Mu-opioid receptor activation in the habenula modulates synaptic transmission and depression-like behaviors.

Opioid system dysregulation in response to stress is known to lead to psychiatric disorders including major depression. Among three different types of opioid receptors, the mu-type receptors (mORs) are highly expressed in the habenula complex, however, the action of mORs in this area and its interaction with stress exposure is largely unknown. Therefore, we investigated the roles of mORs in the habenula using male rats of an acute learned helplessness (aLH) model. First, we found that mOR activation decreased both excitatory and inhibitory synaptic transmission onto the lateral habenula (LHb). Intriguingly, this mOR-induced synaptic depression was reduced in an animal model of depression compared to that of controls. In naïve animals, we found an unexpected interaction between mORs and the endocannabinoid (eCB) signaling occurring in the LHb, which mediates presynaptic alteration occurring with mOR activation. However, we did not observe presynaptic alteration by mOR activation after stress exposure. Moreover, selective mOR activation in the habenula before, but not after, stress exposure effectively reduced helpless behaviors compared to aLH animals. Our observations are consistent with clinical reports suggesting the involvement of mOR signaling in depression, and additionally reveal a critical time window of mOR action in the habenula for ameliorating helplessness symptoms.

Ancistrocladus tectorius Extract Inhibits Obesity by Promoting Thermogenesis and Mitochondrial Dynamics in High-Fat Diet-Fed Mice.

Root extracts of Ancistrocladus tectorius (AT), a shrub native to China, have been shown to have antiviral and antitumor activities, but the anti-obesity effects of AT aerial parts, mainly the leaves and stems, have not been investigated. This study is the first to investigate the anti-obesity effects and molecular mechanism of AT 70% ethanol extract in 3T3-L1 adipocytes and high-fat diet (HFD)-fed C57BL/6J mice. Treatment with AT extract inhibited lipid accumulation in 3T3-L1 cells and decreased the expression of adipogenesis-related genes. AT extract also upregulated the mRNA expression of genes related to mitochondrial dynamics in 3T3-L1 adipocytes. AT administration for 12 weeks reduced body weight and organ weights, including liver, pancreas, and white and brown adipose tissue, and improved plasma profiles such as glucose, insulin, homeostasis model assessment of insulin resistance, triglyceride (TG), and total cholesterol in HFD-fed mice. AT extract reduced HFD-induced hepatic steatosis with levels of liver TG and lipogenesis-related genes. AT extract upregulated thermogenesis-related genes such as Cidea, Pgc1α, Ucp1, Prdm16, Adrb1, and Adrb3 and mitochondrial dynamics-related genes such as Mff, Opa1, and Mfn2 in brown adipose tissue (BAT). Therefore, AT extract effectively reduced obesity by promoting thermogenesis and the mitochondrial dynamics of BAT in HFD-fed mice.

Fear conditioning and extinction distinctively alter bidirectional synaptic plasticity within the amygdala of an animal model of post-traumatic stress disorder.

Synaptic plasticity in the amygdala plays an essential role in the formation and inhibition of fear memory; however, this plasticity has mainly been studied in the lateral amygdala, making it largely uninvestigated in other subnuclei. Here, we investigated long-term potentiation (LTP) and long-term depression (LTD) in the basolateral amygdala (BLA) to the medial division of the central amygdala (CEm) synapses of juvenile C57BL/6N (B6) and 129S1/SvImJ (S1) mice. We found that in naïve B6 and S1 mice, LTP was not induced at the BLA to CEm synapses, whereas fear conditioning lowered the threshold for LTP induction in these synapses of both B6 and S1 mice. Interestingly, fear extinction disrupted the induction of LTP at the BLA to CEm synapses of B6 mice, whereas LTP was left intact in S1 mice. Both low-frequency stimulation (LFS) and modest LFS (mLFS) induced LTD in naïve B6 and S1 mice, suggesting that the BLA to CEm synapses express bidirectional plasticity. Fear conditioning disrupted both types of LTD induction selectively in S1 mice and LFS-LTD, presumably NMDAR-dependent LTD was partially recovered by fear extinction. However, mLFS-LTD which has been known to be endocannabinoid receptor 1 (CB1R)-dependent was not induced after fear extinction in both mouse strains. Our observations suggest that fear conditioning enhances LTP while fear extinction diminishes LTP at the BLA to the CEm synapses of B6 mice with successful extinction. Considering that S1 mice showed strong fear conditioning and impaired extinction, strong fear conditioning in the S1 strain may be related to disrupted LTD, and impaired extinction may be due to constant LTP and weak LFS-LTD at the BLA to CEm synapses. Our study contributes to the further understanding of the dynamics of synaptic potentiation and depression between the subnuclei of the amygdala in juvenile mice after fear conditioning and extinction.

