synaptic areas associated with the detection of female sex pheromones, in addition to elements common to males and females. In order to explore the molecular differences between cells that subserve the sexual dimorphism and odor-specificity of components of the olfactory system, we generated monoclonal antibodies (Mabs) against tissue of the olfactory system of the moth. In 2 fusions, we screened 1105 hybridoma lines and obtained 272 lines that secreted antibodies against Manduca nervous tissue, as assayed immunocytochemically on sections of the primary olfactory center (the antennal lobe) in the brain of Manduca. We describe here 3 classes of Mabs exemplifying the several cell-type-specific antibodies obtained through the screening procedure. Seven hybridoma lines secrete antibodies that specifically recognize cell bodies, axons, and initial segments of dendrites of many or all ORCs of both males and females (classified as olfactory-specific antibodies, OSAs). Electron-microscopic studies of 2 of the Mabs in this class showed that they recognize antigens associated with the cell membrane and that the immunoreactive ORC axons are bundled together in fascicles in the antennal nerve. On immunoblots, one of the OSA Mabs recognizes 3 distinct protein bands of apparent Mrs 42,000, 59,000, and 66,000 Da. When tissue samples enriched in either receptor cell bodies, dendrites, and initial segments of axons or in distal segments of axons and their terminals and synapses were extracted separately, different patterns of bands were detected--42,000 and 59,000 Da bands from cell bodies and initial segments of axons and dendrites, and 42,000 and 66,000 Da bands from distal segments of axons and their terminals--suggesting that the 59,000 Da protein is modified to the 66,000 Da protein during axonal transport. The second Mab we describe here, the male olfactory-specific antibody (MOSA), selectively recognizes the sexually dimorphic ORCs that are present only in males. The antigen recognized by this antibody is found in cell bodies, dendrites, axons, and axon terminals. By electron-microscopic immunocytochemistry, the MOSA immunoreactivity is found in the cytoplasm and appears not to be associated with particular subcellular organelles. This antibody demonstrates that male-specific ORCs are molecularly distinct from other types of ORCs.(ABSTRACT TRUNCATED AT 400 WORDS)     

HUI Mālama O Ke Kai: a positive prevention‐based youth development program based on native hawaiian values and activities

Evaluation of after‐school programs that are culturally and place‐based and promote positive youth development among minority and indigenous youths has not been widely published. The present evaluation is the first of its kind of an after‐school, youth‐risk prevention program called Hui Malāma O Ke Kai (HMK), that emphasizes Native Hawaiian values and activities to promote positive youth development for fifth and sixth graders (N=110) in a rural Native Hawaiian community. Results indicated positive gains on youth self‐reports in Native Hawaiian values, self‐esteem, antidrug use, violence prevention strategies, and healthy lifestyle in Year 1, and in family cohesion, school success, and violence prevention strategies in Year 2. Parent reports of their children indicated positive gains in selected domains. Implications include the support for a promising culturally appropriate program, expansion to middle‐school‐aged youths, and parent involvement. © 2009 Wiley Periodicals, Inc.     

Effects of epidermal growth factor, phorbol ester, and retinoic acid on hormone synthesis and morphology in porcine thyroid follicles cultured in collagen gel.

Epidermal growth factor (EGF), phorbol esters (PEs), and retinoic acid (RA) inhibit differentiated functions of thyrocytes. In the present study the inhibitory effects of these growth-promoting factors on hormone synthesis were studied in thyroid follicles cultured in type-I collagen gel, and morphologic alteration by these factors was examined by light and electron microscopy (EM). Porcine open thyroid follicles obtained by treatment with 0.1% collagenase were embedded in collagen gel and cultured in Ham's F12 medium supplemented with 6H (insulin, hydrocortisone, somatostatin, transferrin, glycyl-his-lys, and thyrotropin) + 0.5% fetal bovine serum (FBS). After 1 week these open follicles developed to closed follicles, and the medium was changed to one containing 6H + 0.5% FBS + 0.1 microM sodium iodide (NaI). Some media were supplemented with either EGF, phorbol 12-myristate 13-acetate (PMA), or all-trans RA. The closed follicles retained ability for hormone synthesis for 2 weeks after the medium change in the presence of 6H + FBS + NaI. The amounts of T4 and T3 secreted into the culture medium from day 9 to day 12 after the medium change were 60% and 45% of those from day 0 to day 4, respectively. EGF reduced production of T4 and T3 by 61% and 69%, respectively; PMA, by 87% and 99%; and RA, by 55% and 44%. In the medium supplemented with 6H + 0.5% FBS, the follicles exhibited intact polarity. Apical surfaces with microvilli were oriented to the follicular lumen and tight junctions were on the apical side of cell-to-cell contacts. Desmosomes were found on both the apical and basal halves of the cell contacts.(ABSTRACT TRUNCATED AT 250 WORDS)     

