Synthesis of enantiopure 2-aryl-2-methoxypropionic acids and determination of their absolute configurations by X-ray crystallography

Racemic 2-aryl-2-methoxypropionic acids were enantioresolved by the use of (S)-(-)-phenylalaninol 4. For instance, racemic 2-methoxy-2-phenylpropionic acid (+/-)-7 was condensed with phenylalaninol (S)-(-)-4 yielding a diastereomeric mixture of amides, which was easily separated by HPLC on silica gel affording the first-eluted amide (-)-13a and the second-eluted amide (+)-13b: alpha = 3.19, Rs = 3.49. The absolute configuration of amide (-)-13a was determined to be (R;S) by X-ray crystallography by reference to the S configuration of the phenylalaninol moiety. Amide (R;S)-(-)-13a was converted to oxazoline (R;S)-(-)-14a, from which enantiopure 2-methoxy-2-phenylpropionic acid (R)-(-)-7 was recovered. Other 2-aryl-2-methoxypropionic acids, (R)-(-)-8, (R)-(-)-9, (R)-(+)-10, (R)-(-)-11, and (R)-(-)-12, were similarly prepared in enantiopure forms with the use of phenylalaninol (S)-(-)-4, and their absolute configurations were clearly determined by X-ray crystallography or by chemical correlation.     

New route to enantiopure MalphaNP acid, a powerful resolution and chiral 1H NMR anisotropy reagent

MalphaNP acid (+/-)-1, 2-methoxy-2-(1-naphthyl)propionic acid, was enantioresolved by the use of phenylalaninol (S)-(-)-4; a diastereomeric mixture of amides formed from acid (+/-)-1 and amine (S)-(-)-4 was easily separated by fractional recrystallization and/or HPLC on silica gel, yielding amides (R;S)-(-)-5a and (S;S)-(+)-5b. Their absolute configurations were determined by X-ray crystallography by reference to the S configuration of the phenylalaninol moiety. Amide (R;S)-(-)-5a was converted to oxazoline (R;S)-(+)-8a, from which enantiopure MalphaNP acid (R)-(-)-1 was recovered. In a similar way, enantiopure MalphaNP acid (S)-(+)-1 was obtained from amide (S;S)-(+)-5b. These reactions provide a new route for the large-scale preparation of enantiopure MalphaNP acid, a powerful chiral reagent for the enantioresolution of alcohols and simultaneous determination of their absolute configurations by (1)H NMR anisotropy.     

Novel chiral isoxazole derivatives: synthesis and pharmacological characterization at human beta-adrenergic receptor subtypes

Isoxazole derivative (+/-)-4 and the three pairs of stereoisomeric 3-bromo-isoxazolyl amino alcohols (S,R)-(-)-7a/(R,R)-(+)-7b, (S,R)-(-)-8a/(R,R)-(+)-8b, and (S,R)-(-)-9a/(R,R)-(+)-9b were synthesized and assayed for their affinity and efficacy at human beta(1)-, beta(2)-, and beta(3)-adrenergic receptors (beta-ARs) in membranes from Chinese hamster ovary (CHO) cells stably transfected with the respective receptor subtype. Whereas derivative (+/-)-4 did not bind at all three beta-ARs, stereoisomers (S,R)-7a-(S,R)-9a behaved as high-affinity ligands at beta(1)- and, particularly, at beta(2)-ARs (K(i) 2.82-66.7 nM). The K(i) values of isomers (R,R)-7b-(R,R)-9b at beta(1)- and beta(2)-subtypes were about 30-100 times higher than those of their (S,R)-7a-9a counterparts, indicating a sizable stereochemical effect. The affinity at beta(3)-ARs was negligible for all the investigated compounds. When submitted to a functional assay, the three stereoisomeric pairs showed a comparable pattern of efficacy at all three beta-AR subtypes. The highest value of efficacy (75-90%) was observed at beta(2)-ARs, whereas all compounds behaved as partial agonists (30-60%) at the beta(3)-subtype. The lowest degree of efficacy (15-35%) was found at beta(1)-ARs. The affinity/efficacy profile of the derivatives under study has been compared with that of the two model compounds, Broxaterol [(+/-)-1] and BRL 37344 [(+/-)-6].     

