Low concentration of hyaluronidase for oocyte denudation can improve fertilization rates and embryo quality.

OBJECTIVE: Hyaluronidase enzyme is an extremely important factor for the process of oocyte denudation, but little is known about its negative effects. METHODS: This prospective randomized study analyzed the results of using different concentrations of hyaluronidase (Diluted: 8IU/mL and Normal: 80IU/mL) used for denudation of sibling-oocytes for 22 women undergoing treatment for assisted reproduction by ICSI. A total of 192 oocytes were injected, being 104 for group I (diluted) and 88 for group II (normal). We analyzed fertilization rate, cleavage, embryo quality at 48 and 72 hours and number of transferred embryos in each group. RESULTS: The diluted enzyme group showed better results in fertilization rates (92.3% vs. 80.6%), mean cleavage (4.18 ± 2.57 vs. 3.09 ± 1.90), in 48-hour embryos A and A + B (60.9% vs. 44.1% and 90.2% vs. 82.3%) and at 72 hours (45.6% vs. 36.8% and 77.1% vs 66.2%), and number of embryos selected for transfer (61.8% vs. 38.1%). The overall pregnancy rate was 59.1%. CONCLUSION: This study demonstrates that the use of 8 IU/mL of hyaluronidase, according to the following protocol, is beneficial and can be successfully used for oocyte denudation, and it is also economically advantageous to the laboratory.

Evaluation of results obtained with corifollitropin alfa after poor ovarian response in previous cycle using recombinant follicular stimulating hormone in the long-term protocol.

OBJECTIVE: This study evaluated the use of Corifollitropin alfa in patients with previous poor response to recombinant follicle stimulating hormone in long-term protocols using gonadotropin-releasing hormone. METHODS: Twenty-seven poor responders to previous treatment with the long term protocol using the recombinant follicle stimulating hormone (Group 1) were selected and then submitted to a second attempt using the same long term protocol with Corifollitropin alfa instead of the recombinant follicle stimulating hormone (Group 2).Ovarian down-regulation was achieved using subcutaneous administration of Leuprolide Acetate. Ovarian stimulation was performed with recombinant follicle stimulating hormone until the administration of human chorionic gonadotropin, followed by follicular aspiration (Group 1). Group 2 was submitted to this same protocol using Corifollitropin alfa instead of recombinant follicle stimulating hormone. RESULTS: There were significant differences in the number of aspirated oocytes, percentage of mature oocytes, amount of injected oocytes and transferred embryos - with all of these parameters being increased in the Corifollitropin alfa group. In addition, the rates of pregnancy and ongoing pregnancy were also significantly higher in the Corifollitropin alfa group. CONCLUSION: The present study demonstrated that the use of Corifollitropin alfa in the long-term protocol could be a highly effective alternative for patients with poor ovarian response, who were unsuccessful in a previous treatment with In Vitro Fertilization - Intracytoplasmic Sperm Injection.

Analytical screening studies on irradiated food packaging.

Foods may be irradiated in their final packaging and this process may affect the composition of the packaging and in turn affect the migration of substances into food. Headspace and liquid injection GC-MS and HPLC with time-of-flight MS have been used to identify and estimate levels of radiolytic products in irradiated finished plastic packaging materials. Fifteen retail packaging materials were studied. Investigations were carried out into the effect of different irradiation types (gamma and electron beam), irradiation doses (1, 3, 7 and 10 kGy) and dose rates (5 kGy s(-1) for electron beam and 0.4 and 1.85 kGy h(-1) for gamma) on the radiolytic products. Any differences seen in comparing the two ionising radiation types were attributed largely to the very different dose rates; for electron beam a 10 kGy dose was delivered in just 2 s whereas using gamma it took 5.4 h. Differences were also seen when comparing the same samples irradiated at different doses. Some substances were not affected by irradiation, others decreased in concentration and others were formed upon increasing doses of irradiation. These results confirm that irradiation-induced changes do occur in substances with the potential to migrate and that the safety of the finished packaging material following irradiation should be assessed.

