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Inventory of the Spanish Institutions and Scientists Involved in Alternatives to the use of Laboratory Animals (Refinement, Reduction or Replacement)+

Inventory: 18i. Inventory of Institutions

Type of Institution: University

Staff:

Pedro M. Fernández Salguero, Email: pmfersal@unex.e, Dr Ld en Ciencias Biológicas, Profesor Titular de Universidad.
Belén Santiago Josefat, Email: sjosefat@unex.es, Ld Ciencias Biológicas, Becaria Predoctoral

Total staff involved in alternative methods is 2 people.

Activities / aims: This university department is mainly involved in basic research; it performs cell biology, molecular biology, biochemistry and toxicology (mechanisms of toxicity, carcinogenicity) testing on a routine basis.

Model systems: The model systems used are transgenic animal models (Knock-out), embryos (mouse GD13.5-15.5), in vitro methods -- micro-organisms (E. coli for cloning and expression of recombinant genes), primary culture of dispersed cells (mouse neurones, fibroblasts, epithelial glands), cell lines culture (fibroblast: NYH3T3, Swiss 3T3) and cell-free systems (cytosolic, nuclear, microsomes).

The endpoints employed are in vivo development of breast gland: cellular and molecular levels (receptors) and in vitro cell morphology (optical and electronic microscopy), cell viability (Trypan blue exclusion; MTT reduction), cellular proliferation (Tyd uptake), cell signalling (inhibitors of MAP and SAP kinases on genes regulated by AHR) and nucleic acids (condensation and fragmentation).

Experimental systems (examples):

1 fibroblast cell lines NIH 3T3, transfection with luciferase and galactosidase, gene regulation at transcription level, complementary test
2 primary culture of mouse cells, viability, proliferation, nucleic acids, role of AHR on proliferation and carcinogenesis, complementary test
3 slices of tissues from cerebella and breast gland, in situ hybridization transmission electronic microscopy, levels of gene expression related to AHR, complementary test

Work lines: Role of the aryl hydrocarbon receptor (AHR) in mouse mammary gland carcinogenesis: interaction between AHR and estrogen receptor. Implication of the AHR in viability of cerebellar granule cells: regulation of cytochrome P450s that mediate reactive oxygen species formation and nitrate oxide synthase-derived-nitrogen reactive species.

Quality assurance / Validation programmes: A Quality Assurance Unit is operating inside the facility. The institution applies Good Laboratory Practices rules. The team are available to participate in EU validation programmes in relation to method development for the identification of genes useful as tumour markers.

Sources of financing: Junta de Extremadura 1997-1999,

Others: Newly formed group

Type of Institution: University

Staff:

Carlos César Pérez García, Email: dmvcpg@unileon.es, Dr Veterinaria, Profesor Titular
Inmaculada Díez Prieto, Email: dmvidp@unileon.es, Dr Veterinaria, Profesora Titular
María J. Cano Rábano, Email: dmvmcr@unileon.es, Dr Veterinaria, Profesora Titular.
Mª Belén García Rodríguez, Email: dmvbrg@unileon.es, Dr Veterinaria, Profesora Titular.
Mª Angeles Ríos Granja, Email: dmvmrg@unileon.es, Dr Veterinaria, Ayudante

Total staff involved in alternative methods is 5 people.

Activities / aims: This university department is mainly involved in basic research and alternatives to animals in education; it performs toxicology (nephrotoxicity) and pathology testing on a routine basis, and physiology studies at a research level.

Model systems: The model systems used are conventional animal models (rodents, dogs) and education models (audiovisuals, mechanical models).

Work lines: Design and development of human disease animal models in nephrology and mineral metabolism. Teaching and training of the basic principles of animal experimentation.

Sources of financing: Junta de Castilla y León

Type of Institution: University

Staff:

Jose Becerra Ratia, Email: becerra@uma.es, Dr Biología, Catedrático.
Total staff involved in alternative methods is 8 people.

Activities / aims: This university department is mainly involved in basic research and non-regulated applied research; it performs cell biology testing on a routine basis and molecular biology studies at a research level.

Model systems: The model systems used are in vitro methods -- primary culture of dispersed cells (rat and human bone marrow) and human volunteers (bone implantation).

