A gain-of-function screen identifies wdb and lkb1 as lifespan-extending genes in Drosophila

The insulin/insulin-like growth factor (IGF) and the target of rapamycin (TOR) signaling pathways are known to regulate lifespan in diverse organisms. However, only a limited number of genes involved in these pathways have been examined regarding their effects on lifespan. Through a gain-of-function screen in Drosophila, we found that overexpression of the wdb gene encoding a regulatory subunit of PP2A, and overexpression of the lkb1 gene encoding a serine/threonine kinase, reduced organ size and extended lifespan. Overexpression of wdb also reduced the level of phosphorylated AKT, while overexpression of lkb1 increased the level of phosphorylated AMPK and decreased the level of phosphorylated S6K. Taken together, our results suggest that wdb- and lkb1-dependent lifespan extension is mediated by downregulation of S6K, a downstream component of the insulin/IGF and TOR signaling pathways.     

Mineralocorticoid receptor activation: a major contributor to salt-induced renal injury and hypertension in young rats

Excessive salt intake is known to preferentially increase blood pressure (BP) and promote kidney damage in young, salt-sensitive hypertensive human and animal models. We have suggested that mineralocorticoid receptor (MR) activation plays a major role in kidney injury in young rats. BP and urinary protein were compared in young (3-wk-old) and adult (10-wk-old) uninephrectomized (UNx) Sprague-Dawley rats fed a high (8.0%)-salt diet for 4 wk. The effects of the MR blocker eplerenone on BP and renal injury were examined in the high-salt diet-fed young UNx rats. Renal expression of renin-angiotensin-aldosterone (RAA) system components and of inflammatory and oxidative stress markers was also measured. The effects of the angiotensin receptor blocker olmesartan with or without low-dose aldosterone infusion, the aldosterone synthase inhibitor FAD286, and the antioxidant tempol were also studied. Excessive salt intake induced greater hypertension and proteinuria in young rats than in adult rats. The kidneys of young salt-loaded rats showed marked histological injury, overexpression of RAA system components, and an increase in inflammatory and oxidative stress markers. These changes were markedly ameliorated by eplerenone treatment. Olmesartan also ameliorated salt-induced renal injury but failed to do so when combined with low-dose aldosterone infusion. FAD286 and tempol also markedly reduced urinary protein. UNx rats exposed to excessive salt at a young age showed severe hypertension and renal injury, likely primarily due to MR activation and secondarily due to angiotensin receptor activation, which may be mediated by inflammation and oxidative stress.     

A novel mechanism in maggot debridement therapy: protease in excretion/secretion promotes hepatocyte growth factor production

Maggot debridement therapy (MDT) is effective for treating intractable wounds, but its precise molecular mechanism, including the association between MDT and growth factors, remains unknown. We administered MDT to nine patients (66.3 ± 11.8 yr, 5 male and 4 female) with intractable wounds of lower extremities because they did not respond to conventional therapies. Significant increases of hepatocyte growth factor (HGF) levels were observed in femoral vein blood during 48 h of MDT (P < 0.05), but no significant change was found for vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), transforming growth factor-β1 (TGF-β1), or tumor necrosis factor-α (TNF-α). We conducted NIH-3T3 cell stimulation assay to evaluate the relation between HGF and protease activity in excretion/secretion (ES) derived from maggots. Compared with the control group, HGF was significantly higher in the 0.05 μg/ml ES group (P < 0.01). Furthermore, protease inhibitors suppressed the increase of HGF (P < 0.05). The HGF expression was increased in proportion to the ES protein concentration of 0.025 to 0.5 μg/ml. In fact, ES showed stronger capability of promoting HGF production and less cytotoxicity than chymotrypsin or bromelain. HGF is an important factor involved in cutaneous wound healing. Therefore, these results suggest that formation of healthy granulation tissue observed during MDT results from the increased HGF. Further investigation to identify molecules enhancing HGF expression by MDT will contribute greatly to drug target discovery for intractable wound healing therapy.     