Microbial hydroxylation of (+/-)- and (-)-(2Z,4E)-5-(1',2'-epoxy-2',6',6'-trimethylcyclohexyl)-3-methyl-2,4-pentadienoic acid into (+/-)- and (-)-xanthoxin acid by Cunninghamella echinulata

Microbial hydroxylation of (+/-)-(2Z,4E)-5-(1',2'-epoxy-2',6',6'-trimethylcyclohexyl)-3-methyl-2,4-pentadienoic acid (3a) with Cercospora cruenta, a fungus producing (+)-abscisic acid, gave a four-stereoisomeric mixture consisting of (+)- and (-)-xanthoxin acid (4a), and (+)- and (-)-epi-xanthoxin acid (5a) by an HPLC analysis with a chiral column. Screening of the microorganisms capable of oxidizing (+/-)-3a showed that Cunninghamella echinulata stereoselectively oxidized (+/-)-3a to xanthoxin acid (4a) with the some degree of enantioselectivity as (-)-3a to (-)-4a.     

Synthesis and antipicornavirus activity of (R)- and (S)-1-[5-(4'-chlorobiphenyl-4-yloxy)-3-methylpentyl]-3-pyridin-4-yl-imidazolidin-2-one

The new pyridyl imidazolidinone derivative, 1-[5-(4'-chlorobiphenyl-4-yloxy)-3-methylpentyl]-3-pyridin-4-yl-imidazolidin-2-one (+/-)-1a, was synthesized and found to have an excellent antiviral activity against EV71 (IC50 = 0.009 microM). Therefore, both the enantiomers, (S)-(+)-1a and (R)-(-)-1a, have been prepared starting from readily available monomethyl (R)-3-methylglutarate (7) as a useful chiral building block and their antiviral activity was evaluated in a plaque reduction assay. Interestingly, we observed that the enantiomer (S)-(+)-1a was 10-fold more active against enterovirus71 (EV71) (IC50 = 0.003 microM) than the corresponding enantiomer (R)-(-)-1a (IC50 = 0.033 microM). Similar results were found against all five strains (1743, 2086, 2231, 4643, and BrCr) of EV71 tested. This demonstrated that the absolute configuration of the chiral carbon atom at the 3-position of the alkyl linker considerably influenced the anti-EV71 activity of these pyridyl imidazolidinones.     

Physiological expression of macrophage apoE in the artery wall reduces atherosclerosis in severely hyperlipidemic mice

We have previously reported that the introduction of macrophage apoE into mice lacking both apoE and the LDL receptor (apoE(-)(/-)/LDLR(-)(/-)) through bone marrow transplantation (apoE(+)(/+)/LDLR(-)(/-)-->apoE(-)(/-)/LDLR(-)(/-)) produces progressive accumulation of apoE in plasma without affecting lipid levels. This model provides a tool to study the effects of physiologically regulated amounts of macrophage apoE on atherogenesis in hyperlipidemic animals. Ten-week-old male apoE(-)(/-)/LDLR(-)(/-) mice were transplanted with either apoE(+)(/+)/LDLR(-)(/-) (n = 11) or apoE(-)(/-)/LDLR(-)(/-) (n = 14) marrow. Although there were no differences between the two groups in lipid levels at baseline or at 5 and 9 weeks after transplantation, apoE levels in the apoE(+)(/+)LDLR(-)(/-)-->apoE(-)(/-)/LDLR(-)(/-) mice increased to 4 times the apoE levels of normal mice. This resulted in a 60% decrease in aortic atherosclerosis in the apoE(+)(/+)/LDLR(-)(/-)-->apoE(-)(/-)/LDLR(-)(/-) compared with the apoE(-)(/-)/LDLR(-)(/-)-->apoE(-)(/-)/LDLR(-)(/-) controls, (15957 +/- 1907 vs. 40115 +/- 8302 micro m(2) +/- SEM, respectively). In a separate experiment, apoE(+)(/+)/LDLR(-)(/-) mice were transplanted with either apoE(+)(/+)/LDLR(-)(/-) or apoE(-)(/-)/LDLR(-)(/-) marrow and placed on a high-fat diet for 8 weeks. In the absence of macrophage apoE, lesion area was increased by 75% in the aortic sinus and by 56% in the distal aorta. These data show that physiologic levels of macrophage apoE in the vessel wall are anti-atherogenic in conditions of severe hyperlipidemia and can affect later stages of plaque development.     

