Prohibitin involvement in the generation of mitochondrial superoxide at complex I in human sperm

Prohibitin (PHB), a major mitochondrial membrane protein, has been shown earlier in our laboratoryto regulate sperm motility via an alteration in mitochondrial membrane potential (MMP) in infertile men with poor sperm quality. To test if PHB expression is associated with sperm mitochondrial superoxide (mROS) levels, here we examined sperm mROS levels, high MMP and lipid peroxidation in infertile men with poor sperm motility (asthenospermia, A) and/or low sperm concentrations (oligoasthenospermia, OA). The diaphorase‐type activity of sperm mitochondrial complex I (MCI) and PHB expression were also determined. We demonstrate that mROS and lipid peroxidation levels are significantly higher in sperm from A and OA subjects than in normospermic subjects, whereas high MMP and PHB expression are significantly lower. A positive correlation between mROS and lipid peroxidation and a negative correlation of mROS with PHB expression, high MMP, and sperm motility were found in these subjects. The finding of similar diaphorase‐type activity levels of sperm MCI in the three groups studied suggests that the catalytic subunits of MCI in the matrix arm may produce mROS on its own. There may be a dysfunction of electron transport at MCI associated with decreased expression of PHB in sperm with poor quality. We conclude that mROS level is increased and associated with decreased PHB expression, and it may regulate sperm motility via increases in low MMP and lipid peroxidation. This is the first report on the involvement of PHB in human sperm motility loss associated with increased generation of mROS at MCI.     

MIXOTROPHY AND NITROGEN UPTAKE BY PFIESTERIA PISCICIDA (DINOPHYCEAE)

The nutritional versatility of dinoflagellates is a complicating factor in identifying potential links between nutrient enrichment and the proliferation of harmful algal blooms. For example, although dinoflagellates associated with harmful algal blooms (e.g. red tides) are generally considered to be phototrophic and use inorganic nutrients such as nitrate or phosphate, many of these species also have pronounced heterotrophic capabilities either as osmotrophs or phagotrophs. Recently, the widespread occurrence of the heterotrophic toxic dinoflagellate, Pfiesteria piscicida Steidinger et Burkholder, has been documented in turbid estuarine waters. Pfiesteria piscicida has a relatively proficient grazing ability, but also has an ability to function as a phototroph by acquiring chloroplasts from algal prey, a process termed kleptoplastidy. We tested the ability of kleptoplastidic P. piscicida to take up ¹⁵N-labeled NH     , NO     , urea, or glutamate. The photosynthetic activity of these cultures was verified, in part, by use of the fluorochrome, primulin, which indicated a positive relationship between photosynthetic starch production and growth irradiance. All four N substrates were taken up by P. piscicida , and the highest uptake rates were in the range cited for phytoplankton and were similar to N uptake estimates for phagotrophic P. piscicida . The demonstration of direct nutrient acquisition by kleptoplastidic P. piscicida suggests that the response of the dinoflagellate to nutrient enrichment is complex, and that the specific pathway of nutrient stimulation (e.g. indirect stimulation through enhancement of phytoplankton prey abundance vs. direct stimulation by saprotrophic nutrient uptake) may depend on P. piscicida 's nutritional state (phagotrophy vs. phototrophy).     

Androgen-induced alterations in vocalizations of femaleXenopus laevis: modifiability and constraints

Summary We examined effects of exogenous androgen (testosterone and dihydrotestosterone) on vocalizations of ovariectomized, adult female South African clawed frogs,Xenopus laevis. When paired with sexually active males, all ovariectomized females exhibited ticking, the unreceptive or release call. Ticking consists of low amplitude, regularly spaced clicks with a mean interclick interval of 154 ms. When androgen-treated and paired with sexually active males, these ovariectomized females also exhibited an aberrant call (atypical ticking) in which click multiples replaced the single clicks of ticking. Mean ICI's for atypical ticking were 37 ms for click doublets and 22 ms for click quadruplets. Androgen treatment decreased the total time spent vocalizing (typical and atypical ticking) by ovariectomized females.All androgen-treated females were then tested repeatedly with sexually receptive females in an attempt to elicit the male-typical vocalization, mate calling. Six of 17 females did not vocalize at all, even when gonadotropin injected. Eight females gave rapid (mean ICI, 36 ms) trains of clicks in an irregular temporal pattern (tick-like calls). Three females gave brief trills with alternating fast and slow components. Comparison of mate calllike vocalizations of androgen-treated females to mate calling of males reveals that calls in females are considerably shorter in duration (female: 0.32 min versus male: 45 min) and slower in tempo (ICI's; fast trill, female: 21 ms, male: 14 ms; slow trill, female: 36 ms, male: 28 ms). Incomplete masculinization of the vocal pattern of females by androgen treatment in adulthood may be due to developmental constraints on the modifiability of the neurons and muscles responsible for calling.Abbreviations C cholesterol - DHT dihydrotestosterone - HCG human chorionic gonadotropin - IBI interburst interval - ICI interclick interval - ovx ovariectomized - T testosterone     

