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Biomedical subjects

Horst Spielmann

Publications and source records attributed to Horst Spielmann.

At least 55 records · Page 3Linked to original sources
PubMed1997

[Reduction of the numbers of animals used for the classification of the acute oral toxicity of chemicals by taking into account cytotoxicity data from the Registry of Cytotoxicity]

Using cytotoxicity data a new classification procedure is introduced which will allow to allocate chemicals to the four toxicity classes for acute oral toxicity according to EU regulation. Simultaneously, the new procedure allows to reduce animals in experiments for determination these four toxicity classes. The cytotoxicity data estimated from in vitro cultivated mammalian cell lines were taken from the "Registry of Cytotoxicity" (RC) in which the mean IC50 (IC50x) of 347 chemicals are stored as well as the acute oral toxicity data (LD50) for rats and mice taken from NIOSH registry. As we have been used to calculate a standard regression line for predicting the dosage range of acute oral toxicity in the two species. The maximum, mean and minimum dosages of oral toxicity were predicted from the RC data and furthermore, these dosages were the basis for allocating chemicals into the four classes of acute oral toxicity defined by the EU. The accuracy for predicting the toxicity classes of the 347 chemicals registered in the RC in comparison to the toxicity classes of the corresponding NIOSH LD50 values amounts 80 percent to 88 percent. We have developed a tier testing strategy for the classification of chemicals into EU toxicity classes which takes into account the cytotoxicity data as predicted mean LD50 in connection with the protocol steps described in Acute Toxic Class (ATC) method. Compared with the ATC method the new combined RC-ATC procedure will allow to reduce animal numbers for allocating chemicals to the EU toxicity classes by about 30 percent. We suggest to validate the RC-ATC procedure in order to achieve regulatory acceptance at the international level.

Journal Article↗
PubMed1997

[Development of a new in vitro test for dermal phototoxicity using a model of reconstituted human epidermis]

Results are reported using the human epidermis model (EpiDermtrade mark) in a test protocol that was adopted from the method developed by ZEBET for phototoxicity testing with the model Skin2. In the new phototoxicity test the cytotoxicity of test materials applied topically to EpiDermtrade mark is determined at five concentrations in the absence and presence of a non-cytotoxic dose of UVA and visible light (sun simulation). Cytotoxicity is determined one day after irradiation in the MTT assay. Experiments performed to optimise test conditions are presented. Using appropriate phototoxic and non-phototoxic test chemicals our results demonstrate that a reconstructed human epidermal model, e.g. EpiDermtrade mark, can be used in phototoxicity testing in the same way as a full skin model.

Journal Article↗
PubMed1996

[Different concepts for risk assessment have to be applied if the data are derived from testing in animals or from testing in vitro]

In vitro tests are well accepted by toxicologist for mechanistic studies but they have so far not been able to replace testing in animals for regulatory purposes, as e.g. risk assessment and risk management. This is basically due to differences in the quality of results obtained from in vivo and in vitro studies. Therefore, a new concept for risk assessment and risk management is proposed which is entirely based upon data from in vitro tests.

Journal Article↗
PubMed1996

[OECD ist accepting test guidelines for validated in vitro toxicity tests in 1996]

Since 1990 in Europe a scientific concept for the validation of in vitro toxicity tests has been developed to facilitate regulatory acceptance of the new methods at the international level. ERGATT and ECVAM have promoted the concept of validation on two workshops in 1990 and 1994. A pre-validation stage is an essential part of this validation concept to achieve a better standardisation of in vitro tests before entering formal validation. Within the NTP in 1995/96 Federal Agencies of the USA represented by the validation center ICCVAM have accepted a validation concept, which basically agrees with the essentials of ECVAM"s European validation concept. Subsequently, in January of 1996 the major industrial nations have at the OECD level agreed to the European/US validation concept. This will at the international level allow mutual acceptance of data obtained with in vitro toxicity tests rather than with animal tests. It seems very likely that in 1996 in vitro testing for skin penetration with human skin will be the first in vitro toxicity test accepted by the OECD. Several in vitro tests for local irritancy testing will follow in 1997/98, since they are currently undergoing validation according to OECD criteria.