Chronic hypertension in ANP knockout mice: contribution of peripheral resistance

Atrial Natriuretic Peptide (ANP) exerts a chronic hypotensive effect which is mediated by a reduction in total peripheral resistance (TPR). Mice with a homozygous disruption of the pro-ANP gene (-/-) fail to synthesize ANP and develop chronic hypertension in comparison to their normotensive wild-type (+/+) siblings. In order to determine whether alterations in basal hemodynamics underlie the hypertension associated with lack of endogenous ANP activity, we used anesthetized mice to measure arterial blood pressure (ABP) and heart rate (HR), as well as cardiac output (CO) by thermodilution technique. -/- (n = 7) and +/+ (n = 10) mice of comparable weight and age were used. Stroke volume (SV) and TPR were derived from CO, HR, and ABP by a standard formula. ABP (mm Hg) was significantly higher in -/- (132+/-4) (P < 0.0001) than in +/+ mice (95+/-2). CO (ml min(-1)), HR(beats min(-1))and SV (microl beat(-1)) did not differ significantly between -/- and +/+ mice (CO -/- = 7.3+/-0.5, +/+ = 8.3+/-0.6; HR -/- = 407+/-22, +/+ = 462+/-21; SV -/- = 17.6+/-1.1, +/+ = 17.6+/-1.7). However, TPR (mm Hg ml(-1) min(-1)) was significantly elevated in -/- mice (18.4+/-0.7) compared to +/+ mice (12.3+/-1) (P = 0.0003). Autonomic ganglion blockade with a mixture of hexamethonium and pentolinium was followed by comparable percent reductions in CO (-/- = 28+/-4, +/+ = 29+/-3), HR (-/- = 9+/-4, +/+ = 16+/-4) and SV(-/- = 21+/-4, +/+ = 15+/-6) in both genotypes. However, the concomitant decrease in ABP (%) in -/- (41+/-2) was significantly greater than in +/+ (23+/-4) mice (P = 0.0009) and was accompanied by a significant reduction in TPR. We conclude that the hypertension associated with lack of endogenous ANP is due to elevated TPR, which is determined by an increase in cardiovascular autonomic tone.     

Synthesis and stereostructure-activity relationship of three asymmetric center pyrethroids: 2-methyl-3-phenylcyclopropylmethyl 3-phenoxybenzyl ether and cyanohydrin ester

2-Methyl-3-phenylcyclopropylmethyl 3-phenoxybenzyl ether 2 and cyanohydrin ester 3, a couple of pyrethroids with three asymmetric centers, were synthesized. Of each of the four diastereomers of 2 and 3, only the (1R*,2R*,3R*)-2a and 3a showed significant insecticidal activities. Dual sets of enantiomers [(1R,2R,3R)-(-)-2a and (1S,2S,3S)-(+)-2a] and [(1R,2R,3R)-(-)-3a and (1S,2S,3S)-(+)-3a] were synthesized through the asymmetric cyclopropanation using the Aratani catalyst. Significant separations of insecticidal activities were observed between both the enantiomers against the tobacco cutworm (Spodoptera litura) and the common mosquito (Culex pipiens pallens); (1S,2S,3S)-(+)-2a and (+)-3a showed higher activities than their antipodes (1R,2R,3R)(-)-2a and (-)-3a. This result is the second example of such synthetic pyrethroids with three asymmetric centers.     

Indispensable role of Stat5a in Stat6-independent Th2 cell differentiation and allergic airway inflammation

