Aldose reductase inhibitory compounds from extracts of Dipsacus asper

Amethanolic extract of Dipsacus asper, having anti-diabetic activity, was examined as a possible aldose reductase (ALR2) inhibitor, a key enzyme involved in diabetic complications. Bioactivity guided fractionation led to the isolation of ten compounds, ursolic acid (1), oleanolic acid-3-O-α-L-arabinopyranoside (2), daucosterol (3), hederagenin-3-O-α-L-arabinopyranoside (4), sweroside(5), caffeic acid (6), esculetin (7), protocatechualdehyde (8), loganin (9), and vanilic acid (10) from the ethyl acetate fraction of D. asper methanol extract. Among them, compounds 4, 6, 7, and 8 exhibited inhibitory effects on aldose reductase, with IC₅₀ values of 23.70, 16.71, 34.36, and 21.81 μM, respectively. This is the first report on the isolation of these compounds from D. asper, and the ALR2 inhibitory activity of hederagenin-3-O-α-L-arabinopyranoside. These results suggest the successful use of the extract of D. asper for ameliorating diabetic complications.     

Combination of 1,4-naphthoquinone with benzothiazoles had selective algicidal effects against harmful algae

A series of naphthoquinone-benzothiazole conjugates were synthesized as algicides, and their efficacies against harmful algal blooming species, such as Chattonella marina, Heterosigma akashiwo and Cochlodinium polykrikoides, were examined. The introduction of substituted benzothiazole at the C2 position of 1,4-naphthoquinone (compounds 1–9) resulted in higher algicidal activity against C. polykrikoides than the C6 conjugates (compounds 10–20). On the other hand, of the C6 conjugates, compounds 11 and 12 exhibited better algicidal activity against H. akashiwo, C. marina, and C. polykrikoides than the C2 conjugates. Further structure-activity analysis indicated that a replacement of the methoxy groups with hydroxyl groups (compounds 21–26) decreased the algicidal activity significantly. Among the various synthetic naphthoquinonebezothiazole conjugates tested, compound 12 was found to affect the most significant decrease in the level of C. polykrikoides growth, with an IC₅₀ of 0.19 μM. Compound 11 was found to be the most potent inhibitor against H. akashiwo and C. polykrikoides, with IC₅₀ values of 0.32 and 0.12 μM, respectively. Overall, these results highlight a possible method for controlling and inhibiting red tide forming algae using NQ derivatives.     

Enhanced production of biomass and lipids by supplying CO2 in marine microalga Dunaliella sp.

Non-food-based biofuel feedstocks are in high demand worldwide. Among the various feedstocks, microalgae are the most promising feedstock for mitigating atmospheric CO₂ and producing biodiesel. In this study, various concentrations of CO₂, from 0.03 to 12%, were used to investigate their effect on the cell growth, biomass and lipid production and fatty acid composition of Dunaliella sp. in a closed photobioreactor. The results showed that the highest biomass and total lipids, 521 mg/L/d and 40 mg/L/d, respectively, were produced with 5% CO₂ aeration during the logarithmic growth phase. The oleic acid (18:1n9c) and elaidic acid (18:1n9t) contents were increased approximately two fold. The physiological responses of Dunaliella sp. at 10% CO₂ were similar to those at 5% CO₂. Therefore, the present results suggest that 5–10% is a suitable CO₂ concentration range for Dunaliella sp. growth to mitigate atmospheric CO₂ and increase biofuel production.     

Enzyme-linked assay of cellulose-binding domain functions from Cellulomonas fimi on multi-well microtiter plate

Cellulose-binding domain (CBD) enriches cellulolytic enzymes on cellulosic surfaces and contributes to the catalytic efficiency by increasing enzyme-substrate complex formations. Thus, high affinity CBDs are essential for the development of efficient cellulose-degrading enzymes. Here, we present a microtiter plate-based assay system to measure the binding affinity of CBDs to cellulose. The assay uses a periplasmic alkaline phosphatase (AP) as a fusion reporter and its activity is detected using a fluorogenic substrate, 4-methylumbelliferyl phosphate. Lignocellulose discs of 6 mm in diameter were used as substrates in 96-well plate. As a result, the enzyme-linked assay detected the binding of CBDs on the cellulosic discs in a highly sensitive manner, detecting from 0.05 to 1.0 μg/mL of APCBD proteins, which is several hundred times more sensitive than conventional protein measurements. The proposed method was applied to compare the binding affinity of different CBDs from Cellulomonas fimi to lignocellulose discs.     