Hexavalent chromium removal in vitro and from industrial wastes, using chromate-resistant strains of filamentous fungi indigenous to contaminated wastes

Two chromate-resistant filamentous fungi, strains H13 and Ed8, were selected from seven independent fungal isolates indigenous to Cr(VI)-contaminated soil because of their ability to decrease hexavalent chromium levels in the growth medium. Morphophysiological studies identified strain H13 as a Penicillium sp. isolate and Ed8 as an Aspergillus sp. isolate. When incubated in minimal medium with glucose as a carbon source and in the presence of 50 microg/mL Cr(VI), these strains caused complete disappearance of Cr(VI) in the growth medium after about 72 h of incubation. Total chromium concentration in growth medium was constant during culture growth, and no accumulation of chromium in fungal biomass was observed. Quantitative determinations of oxidized and reduced chromium species during the reduction process revealed stoichiometric conversion of Cr(VI) to Cr(III). A decrease in Cr(VI) levels from industrial wastes was also induced by Ed8 or H13 biomass. These results indicate that chromate-resistant filamentous fungi with Cr(VI)-reducing capability could be useful for the removal of Cr(VI) contamination.     

Non-random sampling and its role in habitat conservation: a comparison of three wetland macrophyte sampling protocols

Aquatic macrophytes provide essential spawning and nursery habitat for fish, valuable food source for waterfowl, migratory birds and mammals, and contribute greatly to overall biodiversity of coastal marshes of the Laurentian Great Lakes. Two approaches have been used to survey the plant community in coastal wetlands, and these include the grid (GR) and transect (TR) methods. These methods have been used to identify the average species richness at different sites, but their suitability for determining total species richness of a site has not been tested. In this paper, we compare the performance of these two established methods with that of the Stratified method (ST), which uses the sampler’s judgment to guide them to different habitat zones within the wetland. We used the three protocols to compare species richness of six coastal wetlands of the Great Lakes, three pristine marshes in eastern Georgian Bay (Lake Huron) and three degraded wetlands in Lake Ontario, Canada. The greatest species richness was associated with the ST method, irrespective of wetland quality. The ST method was also more efficient (fewer quadrats sampled), and revealed the most number of unique (those found with only one method) and uncommon species (those found in <5% of the quadrats). Despite these statistical differences, we found that sampling method did not significantly affect the performance of a recently developed index of wetland quality, the Wetland Macrophyte Index. These results have important implications for designing macrophyte surveys to track changes in biodiversity and wetland quality.     

Two early mississippian seeds from the price formation of Southwestern Virginia

Lagenospermum imparirameum Arnold, originally described from a few specimens of cupulate seeds borne on two or three times dichotomous branches, is now shown to be borne on more complex branching systems. Details of the cupule and seed morphology are added and an emended diagnosis of the taxon is given. A new species,Gnetopsis hispida, is described as the third occurrence of this genus and the first occurrence in beds of Lower Mississippian Age in North America. The classification, evolutionary implications, and dispersal biology are discussed for each of the seeds     

Differences in induction of hepatic cytochrome p450 isozymes by mice in eight methylenedioxyphenyl compounds