Printing ink compounds in foods: UK survey results.

Three hundred and fifty foodstuffs packaged in printed paper/board were purchased from UK retail outlets. Solvent extracts of all foods and associated quality assurance samples were analysed by gas chromatography-mass spectrometry (GC-MS) to determine the presence and concentrations of 20 printing ink compounds: benzophenone, 4-methylbenzophenone, 2-methylbenzophenone, 3-methylbenzophenone, 4-hydroxybenzophenone, 2-hydroxybenzophenone, 4-phenylbenzophenone, methyl-2-benzoylbenzoate, 1-hydroxycyclohexyl phenyl ketone, 2-isopropylthioxanthone, 4-isopropylthioxanthone, 2,4-diethyl-9H-thioxanthen-9-one, 2,2-dimethoxy-2-phenylacetophenone, 2-methyl-4'-(methylthio)-2-morpholinopropiophenone, 4-(4-methylphenylthio)benzophenone, ethyl-4-dimethylaminobenzoate, 2-ethylhexyl-4-(dimethylamino)benzoate, N-ethyl-p-toluene-sulphonamide, triphenyl phosphate and di-(2-ethylhexyl) fumarate. The presence of one or more of the compounds benzophenone, 4-phenylbenzophenone, methyl-2-benzoylbenzoate, 1-hydroxycyclohexyl phenyl ketone, 2,2-dimethoxy-2-phenylacetophenone, 4-(4-methylphenylthio)benzophenone, ethyl-4-dimethylaminobenzoate, 2-ethylhexyl-4-dimethylaminobenzoate and triphenyl phosphate was confirmed in some food samples. Analysis of the associated packaging material was also carried out to confirm whether or not it was likely that the occurrence of these compounds in the foods was due to migration from the printed paper/board packaging. With the exception of triphenyl phosphate, detected in one foodstuff, all the packaging material contained the substance(s) found in the food.

Modelling the joint effects of a metal and a pesticide on reproduction and toxicokinetics in Lumbricid earthworms.

It is important to understand the aetiology of interactive mixtures effects (i.e. synergism and antagonism) if results from known cases are to be extrapolated to untested combinations. The key role of toxicokinetics in determining internal concentrations at target sites means that understanding chemical uptake in mixtures is an essential requirement for mechanistic understanding of interactions. In this paper, a combined approach using mixture toxicity testing, toxicokinetic studies and modelling has been used to address the link between joint toxicity and internal concentration. The study is conducted in Lumbricid earthworms with a binary mixture of a metal (nickel) and an organophosphate insecticide (chlorpyrifos) not a priori expected to show interactive toxicity. As expected from their dissimilar modes of action and detoxification, exposure to combinations of nickel and chlorpyrifos resulted in additive toxicity. Measurement of internal concentrations indicated that both chemicals were rapidly accumulated (within 3 days) to equilibrium. When exposed as a mixture, Ni uptake followed the same pattern as found for the single chemical. This was not the case for chlorpyrifos which showed a faster rate of uptake and elimination and a slightly higher equilibrium concentration in a mixture. That the difference in chlorpyrifos kinetics in the mixture did not result in interactive toxicity highlights the need to assess chemical toxicodynamics as well as toxicokinetics. Measurement of chlorpyrifos-oxon identified the presence of this toxic form but implementation of more complex approaches encompassing toxicogenomics and epigenetics are ultimately needed to resolve the toxicokinetic to toxicodynamic link for these chemicals.

Comparative transcriptomic responses to chronic cadmium, fluoranthene, and atrazine exposure in Lumbricus rubellus.