It is well-recognized that Stat6 plays a critical role in Th2 cell differentiation and the induction of allergic inflammation. We have previously shown that Stat5a is also required for Th2 cell differentiation and allergic airway inflammation. However, it is the relative importance and redundancy of Stat6 and Stat5a in Th2 cell differentiation and allergic airway inflammation are unknown. In this study we addressed these issues by comparing Stat5a-deficient (Stat5a(-/-)) mice, Stat6(-/-) mice, and Stat5a- and Stat6 double-deficient (Stat5a(-/-) Stat6(-/-)) mice on the same genetic background. Th2 cell differentiation was severely decreased in Stat6(-/-)CD4+ T cells, but Stat6-independent Th2 cell differentiation was still significantly observed in Stat6(-/-)CD4+ T cells. However, even in the Th2-polarizing condition (IL-4 plus anti-IFN-gamma mAb), no Th2 cells developed in Stat5a(-/-)Stat6(-/-) CD4+ T cells. Moreover, Ag-induced eosinophil and lymphocyte recruitment in the airways was severely decreased in Stat5a(-/-)Stat6(-/-) mice compared with that in Stat6(-/-) mice. These results indicate that Stat5a plays an indispensable role in Stat6-independent Th2 cell differentiation and subsequent Th2 cell-mediated allergic airway inflammation.     

Redefining Escherichia coli σ(70) promoter elements: -15 motif as a complement of the -10 motif

Classical elements of σ(70) bacterial promoters include the -35 element ((-35)TTGACA(-30)), the -10 element ((-12)TATAAT(-7)), and the extended -10 element ((-15)TG(-14)). Although the -35 element, the extended -10 element, and the upstream-most base in the -10 element ((-12)T) interact with σ(70) in double-stranded DNA (dsDNA) form, the downstream bases in the -10 motif ((-11)ATAAT(-7)) are responsible for σ(70)-single-stranded DNA (ssDNA) interactions. In order to directly reflect this correspondence, an extension of the extended -10 element to a so-called -15 element ((-15)TGnT(-12)) has been recently proposed. I investigated here the sequence specificity of the proposed -15 element and its relationship to other promoter elements. I found a previously undetected significant conservation of (-13)G and a high degeneracy at (-15)T. I therefore defined the -15 element as a degenerate motif, which, together with the conserved stretch of sequence between -15 and -12, allows treating this element analogously to -35 and -10 elements. Furthermore, the strength of the -15 element inversely correlates with the strengths of the -35 element and -10 element, whereas no such complementation between other promoter elements was found. Despite the direct involvement of -15 element in σ(70)-dsDNA interactions, I found a significantly stronger tendency of this element to complement weak -10 elements that are involved in σ(70)-ssDNA interactions. This finding is in contrast to the established view, according to which the -15 element provides a sufficient number of σ(70)-dsDNA interactions, and suggests that the main parameter determining a functional promoter is the overall promoter strength.     

Possible association of BLM in decreasing DNA double strand breaks during DNA replication

Bloom's syndrome (BS) is a rare genetic disorder and the cells from BS patients show genomic instability and an increased level of sister chromatid exchange (SCE). We generated BLM(-/-) and BLM(-/-)/RAD54(-/-) DT40 cells from the chicken B-lymphocyte line DT40. The BLM(-/-) DT40 cells showed higher sensitivity to methyl methanesulfonate and elevated levels of SCE as expected. The targeted integration frequency was also increased remarkably in BLM(-/-) cells. The SCE frequency increase in BLM(-/-) cells was considerably reduced and the enhanced targeted integration observed in BLM(-/-) cells was almost completely abolished in BLM(-/-)/RAD54(-/-) cells, indicating that a large portion of the SCE in BLM(-/-) cells occurs via homologous recombination, and homologous recombination events increase with the defect of BLM function. The BLM(-/-)/RAD54(-/-) cells showed a slow growth phenotype and an increased incidence of chromosome-type breaks/gaps while each single mutant showed relatively small numbers of chromosome-type breaks/gaps.     

Leukocyte-derived hepatic lipase increases HDL and decreases en face aortic atherosclerosis in LDLr-/- mice expressing CETP