Eight methylenedioxyphenyl (MDP) compounds were examined for their ability to induce cytochrome P450 (P450) in mouse liver. Induction by safrole, isosafrole, and dihydrosafrole was studied in both C57BL/6N (Ah-responsive) and DBA/2N (Ahnonresponsive) male mice after IP administration of 200 mg/kg/day MDP compound for 3 days. Hepatic P450 content, ethylmorphine N-demethylase, ethoxy-resorufin O-deethylase, and acetanilide hydroxylase activities were induced to the same extent in both strains of mice. Benzo(a)pyrene hydroxylase activity, however, was not induced in either C57 or DBA mice. The similarity of results in both strains of mice indicated induction of these P450 isozymes by these three MDP compounds is not mediated by the Ah receptor. Induction of P450 by butylbenzodioxole (n-butyl-BD), tertiarybutylbenzodioxole (t-butyl-BD), methylbenzodioxole (methyl-BD), nitrobenzodioxole (nitro-BD), and bromobenzodioxole (bromo-BD) was examined only in C57BL/6N mice. Methyl-BD, nitro-BD, and bromo-BD did not induce hepatic microsomal proteins or selected P450 monooxygenase activities. In contrast, n-butyl-BD, and t-butyl-BD induced P450 content, ethylmorphine N-demethylase, acetanilide hydroxylase, and ethoxyresorufin O-deethylase activities. Benzo(a)pyrene hydroxylase was not induced by any of the treatments. Induction of these P450 activities is consistent with induction of P450 IIB1 and P450 IA2, but not induction of P450 IA1. Western blot analysis with antibodies to P450 isozymes induced with either phenobarbital (Pb) or 3-methylcholanthrene (3-MC) confirmed that both IIB1 and IA2 were induced, but that IA1 was not induced.     

Rendomab B4, a monoclonal antibody that discriminates the human endothelin B receptor of melanoma cells and inhibits their migration

Metastatic melanoma is an aggressive cancer with a poor prognostic, and the design of new targeted drugs to treat melanoma is a therapeutic challenge. A promising approach is to produce monoclonal antibodies (mAbs) against the endothelin B receptor (ETB), which is known to be overexpressed in melanoma and to contribute to proliferation, migration and vasculogenic mimicry associated with invasiveness of this cancer.

We previously described rendomab-B1, a mAb produced by DNA immunization. It is endowed with remarkable characteristics in term of affinity, specificity and antagonist properties against human ETB expressed by the endothelial cells, but, surprisingly, had poor affinity for ETB expressed by melanoma cells. This characteristic strongly suggested the existence of a tumor-specific ETB form. In the study reported here, we identified a new mAb, rendomab-B4, which, in contrast to rendomab-B1, binds ETB expressed on UACC-257, WM-266-4 and SLM8 melanoma cells. Moreover, after binding to UACC-257 cells, rendomab-B4 is internalized and colocalizes with the endosomal protein EEA-1. Interestingly, rendomab-B4, despite its inability to compete with endothelin binding, is able to inhibit phospholipase C pathway and migration induced by endothelin. By contrast, rendomab-B4 fails to decrease ERK1/2 phosphorylation induced by endothelin, suggesting a biased effect on ETB.

These particular properties make rendomab-B4 an interesting tool to analyze ETB-structure/function and a promising starting point for the development of new immunological tools in the field of melanoma therapeutics.     

In vivo assessment and potential diagnosis of xenobiotics that perturb the thyroid pathway: Proteomic analysis of Xenopus laevis brain tissue following exposure to model T4 inhibitors

As part of a multi-endpoint systems approach to develop comprehensive methods for assessing endocrine stressors in vertebrates, differential protein profiling was used to investigate expression patterns in the brain of the amphibian model (Xenopus laevis) following in vivo exposure to a suite of T4 synthesis inhibitors. We specifically address the application of Two Dimensional Polyacrylamide Gel Electrophoresis (2D PAGE), Isobaric Tags for Relative and Absolute Quantitation (iTRAQ®) and LC–MS/MS to assess changes in relative protein expression levels. 2D PAGE and iTRAQ proved to be effective complementary techniques for distinguishing protein changes in the developing amphibian brain in response to T4 synthesis inhibition. This information served to evaluate the use of distinctive protein profiles as a potential mechanism to screen chemicals for endocrine activity in anurans. Regulatory pathways associated with proteins expressed as a result of chemical effect are reported. To our knowledge, this is also the first account of the anuran larvae brain proteome characterization using proteomic technologies. Correlation of protein changes to other cellular and organism-level responses will aid in the development of a more rapid and cost-effective, non-mammalian screening assay for thyroid axis-disrupting chemicals.