Transcriptional responses of a soil-dwelling organism (the earthworm Lumbricus rubellus) to three chemicals, cadmium (Cd), fluoranthene (FA), and atrazine (AZ), were measured following chronic exposure, with the aim of identifying the nature of any shared transcriptional response. Principal component analysis indicated full or partial separation of control and exposed samples for each compound but not for the composite set of all control and exposed samples. Partial least-squares discriminant analysis allowed separation of the control and exposed samples for each chemical and also for the composite data set, suggesting a common transcriptional response to exposure. Genes identified as changing in expression level (by the least stringent test for significance) following exposure to two chemicals indicated a substantial number of common genes (> 127). The three compound overlapping gene set, however, comprised only 25 genes. We suggest that the low commonality in transcriptional response may be linked to the chronic concentrations (approximately 10% EC50) and chronic duration (28 days) used. Annotations of the three compound overlapping gene set indicated that genes from pathways most often associated with responses to environmental stress, such as heat shock, phase I and II metabolism, antioxidant defense, and cation balance, were not represented. The strongest annotation signature was for genes important in mitochondrial function and energy metabolism.

Reevaluation of ethylene oxide hemolysis and irritation potential.

The in vitro hemolytic and in vivo mucosal irritation potential of ethylene oxide (EO) was investigated with standard procedures used to determine the biocompatibility of medical devices. Test solutions containing EO at concentrations of 25, 50, 100, 250, 500, 1,250, 2,500, 5,000, or 10,000 microg/mL were prepared in saline to simulate a worst-case aqueous extraction of standard medical devices containing 125, 250, 500, 1,250, 2,500, 6,250, 12,500, 25,000, or 50,000 microg/g of EO, respectively. Concentrations of EO up to 500 microg/mL were not hemolytic ( < 5% hemolysis after a 4-h exposure), whereas > or =1250 microg/mL of EO resulted in significant hemolysis. Hamster cheek pouches exposed to cotton pellets saturated with EO at concentrations of up to 2500 microg/mL for 4 h with a recovery period of 14 days were without effects attributable to EO. However, at > or =5000 microg/mL of EO, significant histomorphological alterations of the buccal mucosa were observed and attributed to EO exposure. It was concluded that solutions of EO of up to 500 microg/mL representing an aqueous extract of a general medical device containing at least 2500 microg/g of EO residue do not result in significant hemolysis and irritation.

Membrane type I-matrix metalloproteinase-mediated degradation of type I collagen by oral squamous cell carcinoma cells.

Oral squamous cell carcinomas are highly invasive lesions that destroy adjacent tissues and invade bone and muscle, which is most likely the result of matrix metalloproteinase (MMP) activity. We examined three cell lines derived from squamous cell carcinoma of the tongue for their intrinsic capacities to degrade interstitial collagen with the goal of identifying the matrix-degrading enzymes. SCC-25 and SCC-15 cells degrade reconstituted fibrillar type I collagen in the absence of exogenous growth factors or cytokines when seeded as a colony on dried films. Degradation is confined to the subjacent matrix, is enhanced 2-3-fold by phorbol ester, and is strictly MMP-dependent, as it is blocked by BB-94 and tissue inhibitor of metalloproteinases-2 but not by inhibitors of serine and cysteine proteinases. Both cell lines express active (M(r) 57,000) membrane type I-MMP (MT1-MMP) on their surfaces, as detected by surface biotinylation and immunoprecipitation. Concomitantly, both cell lines activate endogenous MMP-2 when cultured on type I collagen films, as assessed by zymography. Phorbol ester treatment enhances collagen-induced MMP-2 activation, which is accompanied by the appearance of a surface-labeled M(r) 43,000 form of MT1-MMP. Treatment of cells with a synthetic furin inhibitor, which inhibits processing of the MT1-MMP zymogen, blocks collagen degradation. In contrast, CAL 27 cells do not degrade collagen under either basal or phorbol 12-myristate 13-acetate-stimulated conditions. Although proMT1-MMP (M(r) 63,000/65,000) is detectable in these cells by immunoblot analysis, they express greatly reduced levels of active MT1-MMP on their surfaces relative to SCC-25 and SCC-15 cells. Correspondingly, CAL 27 cells cultured on collagen express neither latent nor active gelatinases. Immunoblots of lysates and conditioned media revealed the constitutive expression of proMMP-1 and proMMP-13 in all three cell lines. We conclude that in the absence of exogenous growth factors or accessory stromal cells, degradation of interstitial collagen by oral squamous cell carcinoma cells requires a threshold level of active MT1-MMP on cell surfaces.