Capture of Bactrocera fruit flies in traps baited with liquid versus solid formulations of male lures in Malaysia

Approximately 70 species of Bactrocera fruit flies (Diptera: Tephritidae) are polyphagous economic pests that attack many important agricultural crops. Several of these Bactrocera species are also highly invasive, and many countries operate continuous, large-scale trapping programs to detect incipient infestations. Detection programs rely heavily on traps baited with male lures, with males of some species responding to raspberry ketone (RK; or its synthetic analogue cue-lure [CL]) and males of other species responding to methyl eugenol (ME). These lures (plus naled, an insecticide) are currently applied as liquids, although this procedure is time-consuming and may expose workers to health risks. Recent field tests, conducted largely in Hawaii, have shown that traps baited with a solid formulation (termed a wafer) that contains both RK and ME (plus dichlorvos, an insecticide) capture as many or more B. dorsalis (Hendel) and B. cucurbitae (Coquillett) males as traps baited with the standard liquid lures. While these results are promising, a more complete evaluation of the solid formulation requires testing in a region with a diverse assemblage of Bactrocera species, since interspecific variation in male response to lures has been reported. The objective of the present investigation was to assess the relative effectiveness of liquid versus solid formulations of male lures in Malaysia, a country known to harbor a large assemblage of Bactrocera species. Based on a 12-week sampling period, we found that, contrary to the Hawaiian results, traps baited with the wafer captured significantly fewer males than traps baited with liquid lures for all five ME-responding taxa analyzed and for one of the three RK/CL-responding species analyzed. Possible explanations for the discrepancy between these and earlier findings are offered.     

Resolvin D1 inhibits TGF-β1-induced epithelial mesenchymal transition of A549 lung cancer cells via lipoxin A4 receptor/formyl peptide receptor 2 and GPR32

Epithelial-mesenchymal-transition (EMT) is a key event for tumor cells to initiate metastasis which lead to switching of E-cadherin to N-cadherin. Resolvins are known to promote the resolution of inflammation and phagocytosis of macrophages. However, the role of resolvins in EMT of cancer is not known. Therefore, we examined the effects of resolvins on transforming growth factor, beta 1 (TGF-β1)-induced EMT. Expression of E-cadherin and N-cadherin in A549 lung cancer cells was evaluated by Western blot and confocal microscopy. Involvement of lipoxin A4 receptor/formyl peptide receptor 2 (ALX/FPR2) was examined by gene silencing. TGF-β1 induced expression of N-cadherin in A549 lung cancer cells, and resolvin D1 and D2 inhibited the expression of N-cadherin at low concentrations (1–100 nM). Resolvin D1 and D2 also suppressed the expression of zinc finger E-box binding homeobox 1 (ZEB1). The effects of resolvin D1 and D2 were confirmed in other lung cancer cell lines such as H838, H1299, and H1703. Resolvin D1 and D2 did not affect the proliferation of A549 lung cancer cells. Resolvin D1 and D2 also suppressed the TGF-β1-induced morphological change. Resolvin D1 and D2 also inhibited the TGF-β1-induced migration and invasion of A549 cells. Resolvin D1 is known to act via ALX/FPR2 and GPR32. Thus, we examined the involvement of ALX/FPR2 and GPR32 in the suppressive effects of resolvin D1 on TGF-β1-induced EMT of A549 cells. Gene silencing of ALX/FPR2 and GPR32 blocked the action of resolvin D1. Overexpression of ALX/FPR2 or GPR32 increased the effects of resolvin D1. These results suggest that resolvin D1 inhibited TGF-β1-induced EMT via ALX/FPR2 and GPR32 by reducing the expression of ZEB1.     

Delphinidin prevents hypoxia-induced mouse embryonic stem cell apoptosis through reduction of intracellular reactive oxygen species-mediated activation of JNK and NF-κB, and Akt inhibition

Delphinidin, gallic acid, betulinic acid, and ursolic acid, which are bio-active ingredients in a variety of fruits, vegetables, and herbs, have potent antioxidant activity and various biological activities. However, it is not clear whether these bio-active ingredients can significantly contribute to the protection of embryonic stem (ES) cells from hypoxia-induced apoptosis. In the present study, hypoxia-induced ES cells apoptosis with time, which were abrogated by pretreatment with all ingredients. Hypoxia-induced ROS generation was blocked by pretreatment with all ingredients in a dose-dependent manner, with the maximum ROS scavenging effect observed for delphinidin. Hypoxia increased phosphorylation of JNK and NF-κB were blocked by pretreatment of delphinidin as well as NAC. Hypoxia decreased phosphorylation of Akt^thr308 and ^ser473; these decreases were reversed by pretreatment with delphinidin or NAC. However, Akt inhibition did not affect NF-κB phosphorylation. Delphinidin attenuated the hypoxia-induced increase in Bax, cleaved caspase-9, cleaved caspase-3, and decrease in Bcl-2, which were diminished by pretreatment of Akt inhibitor. Hypoxia induced Bax translocation from the cytosol to mitochondria. Furthermore, hypoxia induced mitochondria membrane potential loss and cytochrome c release in cytosol, which were blocked by delphinidin pretreatment. Hypoxia induced cleavage of procaspase-9 and procaspase-3 which were blocked by delphinidin or SP600125, but Akt inhibitor abolished the protection effect of delphinidin. Moreover, inhibition of JNK and NF-κB abolished hypoxia-induced ES cell apoptosis and inhibition of Akt attenuated delphinidin-induced blockage of apoptosis. The results indicate that delphinidin can prevent hypoxia-induced apoptosis of ES cells through the inhibition of JNK and NF-κB phosphorylation, and restoration of Akt phosphorylation.     