Involvement of tumor necrosis factor-alpha in development of hepatic injury in galactosamine-sensitized mice

Intravenous injection of lipopolysaccharide and D-galactosamine, at doses of 0.2 micrograms/kg and 800 mg/kg, respectively, elicited massive hepatic necrosis within 24 hr in C3H/HeN mice. The plasma L-alanine aminotransferase (ALT, E.C. 2.6.1.2) or L-aspartate aminotransferase (AST, E.C. 2.6.1.1) activities at this point reached more than 2,000 IU/L. However, overt hepatic injury as evaluated by the plasma aminotransferase activities did not develop in mice in which only lipopolysaccharide or only D-galactosamine was injected. No tumor necrosis factor-like activities could be detected in the plasma of galactosamine- and lipopolysaccharide-injected mice as determined by the assay of cytotoxicity to highly tumor necrosis factor-sensitive L-P3 cells through the experimental period of 24 hr. However, passive immunization against mouse tumor necrosis factor-alpha with polyvalent rabbit anti-mouse tumor necrosis factor-alpha antiserum, which was able to neutralize the cytotoxic effects of recombinant mouse tumor necrosis factor-alpha on L-P3 cells, could protect the mice from the development of hepatic injury in a dose-dependent manner. Simultaneous injection of recombinant human tumor necrosis factor-alpha, instead of lipopolysaccharide, with 800 mg/kg of D-galactosamine in lipopolysaccharide-resistant C3H/HeJ mice sensitized the animals more than one thousand-fold to the development of hepatic injury. The livers appeared to be morphologically similar to those of galactosamine- and lipopolysaccharide-injected C3H/HeN mice.     

A Mutation of RNA Polymerase β′ Subunit (RpoC) Converts Heterogeneously Vancomycin-Intermediate Staphylococcus aureus (hVISA) into “Slow VISA”

Various mutations in the rpoB gene, which encodes the RNA polymerase β subunit, are associated with increased vancomycin (VAN) resistance in vancomycin-intermediate Staphylococcus aureus (VISA) and heterogeneously VISA (hVISA) strains. We reported that rpoB mutations are also linked to the expression of the recently found “slow VISA” (sVISA) phenotype (M. Saito, Y. Katayama, T. Hishinuma, A. Iwamoto, Y. Aiba, K Kuwahara-Arai, L. Cui, M. Matsuo, N. Aritaka, and K. Hiramatsu, Antimicrob Agents Chemother 58:5024–5035, 2014, http://dx.doi.org/10.1128/AAC.02470-13). Because RpoC and RpoB are components of RNA polymerase, we examined the effect of the rpoC(P440L) mutation on the expression of the sVISA phenotype in the Mu3fdh2*V6-5 strain (V6-5), which was derived from a previously reported hVISA strain with the VISA phenotype. V6-5 had an extremely prolonged doubling time (DT) (72 min) and high vancomycin MIC (16 mg/liter). However, the phenotype of V6-5 was unstable, and the strain frequently reverted to hVISA with concomitant loss of low growth rate, cell wall thickness, and reduced autolysis. Whole-genome sequencing of phenotypic revertant strain V6-5-L1 and comparison with V6-5 revealed a second mutation, F562L, in rpoC. Introduction of the wild-type (WT) rpoC gene using a multicopy plasmid resolved the sVISA phenotype of V6-5, indicating that the rpoC(P440L) mutant expressed the sVISA phenotype in hVISA. To investigate the mechanisms of resistance in the sVISA strain, we independently isolated an additional 10 revertants to hVISA and VISA. In subsequent whole-genome analysis, we identified compensatory mutations in the genes of three distinct functional categories: the rpoC gene itself as regulatory mutations, peptidoglycan biosynthesis genes, and relQ, which is involved in the stringent response. It appears that the rpoC(P440L) mutation causes the sVISA phenotype by augmenting cell wall peptidoglycan synthesis and through the control of the stringent response.