Stages in the delivery of home-based services to parents at risk of child abuse: a Healthy Start experience.

Hawaii's Healthy Start program of home-based services to families identified at risk for child abuse and neglect is becoming a model for the nation. The present paper provides background information about Healthy Start and describes a three-phase process of engagement and service delivery that characterizes the Healthy Start home visitation model.

The use of discriminant analysis for examining the histological feathers of oral keratoses and lichen planus.

In a retrospective survey of 235 cases in which the diagnosis on biopsy was lichen planus, keratosis or leukoplakia, the histological features were re-assessed as objectively as possible and without reference to the original diagnosis.The tissue changes were recorded under 39 headings, and many were assessed on a roughly quantitative basis. In addition, two clinical features were included; whether the biopsy was from the buccal mucosa (as opposed to some other intraoral site) and whether the lesions involved multiple intraoral sites. For each possible pair of diagnostic categories (keratosis and leukoplakia, lichen planus and keratosis, lichen planus and leukoplakia) the recorded findings were subjected to discriminant analysis in order to provide a quantitative assessment of the value of each individual feature for discriminating between the two diagnostic groups. The computer programme also provided for the application of these calculated values to yield a "score" for each case, and for an assessment of the significance of the separation of the diagnostic groups thus achieved. In general, the values calculated by the computer for the discriminatory value of each tissue change accorded with our subjective impressions, but a number of features that were given a relatively high value had not previously been recognized as important in differential diagnosisA discriminant analysis was also performed on those cases of leukoplakia known to have later developed a carcinoma, in comparison with the leukoplakia cases that did not develop carcinoma. High values were accorded mainly to the well-known features of epithelial atypia, but a similar high value was indicated for the presence of Russell bodies. we had not previously realized that the presence of Russell bodies was of prognostic significance in this context.When the total scores for the groups of cases were analysed, it was found that the separation of each pair of diagnostic groups was significant at the 1% level. The separation of leukoplakia cases that subsequently developed carcinoma, from those that did not develop carcinoma, was significant at the 5% level. In this latter analysis, a better separation might be achieved with larger numbers of cases, but there will always be the complicating factor that an unknown number of leukoplakia cases would develop carcinoma if the patient had received no treatment.

A computer-aided study on the tissue changes in oral keratoses and lichen planus, and an analysis of case groupings by subjective and objective criteria.

In a retrospective survey of 235 cases in which the diagnosis on biopsy was lichen planus, keratosis or leukoplakia, the histological features were re-assessed in as objective a manner as possible. For each case, the tissue changes were recorded under 37 headings, without any attempt at interpretation. The information was then transferred to punched cards, and used for various types of computer-aided analysis. Firstly, the frequency with which each tissue change occurred was assessed for each diagnostic group, and the data presented also show how often each change occurred in those cases that subsequently developed a carcinoma.Re-analysis of the cases on the basis of short lists of tissue changes that were thought to characterize each diagnostic category showed, as expected, that many other factors must have played a part in the original diagnosis.Analysis of the histological findings on an objective basis, using a cluster analysis programme, provided an encouraging degree of separation into the diagnostic groups. When a number of known carcinoma cases were included in the cluster analysis as "markers", a small number of leukoplakia and keratosis cases were placed by the computer into the same cluster as these "markers". Of the original 187 cases originally diagnosed as leukoplakia or keratosis, 4.8% are known to have developed a carcinoma, but of the 11 cases the computer placed in the same cluster as the "marker" cases of carcinoma, 36% have subsequently developed carcinoma. Thus, in the cluster analyses, the computer is tending to "recognize" those cases that later developed carcinoma, and to separate them from the bulk of the cases in which malignant change has not occurred.