In addition to hepatic expression, cholesteryl ester transfer protein (CETP) and hepatic lipase (HL) are expressed by human macrophages. The combined actions of these proteins have profound effects on HDL structure and function. It is not known how these HDL changes influence atherosclerosis. To elucidate the role of leukocyte-derived HL on atherosclerosis in a background of CETP expression, we studied low density lipoprotein receptor-deficient mice expressing human CETP (CETPtgLDLr(-/-)) with a leukocyte-derived HL deficiency (HL(-/-) BM). HL(-/-) bone marrow (BM), CETPtgLDLr(-/-) mice were generated via bone marrow transplantation. Wild-type bone marrow was transplanted into CETPtgLDLr(-/-) mice to generate HL(+/+) BM, CETPtgLDLr(-/-) controls. The chimeras were fed a high-fat, high-cholesterol diet for 14 weeks to promote atherosclerosis. In female HL(-/-) BM, CETPtgLDLr(-/-) mice plasma HDL-cholesterol concentration during high-fat feeding was decreased 27% when compared with HL(+/+) BM, CETPtgLDLr(-/-) mice (P < 0.05), and this was associated with a 96% increase in en face aortic atherosclerosis (P < 0.05). In male CETPtgLDLr(-/-) mice, leukocyte-derived HL deficiency was associated with a 16% decrease in plasma HDL-cholesterol concentration and a 25% increase in aortic atherosclerosis. Thus, leukocyte-derived HL in CETPtgLDLr(-/-) mice has an atheroprotective role that may involve increased HDL levels.     

Tuning mesomorphic properties and handedness of chiral calamitic liquid crystals by minimal modification of the effective core

Two pairs of calamitic liquid crystalline molecules, (+)-2-[4'-(S)-sec-butoxyphenyl]-5-(4'-hexoxyphenyl)toluene ((+)-S-1) and (+)-2-(4'-hexoxyphenyl)-5-[4'-(S)-sec-butoxyphenyl]toluene ((+)-S-2), (-)-2-[4'-(R)-sec-butoxyphenyl]-5-(4'-hexoxyphenyl)toluene ((-)-R-1) and (-)-2-(4'-hexoxyphenyl)-5-[4'-(R)-sec-butoxyphenyl]toluene ((-)-R-2), have been designed and synthesized. Each of the molecules consists of a p-terphenyl core substituted with a methyl group on the middle ring, a chiral sec-butoxy tail, and an achiral n-hexoxy tail. The geometrical difference between (+)-S-1 ((-)-R-1) and (+)-S-2 ((-)-R-2) lies only in the location of the methyl group on the effective mesogenic core. Yet, such a small change in the structure gives rise to remarkable differences in mesogenic properties and handedness. Both (+)-S-1 and (-)-R-1 have an enantiotropic cholesteric phase (N*) and a monotropic twist grain boundary C* phase (TGBC*), whereas (+)-S-2 and (-)-R-2 exhibit only a monotropic N* phase. Moreover, (+)-S-1 ((-)-R-1) and (+)-S-2 ((-)-R-2) have opposite handedness in the N* phase, and (+)-S-1 and (-)-R-1 even have a helical inversion from N* to TGBC* phase through a non-helical chiral mesophase.     

Endoplasmic reticulum stress-associated cone photoreceptor degeneration in cyclic nucleotide-gated channel deficiency

Cyclic nucleotide-gated (CNG) channels play a pivotal role in phototransduction. Mutations in the cone CNG channel subunits CNGA3 and CNGB3 account for >70% of all known cases of achromatopsia. Cones degenerate in achromatopsia patients and in CNGA3(-/-) and CNGB3(-/-) mice. This work investigates the molecular basis of cone degeneration in CNG channel deficiency. As cones comprise only 2-3% of the total photoreceptor population in the wild-type mouse retina, we generated mouse lines with CNG channel deficiency on a cone-dominant background, i.e. CNGA3(-/-)/Nrl(-/-) and CNGB3(-/-)/Nrl(-/-) mice. The retinal phenotype and potential cell death pathways were examined by functional, biochemical, and immunohistochemical approaches. CNGA3(-/-)/Nrl(-/-) and CNGB3(-/-)/Nrl(-/-) mice showed impaired cone function, opsin mislocalization, and cone degeneration similar to that in the single knock-out mice. The endoplasmic reticulum stress marker proteins, including Grp78/Bip, phospho-eIF2α, phospho-IP(3)R, and CCAAT/enhancer-binding protein homologous protein, were elevated significantly in CNGA3(-/-)/Nrl(-/-) and CNGB3(-/-)/Nrl(-/-) retinas, compared with the age-matched (postnatal 30 days) Nrl(-/-) controls. Along with these, up-regulation of the cysteine protease calpains and cleavage of caspase-12 and caspase-7 were found in the channel-deficient retinas, suggesting an endoplasmic reticulum stress-associated apoptosis. In addition, we observed a nuclear translocation of apoptosis-inducing factor (AIF) and endonuclease G in CNGA3(-/-)/Nrl(-/-) and CNGB3(-/-)/Nrl(-/-) retinas, implying a mitochondrial insult in the endoplasmic reticulum stress-activated cell death process. Taken together, our findings suggest a crucial role of endoplasmic reticulum stress in cone degeneration associated with CNG channel deficiency.     