Journal Article↗
PubMed1996

[Replacement of the frog test in physiology teaching by the myograph: practical validation in Germany]

The myograph is a newly developed instrument to replace the classical frog experiments in physiology teaching by testing on volunteers, usually by self-testing of the students. First experiences with the myograph obtained during physiology teaching at four universities in Germany are summarized and evaluated. The myograph was tested in four additional universities. The myograph proves to be superior to the classic frog experiments both from the scientific and educational point of view. Costs of purchasing and maintaining the myograph will not be an obstacle for introducing the new teaching device into higher education. Technical development and practical evaluation of the myograph were funded by the German Ministry of Education and Research BMBF.

Journal Article↗
PubMed1995

[The sophisticated process of validation]

Current concepts on the efficient performance of validation studies are summarized. From the experience of recent attempts to validate alternative methods to animal experiments, the difficult procedure of validation should be refined. The applications of toxicological tests (safety testing) and pharmacological methods (identification of putative drugs) require different validation stategies. Suggestions as to an improved design of test development, evaluation, prevalidation, definitive validation as well as final assessment of the result are presented.

Journal Article↗
PubMed1994

[First results of an EC/COLIPA validation project of in vitro phototoxicity testing methods]

In a joint project six laboratories from the European cosmetics industry (COLIPA) as well as from FRAME (England) and ZEBET (Germany) are validating in vitro methods to be incorporated into new international guidelines for photoirritancy testing. During the first stage of the study it was attempted to select the most promising in vitro photoirritancy tests for further validation. Twenty chemicals with known photoirritation properties (12 phototoxins (PT), 4 non-PTs and 4 UV absorbing non-PTs) were tested under identical UV exposure conditions (5 J/cm2, UV-A sun simulator) in a standardized cytotoxicty assay using 3T3 fibroblasts (endpoint): Neutral red uptake, NRU). The chemicals were also tested with in vitro phototoxicity assays established in industrial laboratories, e.g. the photohaemolyses (Pape et al, 1993), histine oxidation, candida albicans (Johnson et al, 1986), and, furthermore two commercial tests (SOLATEX PItrade mark and Skin2trade mark). Data from the 3T3 NRU photoirritancy test, the red blood cell photohaemolysis test and the Skin2trade mark assay showed a better overall correlation to human in vivo data than results from the other tests. These simple assays therefore, seem very promising for further validation under blind conditions. The protocols of the other tests have to be improved and standardized to permit better interlaboratory comparison.

Journal Article↗
PubMed1994

[Prediction of acute toxicity]

The updated register of cytotoxicity data (RCD) is containing 361 chemicals with a total of 1912 IC50 values, for each of which a mean IC50 (IC50) has been calculated. In addition, the RCD is containing oral and i.v. LD50 values for rats and mice from the NIOSH resister RTECS. For 347 of the data pairs IC50/LD50 p.o. and for 148 of the data pairs IC50/LD50 i.v. linear standard regression lines have been calculated, which allow to predict an approximative LD50 value from a given IC50. For a group of 26 neurotropic agents documented in the RCD the prediction of LD50 values from IC50 data was surprisingly good. This suggests that even for neurotropic agents there is a sufficient positive correlation between in vitro and in vivo toxicity. The unexpected result is discussed with respect to basal cytotoxicity which seems to be reflected by IC50 and which is related to basic functions of the cells, as e.g. proliferation, DNA- and protein synthesis, rather than to specific endpoints of neurotoxicity. The results obtained with 26 neurotropic agents support the concept that acute toxicity in vivo can sufficiently be predicted from the data of the updated RDC. An attempt is made to predict the approximate LD50 values, both p.o. and i.v., data from IC50 values of the RDC for an established neurotropic agent with insufficient acute in vivo toxicity data.

Journal Article↗
PubMedDate not supplied

Improvement of an in vitro stem cell assay for developmental toxicity: the use of molecular endpoints in the embryonic stem cell test.