Mitochondrial dysfunction and increased reactive oxygen species impair insulin secretion in sphingomyelin synthase 1-null mice

Sphingomyelin synthase 1 (SMS1) catalyzes the conversion of ceramide to sphingomyelin. Here, we generated and analyzed SMS1-null mice. SMS1-null mice exhibited moderate neonatal lethality, reduced body weight, and loss of fat tissues mass, suggesting that they might have metabolic abnormality. Indeed, analysis on glucose metabolism revealed that they showed severe deficiencies in insulin secretion. Isolated mutant islets exhibited severely impaired ability to release insulin, dependent on glucose stimuli. Further analysis indicated that mitochondria in mutant islet cells cannot up-regulate ATP production in response to glucose. We also observed additional mitochondrial abnormalities, such as hyperpolarized membrane potential and increased levels of reactive oxygen species (ROS) in mutant islets. Finally, when SMS1-null mice were treated with the anti-oxidant N-acetyl cysteine, we observed partial recovery of insulin secretion, indicating that ROS overproduction underlies pancreatic β-cell dysfunction in SMS1-null mice. Altogether, our data suggest that SMS1 is important for controlling ROS generation, and that SMS1 is required for normal mitochondrial function and insulin secretion in pancreatic β-cells.     

Behavioral changes in the termite, Coptotermes formosanus (Isoptera), inoculated with six fungal isolates

The studies of pathogen-prevention behaviors of termites have focused on hygiene behavior directed only against highly virulent pathogens. Therefore, we compared behavioral changes in the subterranean termite Coptotermes formosanus following contact with entomopathogenic fungi with different levels of virulence. The fungal virulence was inferred from the daily mortality and the LD₅₀ value in previous data. When untreated termites were allowed to contact their fungus-inoculated nestmates, mutual grooming was frequent during 30 min after inoculation. The inoculated termites were often attacked and eaten by their uninoculated nestmates, and then buried after death. Notably, there was no influence of fungal virulence on these pathogen-prevention behaviors. However, the fungal isolates and genera affected not only the frequency of the behaviors but also the horizontal transmission pattern, the number of dead individuals and the survival period before the first death following infection.     

Hypoxia-Inducible Factor 1 Regulation through Cross Talk between mTOR and MT1-MMP

Hypoxia-inducible factor 1 (HIF-1) plays a key role in the cellular adaptation to hypoxia. Although HIF-1 is usually strongly suppressed by posttranslational mechanisms during normoxia, HIF-1 is active and enhances tumorigenicity in malignant tumor cells that express the membrane protease MT1-MMP. The cytoplasmic tail of MT1-MMP, which can bind a HIF-1 suppressor protein called factor inhibiting HIF-1 (FIH-1), promotes inhibition of FIH-1 by Mint3 during normoxia. To explore possible links between HIF-1 activation by MT1-MMP/Mint3 and tumor growth signals, we surveyed a panel of 252 signaling inhibitors. The mTOR inhibitor rapamycin was identified as a possible modulator, and it inhibited the mTOR-dependent phosphorylation of Mint3 that is required for FIH-1 inhibition. A mutant Mint3 protein that cannot be phosphorylated exhibited a reduced ability to inhibit FIH-1 and promoted tumor formation in mice. These data suggest a novel molecular link between the important hub proteins MT1-MMP and mTOR that contributes to tumor malignancy.     

Galectin-9 Enhances Cytokine Secretion,but Suppresses Survival and Degranulation,in Human Mast Cell Line

Galectin-9 (Gal-9), a lectin having a β-galactoside-binding domain, can induce apoptosis of Th1 cells by binding to TIM-3. In addition, Gal-9 inhibits IgE/Ag-mediated degranulation of mast cell/basophilic cell lines by binding to IgE, thus blocking IgE/Ag complex formation. However, the role of Gal-9 in mast cell function in the absence of IgE is not fully understood. Here, we found that recombinant Gal-9 directly induced phosphorylation of Erk1/2 but not p38 MAPK in a human mast cell line, HMC-1, which does not express FcεRI. Gal-9 induced apoptosis and inhibited PMA/ionomycin-mediated degranulation of HMC-1 cells. On the other hand, Gal-9 induced cytokine and/or chemokine production by HMC-1 cells, dependent on activation of ERK1/2 but not p38 MAPK. In addition, the lectin activity of Gal-9 was required for Gal-9-mediated cytokine secretion by HMC-1 cells. These observations suggest that Gal-9 has dual properties as both a regulator and an activator of mast cells.     