Functional relation among RecQ family helicases RecQL1, RecQL5, and BLM in cell growth and sister chromatid exchange formation

Human RECQL1 and RECQL5 belong to the RecQ family that includes Bloom syndrome, Werner syndrome, and Rothmund-Thomson syndrome causative genes. Cells derived from individuals suffering from these syndromes show significant levels of genomic instability. However, neither RECQL1 nor RECQL5 has been related to a disease, and nothing is known about the functions of RecQL1 and RecQL5. We generated here RECQL1(-/-), RECQL5(-/-), RECQL1(-/-)/RECQL5(-/-), RECQL1(-/-)/BLM(-/-), and RECQL5(-/-)/BLM(-/-) cells from chicken B-lymphocyte line DT40 cells. Although BLM(-/-) DT40 cells showed a slow-growth phenotype, a higher sensitivity to methyl methanesulfonate than the wild type, and an approximately 10-fold increase in the frequency of sister chromatid exchange (SCE) compared to wild-type cells, RECQL1(-/-), RECQL5(-/-), and RECQL1(-/-)/RECQL5(-/-) cells showed no significant difference from the wild-type cells in growth, sensitivity to DNA-damaging agents, and the frequency of SCE. However, both RECQL1(-/-)/BLM(-/-) and RECQL5(-/-)/BLM(-/-) cells grew more slowly than BLM(-/-) cells because of the increase in the population of dead cells, indicating that RecQL1 and RecQL5 are somehow involved in cell viability under the BLM function-impaired condition. Surprisingly, RECQL5(-/-)/BLM(-/-) cells showed a higher frequency of SCE than BLM(-/-) cells, indicating that RecQL5 suppresses SCE under the BLM function-impaired condition.     

Preparation of optically active 2-aminobutanoic acid via optical resolution by replacing crystallization

An attempt was made to use a simple procedure to obtain (R)- and (S)-2-aminobutanoic acids [(R)- and (S)-1] which are non-proteinogenic alpha-amino acids and are useful as chiral reagents in asymmetric syntheses. Compound (RS)-1 p-toluenesulfonate [(RS)-2], which is known to exist as a conglomerate, was optically resolved by replacing crystallization with (R)- and (S)-methionine p-toluenesulfonate [(R)- and (S)-3] as optically active co-solutes. When (S)-3 was employed as the co-solute, (R)-2 was preferentially crystallized from a supersaturated solution of (RS)-2 in 1-propanol, as was (S)-2 in the presence of (R)-3. (R)- and (S)-2 recrystallized from 1-propanol were treated with triethylamine in methanol to give (R)- and (S)-1 in optically pure forms.     

Longidorus ferrisi n. sp. from California Citrus

In October 1999, the authors received fixed specimens of a species of Longidorus from Howard Ferris found about the roots of a citrus tree in Oakville, Napa County, CA. After determining it to be new a species, we requested additional specimens. The samples contained roughly equal numbers of males and females. Longidorus ferrisi n. sp. is most similar to L. elongatus, but can be distinguished by a greater c-ratio (111-187 vs 73-141), a lesser c′ (0.7-1.1 vs 1.0-1.3), a more offset head, a more posterior guide ring (35-40 vs 30-33 μm), the presence of sperm in the uterus in mature females, and the approximate 1:1 ratio of females to males. Other similar species include L. artemisiae, L. crassus, L. glycines, and L. milanis. Longidorus ferrisi n. sp. differs from L. artemisiae by a lesser a-ratio (74-102 vs 109-155), a lesser c′ value (0.7-1.1 vs 1.0-1.6), a more posterior guide ring (35-40 vs 27-34 μm), a longer odontostyle (91-108 vs 84-98 μm), a wider lip region (16-19 vs 14-17 μm), wider mid-body (53-69 vs 41-52 μm), and longer spicules (57-65 vs 39-49 μm). The new species differs substantially from L. crassus by its lip shape and the presence of males, and differs from L. glycines by a shorter body (4.33-5.97 vs 6.14-8.31 mm), a lesser c′ value (0.7-1.1 vs 0.9-1.4), a narrower lip region (16-19 vs 20-23 μm), wider mid-body (53-69 vs 39-57 μm), longer spicules (53-69 vs 45-53 μm), and fewer supplements (7-11 vs 11-17). Longidorus ferrisi n. sp. differs from L. milanis by a longer body (4.33-5.97vs 3.00-4.90 mm), a greater c value (111-187 vs 86-130), a wider mid-body (53-69 vs 43-56 μm), a different head shape, and longer spicules (53-69 vs 41-54 μm). The nuclear 18S ribosomal DNA sequence of this species revealed that this species is unique with respect to all sequenced Longidorus species.     