Biodegradation Potential of Polyethylene Terephthalate by the Two Insect Gut Symbionts Xanthomonas sp. HY-74 and Bacillus sp. HY-75.

Polyethylene terephthalate (PET) is a plastic material that is widely used in beverage bottles, food packaging, and other consumer products, which is highly resistant to biodegradation. In this study, we investigated the effects of two insect gut symbionts, Xanthomonas sp. HY-74 and Bacillus sp. HY-75, during PET biodegradation. Both strains degraded PET-containing agar plates, and the sole nutrition source assay showed that HY-74 had different degradation rates depending on the presence of specific carbon and nitrogen sources, whereas HY-75 exhibited comparable degradation across all tested conditions. The two strains biodegraded the PET film with 1.57 ± 0.21% and 1.42 ± 0.46% weight loss after 6 weeks, respectively. Changes in the morphology and structure of the PET films, such as erosion, scratching, and surface roughening, were determined using scanning electron microscopy (SEM). Further, the two strains biodegraded PET powder, broke it into its degradation products, and changed the surface functional groups. This is the first study to investigate the biodegradation of PET by Hymenoptera gut-derived microbes and offers promising insights into the potential applications of insect gut symbionts in PET waste management.

Assessment of nematicidal and plant growth-promoting effects of Burkholderia sp. JB-2 in root-knot nematode-infested soil.

Root-knot nematodes (RKN), Meloidogyne spp., are plant-parasitic nematodes that are responsible for considerable economic losses worldwide, because of the damage they cause to numerous plant species and the inadequate biological agents available to combat them. Therefore, developing novel and eco-friendly nematicides is necessary. In the present study, Burkholderia sp. JB-2, isolated from RKN-infested rhizosphere soil in South Korea, was evaluated to determine its nematicidal and plant growth-promoting effects under in vitro and in vivo conditions. Cell-free filtrates of the JB-2 strain showed high levels of nematicidal activity against second-stage juveniles (J2) of M. incognita, with 87.5% mortality following two days of treatment. In addition, the assessment of the activity against other six plant parasitic nematodes (M. javanica, M. hapla, M. arenaria, Ditylenchus destructor, Aphelenchoides subtenuis, and Heterodera trifolii) showed that the cell-free filtrates have a broad nematicidal spectrum. The three defense-responsive (MiMIF-2, MiDaf16-like1, and MiSkn1-like1) genes were activated, while Mi-cm-3 was downregulated when treated with cell-free filtrates of JB-2 cultures on J2. The greenhouse experiments suggested that the cell-free filtrates of the JB-2 strain efficiently controlled the nematode population in soil and egg mass formations of M. incognita in tomato (Solanum lycopersicum L., cv. Rutgers). An improvement in the host plant growth was observed, in which the shoot length and fresh weights of shoots and roots increased. The treatment with 10% of JB-2 cell-free filtrates significantly upregulated the expression levels of plant defenses (SlPR1, SlPR5, and SlPAL) and growth-promoting (ACO1, Exp18, and SlIAA1) genes compared with the corresponding parameters of the control group. Therefore, JB-2 could be a promising candidate for the sustainable management of RKN.

Enzyme Feed Additive with Arazyme Improve Growth Performance, Meat Quality, and Gut Microbiome of Pigs.