Food Sources for Ruditapes philippinarum in a Coastal Lagoon Determined by Mass Balance and Stable Isotope Approaches

The relationship between the food demand of a clam population (Ruditapes philippinarum (Adams & Reeve 1850)) and the isotopic contributions of potential food sources (phytoplankton, benthic diatoms, and organic matter derived from the sediment surface, seagrass, and seaweeds) to the clam diet were investigated. In particular, we investigated the manner in which dense patches of clams with high secondary productivity are sustained in a coastal lagoon ecosystem (Hichirippu Lagoon) in Hokkaido, Japan. Clam feeding behavior should affect material circulation in this lagoon owing to their high secondary productivity (ca. 130 g C m⁻² yr⁻¹). Phytoplankton were initially found to constitute 14–77% of the clam diet, although phytoplankton nitrogen content (1.79–4.48 kmol N) and the food demand of the clam (16.2 kmol N d^–1) suggest that phytoplankton can constitute only up to 28% of clam dietary demands. However, use of isotopic signatures alone may be misleading. For example, the contribution of microphytobenthos (MPB) were estimated to be 0–68% on the basis of isotopic signatures but was subsequently shown to be 35±13% (mean ± S.D.) and 64±4% (mean ± S.D.) on the basis of phytoplankton biomass and clam food demand respectively, suggesting that MPB are the primary food source for clams. Thus, in the present study, the abundant MPB in the subtidal area appear to be a key food source for clams, suggesting that these MPB may sustain the high secondary production of the clam.     

Adoption of lycaenid Niphanda fusca (Lepidoptera: Lycaenidae) caterpillars by the host ant Camponotus japonicus (Hymenoptera: Formicidae)

Caterpillars of the parasitic lycaenid butterfly are often adopted by host ants. It has been proposed that this adoption occurs because the caterpillars mimic the cuticular hydrocarbons of the host ant. This study aimed to examine whether caterpillars of the Japanese lycaenid butterfly Niphanda fusca induce adoption by mimicking their host ant Camponotus japonicus. Behavioral observations conducted in the laboratory showed that most second‐instar caterpillars were not adopted, whereas most third‐instar caterpillars were successfully adopted by host workers. A chemical comparison detected no characteristic differences in the cuticular hydrocarbon profiles between second‐ and third‐instar caterpillars. However, morphological features of the caterpillars differed between the second and third instars; third‐instar caterpillars developed exocrine glands (ant organs) such as tentacle organs and a dorsal nectary organ. These results suggest that multiple chemical signatures, not only cuticular hydrocarbons, may be important for invasion of the host ant nest.     

Fish oil diet affects on oxidative senescence of red blood cells linked to degeneration of spleen cells in mice

The effect of dietary polyunsaturated fatty acids and α-tocopherol supplementation on erythrocyte lipid peroxidation and immunocompetent cells in mice was studied comparatively using seven dietary oils (15% oil/diet, w/w) including fish oil rich in eicosapentaenoic acid (EPA, 20:5, n–3) and docosahexaenoic acid (DHA, 22:6, n–3). A 43% increase in spleen weight, about twice as many spleen cells and no change in the subpopulations of spleen cells, as well as a significant depression of mitogen-induced blastogenesis of both T and B cells in the spleen were observed in mice fed fish oil for 30 days in comparison with soybean oil diet-fed mice. In the fish oil diet-fed mice, membranous lipid hydroperoxide (hydroperoxides of phosphatidylcholine and phosphatidylethanolamine) accumulation as a marker of oxidative senescence in red blood cells (RBC) was 2.7–3.5 times higher than that in mice fed soybean oil, although there was no difference in the plasma phosphatidylcholine hydroperoxide concentration. In spite of the supplementation of α-tocopherol to up to 10 times the level in the basal diet, the degeneration of spleen cells and the stimulated oxidative senescence of RBC found by the fish oil feeding could not be prevented. The results suggest that oral intake of excess polyunsaturated fatty acids, i.e. EPA and DHA, in a fish oil diet can lead to acceleration of membrane lipid peroxidation resulting in RBC senescence linked to the lowering of immune response of spleen cells, and that supplementation of α-tocopherol as antioxidant does not always effectively prevent such oxidative degeneration as observed in spleen cells and RBC in vivo.