Use of wastewater as a nutrient solution in a closed gravel hydroponic culture of giant reed (Arundo donax)

The objective of this experimental work was to examine the efficiency of giant reed (Arundo donax L.), as a source of biomass production and as a biofiltering device for sewage effluents. Two giant reed populations were cultivated in a closed gravel hydroponic system, where pig's waste was used as a nutrient solution. The results showed that stem biomass production varied from 12 to 23 kg DM m(-2) yr(-1), more than the ordinary production in the soil. According to stem analysis, for the first two years, there was an average infiltration rate of 31 g m(-2) yr(-1) total N, 7.5 g m(-2) yr(-1) total P, 18.8 g m(-2) yr(-1) K, 2.1 g m(-2) yr(-1) Ca, 2.1 g m(-2) yr(-1) Mg, 0.27 g m(-2) yr(-1) Fe, 0.02 g m(-2) yr(-1) Mn, 0.14 g m(-2) yr(-1) Zn and 0.08 g m(-2) yr(-1) Cu. During the third year, when a nutrient solution with added P was used, the average infiltration rate for most elements increased by 46% and for P by 169%.     

Mitochondrial reactive oxygen production is dependent on the aromatic hydrocarbon receptor

2,3,7,8-Tetrachlorodibenzo-p-dioxin (dioxin; TCDD) is a pervasive environmental contaminant that induces hepatic and extrahepatic oxidative stress. We have previously shown that dioxin increases mitochondrial respiration-dependent reactive oxygen production. In the present study we examined the dependence of mitochondrial reactive oxygen production on the aromatic hydrocarbon receptor (AHR), cytochrome P450 1A1 (CYP1A1), and cytochrome P450 1A2 (CYP1A2), proteins believed to be important in dioxin-induced liver toxicity. Congenic Ahr(-/-), Cyp1a1(-/-) and Cyp1a2(-/-) knockout mice, and C57BL/6J inbred mice as their Ahr/Cyp1a1/Cyp1a2(+/+) wild-type (wt) counterparts, were injected intraperitoneally with dioxin (15 microg/kg body weight) or corn-oil vehicle on 3 consecutive days. Liver mitochondria were examined 1 week following the first treatment. The level of mitochondrial H(2)O(2) production in vehicle-treated Ahr(-/-) mice was one fifth that found in vehicle-treated wt mice. Whereas dioxin caused a rise in succinate-stimulated mitochondrial H(2)O(2) production in the wt, Cyp1a1(-/-), and Cyp1a2(-/-) mice, this increase did not occur with the Ahr(-/-) knockout. The lack of H(2)O(2) production in Ahr(-/-) mice was not due to low levels of Mn(2+)-superoxide dismutase (SOD2) as shown by Western immunoblot analysis, nor was it due to high levels of mitochondrial glutathione peroxidase (GPX1) activity. Dioxin decreased mitochondrial aconitase (an enzyme inactivated by superoxide) by 44% in wt mice, by 26% in Cyp1a2(-/-) mice, and by 24% in Cyp1a1(-/-) mice; no change was observed in Ahr(-/-) mice. Dioxin treatment increased mitochondrial glutathione levels in the wt, Cyp1a1(-/-), and Cyp1a2(-/-) mice, but not in Ahr(-/-) mice. These results suggest that both constitutive and dioxin-induced mitochondrial reactive oxygen production is associated with a function of the AHR, and these effects are independent of either CYP1A1 or CYP1A2.