The supplementation of pig diets with exogenous enzymes is widely used with the expectation that it will improve the efficiency of nutrient utilization, thereby, improving growth performance. This study aims to evaluate the effects of a 0.1% (v/v) multi-enzyme (a mixture of arazyme (2,500,000 Unit/kg), xylanase (200,000 Unit/kg) and mannanase (200,000 Unit/kg)) supplementation derived from invertebrate symbiotic bacteria on pig performance. Here, 256 growing pigs were assigned to control and treatment groups, respectively. The treatment group exhibited a significantly reduced average slaughter age; the final body weight and average daily gain increased compared with that of the control group. In the treatment group, the longissimus muscle showed a remarkable decrease in cooking loss, shear force, and color values with increased essential and non-essential amino acid concentrations. Furthermore, the concentrations of mono- and polyunsaturated fatty acids in the treatment group increased. Feed additive supplementation increased the family of Ruminococcaceae and genera Lactobacillus, Limosilactobacillus, Turicibacter, and Oscillibacter, which play a positive role in the host physiology and health. Predicted metabolic pathway analysis confirmed that operational taxonomic units and predicted amino acid biosynthesis pathways were strongly associated. The results suggest that applying exogenous enzymes derived from invertebrate symbiotic bacteria enhances animal performance.

Stress-induced translation of KCNB1 contributes to the enhanced synaptic transmission of the lateral habenula.

The lateral habenula (LHb) is a well-established brain region involved in depressive disorders. Synaptic transmission of the LHb neurons is known to be enhanced by stress exposure; however, little is known about genetic modulators within the LHb that respond to stress. Using recently developed molecular profiling methods by phosphorylated ribosome capture, we obtained transcriptome profiles of stress responsive LHb neurons during acute physical stress. Among such genes, we found that KCNB1 (Kv2.1 channel), a delayed rectifier and voltage-gated potassium channel, exhibited increased expression following acute stress exposure. To determine the roles of KCNB1 on LHb neurons during stress, we injected short hairpin RNA (shRNA) against the kcnb1 gene to block its expression prior to stress exposure. We observed that the knockdown of KCNB1 altered the basal firing pattern of LHb neurons. Although KCNB1 blockade did not rescue despair-like behaviors in acute learned helplessness (aLH) animals, we found that KCNB1 knockdown prevented the enhancement of synaptic strength in LHb neuron after stress exposure. This study suggests that KCNB1 may contribute to shape stress responses by regulating basal firing patterns and neurotransmission intensity of LHb neurons.

Amentoflavone-Enriched Selaginella rossii Protects against Ultraviolet- and Oxidative Stress-Induced Aging in Skin Cells.

Selaginellaceae plants are used in cosmetics to limit skin aging. This study is the first to investigate the anti-aging effects of Selaginella rossii (SR) on ultraviolet B (UVB)- and oxidative stress-induced skin cells. The 95% ethanol extract of Selaginella rossii (SR95E) contained much higher amounts of amentoflavone (AMF), an active compound, than other Selaginellaceae plants and was more effective in inhibiting matrix metalloproteinase (MMP)-1 expression in CCD-986sk fibroblasts. SR95E significantly decreased UVB-induced MMP-1, MMP-2, MMP-3 and MMP-9 expression and enhanced procollagen type I C-peptide content and mRNA expression of collagen type I alpha (COL1A)1 and COL1A2 in CCD-986sk fibroblasts. In HaCaT keratinocytes, SR95E treatment also dose-dependently decreased UVB-induced MMP-1 concentration and MMP-1, MMP-2, MMP-3 and MMP-9 mRNA expression. Moreover, SR95E treatment markedly inhibited UVB-induced c-Jun N-terminal kinase and p38 mitogen-activated protein kinase signaling and nuclear factor kappa-B signaling in HaCaT cells. Furthermore, SR95E and AMF markedly regulated the 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH)-induced expression of cellular senescence-related markers, including p16, p21 and LMNB1, in HaCaT cells. Overall, this study indicates that SR may have potential as a functional material on preventing UVB- and AAPH-induced skin aging and wrinkles.

Novel, acidic, and cold-adapted glycoside hydrolase family 8 endo-ß-1,4-glucanase from an Antarctic lichen-associated bacterium, Lichenicola cladoniae PAMC 26568.

