Biomedical subjects

C Gennings

Publications and source records attributed to C Gennings.

42 records · Page 3Linked to original sources

PubMedDate not supplied

Optimization of an estrogen receptor-alpha transcriptional activation assay for testing a diverse spectrum of chemicals.

Estrogen receptor (ER) transactivation assays were initially designed to study endogenous mechanisms of steroid hormone action, but more recently have been used to assess industrial chemicals for potential estrogenic activity. Given the diverse spectrum of physicochemical properties of these chemicals, we examined the effects of pH (a weak organic and strong inorganic acid and base), hyperosmolality (NaCl, mannitol) and two different vehicles (DMSO, Triton X-100) on responses to estradiol-17beta (E2) in an ER transactivation assay. MCF-7 human breast cancer cells were transiently transfected with a chimeric estrogen receptor (Gal4-HEG0) and a Gal4-regulated luciferase reporter gene (17m5-G-Luc), treated with E2 under various test conditions, and then assessed for ER-mediated luciferase activity. Maximal E2-induced reporter activity was observed at pH 7.8 (pre-incubation), but was markedly reduced at pH < or =7.5, or > or =8.0 (P < 0.001), even though there was no evidence of cytotoxicity. Hyperosmolality induced by addition of mannitol (> or =25 mM) resulted in significant decreases in overall assay responsiveness, whereas NaCl (> or =80 mM) decreased the sensitivity of the assay by increasing the no-observed-effect level for E2 compared to control cultures (330 mOsm). The maximal DMSO concentration that resulted in consistently high E(2)-induced reporter activity was 0.1%, whereas concentrations of Triton X-100 above 1 ppm inhibited E2-induced reporter responses and were cytotoxic above 10 ppm. These results indicate that various physicochemical factors have the potential to confound assay data if not kept within predefined operational limits.

Cells, Cultured↗

PubMedDate not supplied

Autoregulation of cerebral blood flow in fetuses with congenital heart disease: the brain sparing effect.

Fetuses with congenital heart disease (CHD) have circulatory abnormalities that may compromise cerebral oxygen delivery. We believe that some CHD fetuses with decreased cerebral oxygen supply have autoregulation of blood flow that enhances cerebral perfusion (brain sparing). We hypothesize that cerebral autoregulation occurs in CHD fetuses, and the degree of autoregulation is dependent on the specific CHD and correlates with intrauterine head circumferences. CHD fetuses were compared to normal fetuses. Data included cardiac diagnosis, cerebral and umbilical artery Doppler, head circumference, weight, and gestational age. The cerebral-to-placental resistance ratio (CPR) was assessed as a measure of cerebral autoregulation. CPR = cerebral/umbilical resistance index (RI) and RI = systolic-diastolic/systolic velocity (normal CPR > 1). CPR > 1 was found in 95% of normal vs 44% of CHD fetuses. The incidence of CPR < 1 was greatest in hypoplastic left or right heart fetuses. Compared to normal, cerebral RI was decreased in CHD fetuses. The CPR vs gestational age relationship, and the relationship among weight, head circumference, and CPR differed across normal and CHD fetuses. Fetuses > 2 kg with CHD and a CPR < 1 had smaller head circumferences than normal. Brain sparing occurs in CHD fetuses. Fetuses with single ventricular physiology are most affected. Inadequate cerebral flow in CHD fetuses, despite autoregulation, may alter brain growth.

Blood Flow Velocity↗

PubMedDate not supplied

Economical designs for detecting and characterizing departure from additivity in mixtures of many chemicals.

A well-chosen experimental design can result in efficient estimation of model parameters and often savings of resources. When the interaction of components in a mixture is under study, a useful experimental layout is a factorial design-where all combinations of the levels of each component are observed together. However, as the number of components in the mixture becomes large, these types of designs become infeasible. Using the definition of additivity as proposed by Berenbaum, some authors have instead used an experimental design necessary to estimate coefficients in an additivity model where only dose-response (concentration-effect) information of single compounds is required. Two approaches for using an additivity model are described. Both compare what is observed with what is predicted under the assumption of additivity. One is based on a comparison of responses to a mixture of interest; the other is based on a comparison of locations of mixtures that yield the same predicted response when the components are at a fixed proportion. An example using a threshold model for the dose-response relationship is provided.

Animals↗

PubMedDate not supplied

Experimental designs, statistics and interpretation.

The Working Group on Experimental Designs, Statistics and Interpretation considered the use of statistics in combination toxicology, the terminology used to describe the interaction(s) of chemicals, the use of efficient experimental designs to minimize animal use, the diverse interests and goals covered by combination toxicology and approaches useful for complex mixtures. The importance of the use of appropriate experimental designs and statistical methodology was recognized. Given the present lack of consensus on terminology and methodology, it is recommended that investigators provide in their publications the definition of additivity and the mathematical model being used.

Animal Testing Alternatives↗

PubMedDate not supplied

Incorporation of S-9 activation into an ER-alpha transactivation assay.

We evaluated the feasibility of incorporating an exogenous metabolic activating system into an estrogen receptor-alpha transactivation assay. 17beta-estradiol (E2), and the proestrogenic pesticide methoxychlor (MXC) were evaluated for activity in the presence and absence of Aroclor-1254 induced rat liver S-9 fractions. Both E2 and MXC responded consistently in the assay with average EC(50) values of 9.6 x 10(-11) M and 1.2 x 10(-5) M, respectively. In the presence of a 0.1% S-9 fraction, the EC(50) for E2 was increased to 1.4 x 10(-9) M and that for MXC decreased to 4.9 x 10(-7) M, with both compounds demonstrating increased secondary metabolite formation as evidenced by HPLC analysis. Consistent with these data, metabolites of E2 and MXC exhibited decreased and increased potencies, respectively, in the assay system relative to the parent molecules. S-9 was compatible with the MCF-7 reporter assay and has the potential to enhance detection of proestrogenic materials.

Animals↗

PubMedDate not supplied

Comparing the response surfaces of two cholinolytics when used in combination with physostigmine as a pretreatment against organophosphate challenge.

Physostigmine (PHY) is currently being evaluated as a potential pretreatment against nerve agent challenge. Although it might provide increased protection against nerve agent-induced incapacitation, it is nonetheless behaviorally toxic itself. Addition of a cholinolytic adjunct is expected to significantly reduce this behavioral toxicity. The efficacy of pretreatment regimens (PRG) (composed of PHY in combination with either scopolamine (SCP) or trihexyphenidyl (artane; ART] was evaluated in guinea pigs challenged with soman (98 ug/kg, sc; 3.5 LD50) to determine the dose of PHY and adjunct required for optimal efficacy as well as the better of the two adjuncts. Three different endpoints were measured on each animal: (1) whether or not the animal survived up to 24 hours post challenge, (2) severity of incapacitation, and (3) time to recovery. The survival data revealed no significant differences between the two adjuncts and PHY against soman, but the data suggests that a pretreatment combination of PHY and ART is likely to give a greater therapeutic index than one containing SCP. Furthermore, since ART is less toxic behaviorally, it would most likely be better tolerated as a pretreatment with PHY in non-agent exposed subjects. The findings in this report are not to be construed as an official Department of the Army position unless so designated by other authorized documents. In conducting the work described in this report, the investigators adhered to the "Guide for the Care and Use of Laboratory Animals" as promulgated by the Committee on Revision of the Guide for Laboratory Animal Facilities and Care of the Institute of Laboratory Animal Resources, National Research Council.

Animals↗