Lake restoration by biomanipulation using piscivore and Daphnia stocking; results of the biomanipulation in Japan

Biomanipulation has been employed in numerous locations throughout the world as a means for reducing phytoplankton biomass; however, it has not been employed very often in Japan. A common approach involves the introduction of piscivorous fish to reduce the abundance of planktivorous fish. In our study, to first apply biomanipulation, we stocked Lake Shirakaba (a high-altitude, protected area in a park) in central Japan with rainbow trout fingerlings and cladoceran Daphnia (Daphnia galeata) in 2000. A “pre-biomanipulation” data set (1997–1999) and “a post-biomanipulation” data set (2000–2006) allowed us to evaluate the lake's response to biomanipulation. After the biomanipulation, zoo-planktivorous pond smelt disappeared and a large population of Daphnia had been established, which substantially reduced the number of the previously dominant small cladocerans and rotifers. Water transparency increased from about 2 m (before biomanipulation) to more than 4 m (after biomanipulation). Reductions in algal biomass and increased transparency led to expansion of the submerged macrophyte Elodea nuttallii. Total phosphorus concentrations declined as well over this time period. Based on these results, we concluded that biomanipulation using piscivore and Daphnia stocking succeeded in improving lake water quality by reducing algal abundance and providing favorable conditions for the establishment of rooted plants.     

Loss of VHL promotes progerin expression,leading to impaired p14/ARF function and suppression of p53 activity

Renal cell carcinomas (RCCs) are frequently occurring genitourinary malignancies in the aged population. A morphological characteristic of RCCs is an irregular nuclear shape, which is used to index cancer grades. Other features of RCCs include the genetic inactivation of the von Hippel-Lindau gene, VHL, and p53 genetic-independent inactivation. An aberrant nuclear shape or p53 suppression has not yet been demonstrated. We examined the effect of progerin (an altered splicing product of the LMNA gene linked to Hutchinson Gilford progeria syndrome; HGPS) on the nuclear deformation of RCCs in comparison to that of HGPS cells. In this study, we showed that progerin was suppressed by pVHL and was responsible for nuclear irregularities as well as p53 inactivation. Thus, progerin suppression can ameliorate nuclear abnormalities and reactivate p53 in response to genotoxic addition. Furthermore, we found that progerin was a target of pVHL E3 ligase and suppressed p53 activity by p14/ARF inhibition. Our findings indicate that the elevated expression of progerin in RCCs results from the loss of pVHL and leads to p53 inactivation through p14/ARF suppression. Interestingly, we showed that progerin was expressed in human leukemia and primary cell lines, raising the possibility that the expression of this LMNA variant may be a common event in age-related cancer progression.     

Cyclin-dependent kinase 4 signaling acts as a molecular switch between syngenic differentiation and neural transdifferentiation in human mesenchymal stem cells

Multipotent mesenchymal stem/stromal cells (MSCs) are capable of differentiating into a variety of cell types from different germ layers. However, the molecular and biochemical mechanisms underlying the transdifferentiation of MSCs into specific cell types still need to be elucidated. In this study, we unexpectedly found that treatment of human adipose- and bone marrow-derived MSCs with cyclin-dependent kinase (CDK) inhibitor, in particular CDK4 inhibitor, selectively led to transdifferentiation into neural cells with a high frequency. Specifically, targeted inhibition of CDK4 expression using recombinant adenovial shRNA induced the neural transdifferentiation of human MSCs. However, the inhibition of CDK4 activity attenuated the syngenic differentiation of human adipose-derived MSCs. Importantly, the forced regulation of CDK4 activity showed reciprocal reversibility between neural differentiation and dedifferentiation of human MSCs. Together, these results provide novel molecular evidence underlying the neural transdifferentiation of human MSCs; in addition, CDK4 signaling appears to act as a molecular switch from syngenic differentiation to neural transdifferentiation of human MSCs.