Endo-ß-1,4-glucanase is a crucial glycoside hydrolase (GH) involved in the decomposition of cellulosic materials. In this study, to discover a novel cold-adapted ß-1,4-D-glucan-degrading enzyme, the gene coding for an extracellular endo-ß-1,4-glucanase (GluL) from Lichenicola cladoniae PAMC 26568, an Antarctic lichen (Cladonia borealis)-associated bacterium, was identified and recombinantly expressed in Escherichia coli BL21. The GluL gene (1044-bp) encoded a non-modular polypeptide consisting of a single catalytic GH8 domain, which shared the highest sequence identity of 55% with that of an uncharacterized protein from Gluconacetobacter takamatsuzukensis (WP_182950054). The recombinant endo-ß-1,4-glucanase (rGluL: 38.0 kDa) most efficiently degraded sodium carboxymethylcellulose (CMC) at pH 4.0 and 45°C, and showed approximately 23% of its maximum degradation activity even at 3°C. The biocatalytic activity of rGluL was noticeably enhanced by >1.3-fold in the presence of 1 mM Mn2+ or NaCl at concentrations between 0.1 and 0.5 M, whereas the enzyme was considerably downregulated by 1 mM Hg2+ and Fe2+ together with 5 mM N-bromosuccinimide and 0.5% sodium dodecyl sulfate. rGluL is a true endo-ß-1,4-glucanase, which could preferentially decompose D-cellooligosaccharides consisting of 3 to 6 D-glucose, CMC, and barley ß-glucan, without other additional glycoside hydrolase activities. The specific activity (15.1 U mg-1) and k cat/K m value (6.35 mg-1 s-1mL) of rGluL toward barley ß-glucan were approximately 1.8- and 2.2-fold higher, respectively, compared to its specific activity (8.3 U mg-1) and k cat/K m value (2.83 mg-1 s-1mL) toward CMC. The enzymatic hydrolysis of CMC, D-cellotetraose, and D-cellohexaose yielded primarily D-cellobiose, accompanied by D-glucose, D-cellotriose, and D-cellotetraose. However, the cleavage of D-cellopentaose by rGluL resulted in the production of only D-cellobiose and D-cellotriose. The findings of the present study imply that rGluL is a novel, acidic, and cold-adapted GH8 endo-ß-1,4-glucanase with high specific activity, which can be exploited as a promising candidate in low-temperature processes including textile and food processes.

Arazyme in combination with dietary carbohydrolases influences odor emission and gut microbiome in growing-finishing pigs.

This study evaluated the effects of supplementing feed with arazyme and dietary carbohydrolases derived from invertebrate gut-associated symbionts on the noxious gas emissions, gut microbiota, and host-microbiome interactions of pigs. Here, 270 and 260 growing pigs were assigned to control and treatment groups, respectively. The tested feed additives contained a mixture of arazyme (2,500,000 Unit/kg) and synergetic enzymes, xylanase (200,000 Unit/kg) and mannanase (200,000 Unit/kg), derived from insect gut-associated symbionts in a 7.5:1:1 ratio. The control group was fed a basal diet and the treatment group was fed the basal diet supplemented with 0.1 % enzyme mixture (v/v) for 2 months. Odorous gases were monitored in ventilated air from tested houses. Fecal samples were collected from steel plate under the cage at the completion of the experiment to determine chemical composition, odor emissions, and bacterial communities. There was a significant decrease in the concentration of NH3 (22.5 vs. 11.2 ppm; P < 0.05), H2S (7.35 vs. 3.74 ppm; P < 0.05), trimethylamine (TMA) (0.066 vs. 0.001 ppm; P < 0.05), and p-cresol (0.004 ppm vs. 0 ppm; P < 0.05) at 56 d in treatment group compared with the control group. Moreover, fecal analysis results showed that exogenous enzyme supplementation caused a reduction in VFAs and indole content with approximately >60 % and 72.7 %, respectively. The result of gas emission analysis showed that NH3 (9.9 vs. 5.3 ppm; P < 0.05) and H2S (5.8 vs. 4.1 ppm; P < 0.05) were significantly reduced in the treatment group compared to the control group. The gut microbiota of the treatment group differed significantly from that of the control group, and the treatment group altered predicted metabolic pathways, including sulfur and nitrogen related metabolism, urea degradation. The results demonstrated that supplementing feed with arazyme with dietary carbohydrolases effectively controls noxious gas emissions and improves health and meat quality of pigs.