The embryonic stem cell test (EST) takes advantage of the potential of murine embryonic stem (ES) cells to differentiate in culture to test embryotoxicity in vitro. The EST represents a reliable, scientifically validated in vitro system for the classification of compounds according to their teratogenic potential based on the morphological analysis of beating cardiomyocytes in embryoid body (EB) outgrowths compared to cytotoxic effects on undifferentiated murine ES cells and differentiated 3T3 fibroblasts. In order to identify more objective endpoints of differentiation other than the microscopic evaluation of "beating areas" and to adapt the EST to applications in high-throughput screening systems we improved and expanded the EST protocol by establishing molecular endpoints of differentiation. The quantitative expression of sarcomeric myosin heavy chain (MHC) and alpha-actinin genes under the influence of test compounds was studied employing intracellular flow cytometry. Strong embryotoxicants exerted a dose-dependent effect on both the expression levels of MHC and alpha-actinin and the differentiation into beating cardiomyocytes. Furthermore, quantitative FACS (fluorescence-activating cell sorting) analysis showed the same sensitivity for the classification of substances as the conventional endpoint but allowed a significant reduction of the test period. Within 7 days, maximal expression of sarcomeric marker proteins was observed. Our findings indicate that structural proteins of the sarcomere apparatus, alpha-actinin and myosin heavy chain (MHC), seem to be promising candidates to predict developmental toxicity in vivo from in vitro data. Thus, the improved EST holds promise as a new predictive screen for risk assessment with respect to developmental toxicity using stem cell technology and technological advances in the field of gene expression analysis.

Actinin↗
PubMedDate not supplied

Validation and regulatory acceptance of new carcinogenicity tests.

The 3 Rs concept, which was developed by Russell and Burch in 1959, was implemented into the legal framework in the European Union(EU) for the protection of vertebrate animals used for experimental and other scientific purposes, when Directive 86/609/EEC was adopted in 1986. To reduce or replace animal testing for regulatory purposes, nonanimal tests must be independently validated to prove that they can provide information that is relevant and reliable for hazard prediction in vivo. At the end of the 1980s, no scientific concept existed for the formal validation ofin vitro toxicity tests, so a small group of European and American scientists developed a set of principles for experimental validation, which was accepted internationally by the OECD in 1996. A major breakthrough was the acceptance in the EU of the scientifically validated in vitro toxicity test for phototoxic potential in 2000, which was accepted by the OECD at the worldwide level in 2002. Taking the progress in the development and validation of alternative toxicity tests during the past decade into account, the current concepts of developing alternatives to the standard 2-year rodent bioassay for carcinogenicity testing are discussed.

Animal Testing Alternatives↗
PubMedDate not supplied

The ECVAM international validation study on in vitro embryotoxicity tests: results of the definitive phase and evaluation of prediction models. European Centre for the Validation of Alternative Methods.

From 1996 to 2000, ZEBET (Centre for Documentation and Evaluation of Alternative Methods to Animal Experiments at the BgVV, Berlin, Germany) coordinated the European Centre for the Validation of Alternative Methods (ECVAM) prevalidation and validation study on three embryotoxicity tests: a) a test employing embryonic stem cell lines (EST); b) the micromass (MM) test; and c) the postimplantation rat whole-embryo culture assay (WEC test). The main objectives of the study were to assess the performance of these three in vitro tests in discriminating between non- embryotoxic, weakly embryotoxic and strongly embryotoxic compounds. Phase I of the study (1997) was designed as a prevalidation phase, for test protocol optimisation, and for the establishment of a comprehensive database of in vivo and in vitro data on embryotoxic compounds. Phase II (1998-2000) involved a formal validation trial, conducted under blind conditions on 20 test compounds selected from the database, which were coded and distributed to the participating laboratories. In the preliminary phase of the validation study, six chemicals out of the 20, which showed embryotoxic potential, were tested. These results were used to define new biostatistically based prediction models (PMs) for the MM and WEC tests, and to evaluate those developed previously for the EST. As a next step, the PMs were evaluated by using the results for the remaining 14 chemicals of the definitive phase of the validation study. The three in vitro embryotoxicity tests proved to be applicable to testing a diverse group of chemicals with different embryotoxic potentials (non-embryotoxic, weakly embryotoxic, and strongly embryotoxic). The reproducibility of the three in vitro embryotoxicity tests were acceptable according to the acceptance criteria defined by the Management Team. The concordances between the embryotoxic potentials derived from the in vitro data and from the in vivo data were good for the EST and the WEC (PM2) test, and sufficient for the MM test and the WEC (PM1) tests according to the performance criteria defined by the Management Team before the formal validation study. When applying the PM of the EST to the in vitro data obtained in the definitive phase of the formal validation study, chemicals were classified correctly in 78% of the experiments. For the MM and the WEC tests, the PMs provided 70% and 80% (PM2) correct classifications, respectively. And, very importantly, an excellent predictivity (100%, except for PM1 of the WEC test, with 79%, considered as good) was obtained with strong embryotoxic chemicals in each of the three in vitro tests.

3T3 Cells↗