Amentoflavone-Enriched Selaginella rossii Warb. Suppresses Body Weight and Hyperglycemia by Inhibiting Intestinal Lipid Absorption in Mice Fed a High-Fat Diet.

Many Selaginellaceae species are used as traditional medicines in Asia. This study is the first to investigate the anti-obesity and anti-diabetic effects of Selaginella rossii (SR) in high-fat diet (HFD)-fed C57BL/6J mice. Seven-day oral administration of ethanol extract (100 mg/kg/day) or ethyl acetate (EtOAc) extract (50 mg/kg/day) from SR improved oral fat tolerance by inhibiting intestinal lipid absorption; 10-week long-term administration of the EtOAc extract markedly reduced HFD-induced body weight gain and hyperglycemia by reducing adipocyte hypertrophy, glucose levels, HbA1c, and plasma insulin levels. Treatment with SR extracts reduced the expression of intestinal lipid absorption-related genes, including Cd36, fatty acid-binding protein 6, ATP-binding cassette subfamily G member 8, NPC1 like intracellular cholesterol transporter 1, and ATP-binding cassette subfamily A member 1. In addition, the EtOAc extract increased the expression of protein absorption-related solute carrier family genes, including Slc15a1, Slc8a2, and Slc6a9. SR extracts reduced HFD-induced hepatic steatosis by suppressing fatty acid transport to hepatocytes and hepatic lipid accumulation. Furthermore, amentoflavone (AMF), the primary compound in SR extracts, reduced intestinal lipid absorption by inhibiting fatty acid transport in HFD-fed mice. AMF-enriched SR extracts effectively protected against HFD-induced body weight gain and hyperglycemia by inhibiting intestinal lipid absorption.

Interaction Between Glucocorticoid Receptors and FKBP5 in Regulating Neurotransmission of the Hippocampus.

FK501 binding protein 51 (FKBP5) is a stress response prolyl isomerase that inhibits the translocation of the glucocorticoid receptor (GR) heterocomplex to the nucleus. Previous studies have shown that the expression levels of FKBP5 are positively correlated with psychiatric disorders, including depression and post-traumatic stress disorder. In rodents, FKBP5 deletion in the brain leads to be resilient to stress-induced depression. The hippocampus is known to be one of the primary locations mediating stress responses in the brain by providing negative feedback signals to the hypothalamus-pituitary-adrenal gland axis. Therefore, we aimed to investigate the role of FKBP5 and its interaction with GRs in the hippocampus. We observed that FKBP5 deletion in the hippocampus resulted in a minimal change in synaptic transmission. In the hippocampus, GR activation alters the release probability in inhibitory synapses as well as the postsynaptic contribution of glutamate receptors in excitatory synapses; however, no such alterations were induced in the absence of FKBP5. FKBP5 deficiency causes insensitivity to activated GRs in the hippocampus suggesting that FKBP5 mediates synaptic changes caused by GR activation. Our study provides electrophysiological evidence of stress resilience observed in FKBP5-deficient mice.

Lateral Septum Somatostatin Neurons are Activated by Diverse Stressors.

The lateral septum (LS) is a forebrain structure that has been implicated in a wide range of behavioral and physiological responses to stress. However, the specific populations of neurons in the LS that mediate stress responses remain incompletely understood. Here, we show that neurons in the dorsal lateral septum (LSd) that express the somatostatin gene (hereafter, LSdSst neurons) are activated by diverse stressors. Retrograde tracing from LSdSst neurons revealed that these neurons are directly innervated by neurons in the locus coeruleus (LC), the primary source of norepinephrine well-known to mediate diverse stress-related functions in the brain. Consistently, we found that norepinephrine increased excitatory synaptic transmission onto LSdSst neurons, suggesting the functional connectivity between LSdSst neurons and LC noradrenergic neurons. However, optogenetic stimulation of LSdSst neurons did not affect stress-related behaviors or autonomic functions, likely owing to the functional heterogeneity within this population. Together, our findings show that LSdSst neurons are activated by diverse stressors and suggest that norepinephrine released from the LC may modulate the activity of LSdSst neurons under stressful circumstances.

Polyurethane biodegradation by Serratia sp. HY-72 isolated from the intestine of the Asian mantis Hierodula patellifera.

Polyurethane (PU), currently replacing existing synthetic materials worldwide, is a synthetic polymer derived from polyols, isocyanates, and a chain extender added by condensation reactions. PU wastes which are difficult to recycle, are commonly discarded in landfills and flow into ecosystems, thereby causing serious environmental problems. In recent years, insect-associated microbes have become a promising, eco-friendly strategy as an alternative to plastic recycling. This study aimed to evaluate the potential of Serratia sp. HY-72 strain isolated from the intestine of the Asian mantis (Hierodula patellifera) for PU degradation. The 65 kDa family I.3 lipase which degrades PU was identified and characterized, with a specific activity of 2,883 U mg-1. The bacterial filtrates and the recombinant lipase degraded Impranil (a colloidal polyester-PU dispersion, 100 g l-1) by 85.24 and 78.35% after 72 h incubation, respectively. Fourier transform infrared spectroscopy analysis revealed changes in Impranil functional groups, with decreased C=O functional group and aliphatic chain signals, and increased N-H bending with C-N stretching and C-O stretching. The current study also revealed that the HY-72 strain biodegraded the commercial PU foams (polyester- and polyether- PU) with 23.95 and 10.95% weight loss after 2 weeks, respectively with changes in surface morphology and structure such as cracks, roughness, and surface roughening. Altogether, this is one of the few studies reporting biodegradation of PU by the insect-associated microbe. These findings suggest that the insect-associated microbe could be a promising resource for biodegradation and recycling of plastic waste.

Novel Bi-Modular GH19 Chitinase with Broad pH Stability from a Fibrolytic Intestinal Symbiont of Eisenia fetida, Cellulosimicrobium funkei HY-13.

Endo-type chitinase is the principal enzyme involved in the breakdown of N-acetyl-d-glucosamine-based oligomeric and polymeric materials through hydrolysis. The gene (966-bp) encoding a novel endo-type chitinase (ChiJ), which is comprised of an N-terminal chitin-binding domain type 3 and a C-terminal catalytic glycoside hydrolase family 19 domain, was identified from a fibrolytic intestinal symbiont of the earthworm Eisenia fetida, Cellulosimicrobium funkei HY-13. The highest endochitinase activity of the recombinant enzyme (rChiJ: 30.0 kDa) toward colloidal shrimp shell chitin was found at pH 5.5 and 55 °C and was considerably stable in a wide pH range (3.5-11.0). The enzyme exhibited the highest biocatalytic activity (338.8 U/mg) toward ethylene glycol chitin, preferentially degrading chitin polymers in the following order: ethylene glycol chitin > colloidal shrimp shell chitin > colloidal crab shell chitin. The enzymatic hydrolysis of N-acetyl-ß-d-chitooligosaccharides with a degree of polymerization from two to six and colloidal shrimp shell chitin yielded primarily N,N'-diacetyl-ß-d-chitobiose together with a small amount of N-acetyl-d-glucosamine. The high chitin-degrading ability of inverting rChiJ with broad pH stability suggests that it can be exploited as a suitable biocatalyst for the preparation of N,N'-diacetyl-ß-d-chitobiose, which has been shown to alleviate metabolic dysfunction associated with type 2 diabetes.

Novel Anti-Fungal d-Laminaripentaose-Releasing Endo-ß-1,3-glucanase with a RICIN-like Domain from Cellulosimicrobium funkei HY-13.

Endo-ß-1,3-glucanase plays an essential role in the deconstruction of ß-1,3-d-glucan polysaccharides through hydrolysis. The gene (1650-bp) encoding a novel, bi-modular glycoside hydrolase family 64 (GH64) endo-ß-1,3-glucanase (GluY) with a ricin-type ß-trefoil lectin domain (RICIN)-like domain from Cellulosimicrobium funkei HY-13 was identified and biocatalytically characterized. The recombinant enzyme (rGluY: 57.5 kDa) displayed the highest degradation activity for laminarin at pH 4.5 and 40 °C, while the polysaccharide was maximally decomposed by its C-terminal truncated mutant enzyme (rGluYΔRICIN: 42.0 kDa) at pH 5.5 and 45 °C. The specific activity (26.0 U/mg) of rGluY for laminarin was 2.6-fold higher than that (9.8 U/mg) of rGluYΔRICIN for the same polysaccharide. Moreover, deleting the C-terminal RICIN domain in the intact enzyme caused a significant decrease (>60%) of its ability to degrade ß-1,3-d-glucans such as pachyman and curdlan. Biocatalytic degradation of ß-1,3-d-glucans by inverting rGluY yielded predominantly d-laminaripentaose. rGluY exhibited stronger growth inhibition against Candida albicans in a dose-dependent manner than rGluYΔRICIN. The degree of growth inhibition of C. albicans by rGluY (approximately 1.8 µM) was approximately 80% of the fungal growth. The superior anti-fungal activity of rGluY suggests that it can potentially be exploited as a supplementary agent in the food and pharmaceutical industries.

Combination Treatment of Arazyme and Soy Leaf Extract Attenuates Hyperglycemia and Hepatic Steatosis in High-Fat Diet-Fed C57BL/6J Mice.

Arazyme and extracts of soy leaves (ESLs) are used as ingredients for functional foods; however, their combined administration has not been studied. This study assessed the combined effect of Arazyme and ESLs in high-fat-diet (HFD)-induced obese C57BL/6J mice fed 2 mg/kg Arazyme, 50 mg/kg ESLs, or a combination of 2 mg/kg Arazyme and 50 mg/kg ESLs by oral gavage for 13 weeks. Individually, Arazyme and ESLs had no effect on the HFD-induced phenotypes. The combination of Arazyme and ESLs significantly suppressed body weight gain, improved glucose and insulin tolerance, and suppressed hepatic steatosis by reducing lipid synthesis and enhancing lipid utilization gene expression. Furthermore, the combination significantly reduced HFD-induced plasma bile acid reabsorption by suppressing bile acid transporter expression, including the ATP biding cassette subfamily B member 11 (Abcb11), solute carrier family 10 member 1 (Slc10a1), Slc10a2, Slc51a, and Slc51b in the liver and gut. Moreover, the combination of Arazyme and ESLs significantly prevented HFD-induced islet compensation in the pancreas. These results suggest that the incorporation of Arazyme combined with ESLs reduces HFD-induced body weight, hyperglycemia, and hepatic steatosis by regulating liver-gut bile acid circulation in HFD-fed mice. This combination can markedly reduce treatment doses and enhance their therapeutic effects, thereby reducing therapeutic healthcare costs.

Development of a novel denture care agent with highly active enzyme, arazyme.

BACKGROUND: The importance of efficient denture deposit removal and oral hygiene has been further underscored by the continuous increase of denture wearers. Denture hygiene management has also become an important aspect associated with denture-induced stomatitis. This study aims to evaluate the denture cleaning effect of arazyme, the metalloprotease produced from the Serratia proteamaculans HY-3. We performed growth inhibition tests against oral opportunistic pathogens to be used as a potential oral health care agent. METHODS: The proteolytic activities of arazyme was evaluated over broad ranges of temperature, pH, and denture components compared to those of subtilisin in commercially available denture cleansers. The washing effects of arazyme were also measured by using homogeneously soiled EMPA 105 cottons. To investigate the denture cleaning capability of arazyme, artificially contaminated dentures were treated with arazyme, subtilisin (Everlase 6.0T), and Polident®, respectively. The growth kinetics of Candida albicans, Enterococcus faecalis, Staphylococcus epidermis, and Streptococcus mutans were evaluated in the presence of different concentrations of arazyme to estimate the prevention effects of arazyme against major oral opportunistic pathogens. RESULTS: Arazyme showed strong proteolytic activities over wide temperature and pH ranges compared with the serine protease of the subtilisin family. Arazyme demonstrated efficient removal and decomposition of artificially contaminated dentures and showed explicit washing effects against soiled cottons. Moreover arazyme inhibited the growth of oral opportunistic pathogens, including C. albicans, E. faecalis, S. epidermis, and S. mutans, with more than 80% inhibition against C. albicans, the major cause of denture stomatitis, with 250 mg/mL arazyme. CONCLUSIONS: Arazyme shows promise as a biological oral health care agent with effective cleaning and antimicrobial activities and is a potential source for developing novel denture care agents.

Identification and Characterization of a Novel, Cold-Adapted d-Xylobiose- and d-Xylose-Releasing Endo-ß-1,4-xylanase from an Antarctic Soil Bacterium, Duganella sp. PAMC 27433.

Endo-ß-1,4-xylanase is a key enzyme in the degradation of ß-1,4-d-xylan polysaccharides through hydrolysis. A glycoside hydrolase family 10 (GH10) endo-ß-1,4-xylanase (XylR) from Duganella sp. PAMC 27433, an Antarctic soil bacterium, was identified and functionally characterized. The XylR gene (1122-bp) encoded an acidic protein containing a single catalytic GH10 domain that was 86% identical to that of an uncultured bacterium BLR13 endo-ß-1,4-xylanase (ACN58881). The recombinant enzyme (rXylR: 42.0 kDa) showed the highest beechwood xylan-degrading activity at pH 5.5 and 40 °C, and displayed 12% of its maximum activity even at 4 °C. rXylR was not only almost completely inhibited by 5 mM N-bromosuccinimide or metal ions (each 1 mM) including Hg2+, Ca2+, or Cu2+ but also significantly suppressed by 1 mM Ni2+, Zn2+, or Fe2+. However, its enzyme activity was upregulated (>1.4-fold) in the presence of 0.5% Triton X-100 or Tween 80. The specific activities of rXylR toward beechwood xylan, birchwood xylan, oat spelts xylan, and p-nitrophenyl-ß-d-cellobioside were 274.7, 103.2, 35.6, and 365.1 U/mg, respectively. Enzymatic hydrolysis of birchwood xylan and d-xylooligosaccharides yielded d-xylose and d-xylobiose as the end products. The results of the present study suggest that rXylR is a novel cold-adapted d-xylobiose- and d-xylose-releasing endo-ß-1,4-xylanase.

Viburnum stellato-tomentosum Extract Suppresses Obesity and Hyperglycemia through Regulation of Lipid Metabolism in High-Fat Diet-Fed Mice.

The potential biological activities of Viburnum stellato-tomentosum (VS), a plant mainly found in Costa Rica, have yet to be reported. Supplementation of VS extract for 17 weeks significantly decreased body weight gain, fat weight, fasting glucose, insulin, homeostasis model assessment of insulin resistance (HOMA-IR), and triglyceride levels in high-fat diet (HFD)-fed C57BL/6J mice. The molecular mechanisms underlying the anti-obesity and glucose-lowering effects of VS extract were investigated. VS extract suppressed adipocyte hypertrophy by regulating lipogenesis-related CCAAT/enhancer-binding protein α (C/EBPα) and insulin sensitivity-related peroxisome proliferator-activated receptor γ (Pparg) expression in adipose tissue (AT) and hepatic steatosis by inhibiting C/EBPα and lipid transport-related fatty acid binding protein 4 (FABP4) expression. VS extract enhanced muscular fatty acid ß-oxidation-related AMP-activated protein kinase (AMPK) and PPARα expression with increasing Pparg levels. Furthermore, VS extract contained a much higher content of amentoflavone (AMF) (29.4 mg/g extract) compared to that in other Viburnum species. AMF administration decreased Cebpa and Fabp4 levels in the AT and liver, as well as improved insulin signaling-related insulin receptor substrate 1 (Irs1) and glucose transporter 1 (Glut1) levels in the muscle of HFD-fed mice. This study elucidated the in vivo molecular mechanisms of AMF for the first time. Therefore, VS extract effectively diminished obesity and hyperglycemia by suppressing C/EBPα-mediated lipogenesis in the AT and liver, enhancing PPARα-mediated fatty acid ß-oxidation in muscle, and PPARγ-mediated insulin sensitivity in AT and muscle.