The endpoints employed in vitro are morphology (histology, histochemistry, immunocytochemistry, etc.), cellular proliferation (BrdU, PCNA), metabolic activity (bone formation) and biotransformation systems (biomaterials for bone formation).

Experimental systems (examples):

1 bone marrow cell cultures, bone marrow formation, osteogenic capacity in vitro

Work lines: Regeneration in fish. Bone regeneration and bone repair. Osteogenesis and growth factors.

Quality assurance / Validation programmes: The institution applies Good Laboratory Practices rules according to the EU. The team are available to participate in EU validation programmes.

Sources of financing: DGYCIT, FIS, MAPFRE, Junta de Andalucía (PAI)

Type of Institution: University

Staff:

Eugenio Vilanova Gisbert, Email: evilanova@umh.es, Dr Ciencias Químicas, Catedrático, 40 % time devoted to altern. meth., duties related to altern. meth.: Miembro Comisión Promotora de REMA, Presidente Asociación Española de Toxicología

José Barril Antuña
Maricruz Pellin Mira
Victoria Carrera González
Miguel Angel Sogorb
Maria A. Escudero
Mónica Bernabeu
Jorge Estevez
Adolfo Monroy

Total staff involved in alternative methods is 10 people.

Activities / aims: This university department is mainly involved in basic research, non-regulated applied research and alternatives to animals in education; it performs biochemistry, biokinetics and biotransformation and toxicology (mechanisms of toxicity, neurotoxicity, ecotoxicity) testing on a routine basis. Pesticides, drugs of abuse, environmental pollutants, wastes and solvents are routinely evaluated.

Model systems: The model systems used are conventional animal models and invertebrates (Daphnia), in vitro methods -- primary culture of dispersed cells (bovine chromaffine cells), cell-free systems (membranes, cytosolic), education models (data bases, multimedia) and mathematical modelling (toxicokinetics).

The endpoints employed are in vivo general observation, walking alterations, electrophysiology, histopathology, enzyme modifications) and in vitro cell viability, biotransformation systems, neurotransmitter secretion and enzyme inhibition.

Experimental systems (examples):

1 chick and human lymphocytes, bovine chromaffine cells, catecholamine secretion, protein phosphorylation ionic current, esterases, mechanisms and effects of esterases inhibition by organophosphate, replacement test
2 Daphnia, enzyme inhibition, death, ecotoxicity model for waste water, replacement test
3 adult hen, locomotion, reflex, enzyme inhibition, delayed neurotoxicity, drastic reduction in animal use

4 slaughterhouse material: extracts from nervous tissue and peripheral nerves, esterase organophosphorylation kinetics, toxic molecular mechanisms, reduction

Work lines: Biotransformation of organophosphorus pesticides; mechanism and role of phosphotriesterases. Characterization of esterases as target for the induction and promotion of toxic neurodegenerative syndromes. The characterization and role of the so called soluble-neuropathy target esterase (S-NTE). Chromaffin cells as model for studying mechanism of molecular interaction and organophosphorus compounds toxicity. Characterization of Daphnia magna as model for testing pesticides and other environmental pollutants ecotoxicity. Toxicological evaluation of agrochemical and of industrial solvents.

Quality assurance / Validation programmes: The team are available to participate in EU validation programmes.

Sources of financing: FIS, CICYT, industries

Type of Institution: University

Staff:

Carlos M. Carceles Rodríguez, Email: carceles@fcu.um.es, Dr Veterinaria, Prof Titular
Elisa Escudero Pastor, Email: escudero@fcu.um.es, Dra Veterinaria, Prof.Titular U

Total staff involved in alternative methods is 2 people.

Activities / aims: This university department is mainly involved in non-regulated applied research; it performs biokinetics and biotransformation testing on a routine basis. Pharmaceuticals are routinely evaluated; wastes also being occasionally studied.

Model systems: The model systems used are conventional animal models and in vitro methods -- micro-organisms (Hicrococus luteus).

Work lines: Pharmacokinetics of drugs (antibiotics and anthelmintics) in animals

Quality assurance / Validation programmes: The institution applies Good Laboratory Practices rules according to the EU. The team have previously been involved in alternative method validation programmes

Sources of financing: CICYT

Type of Institution: University

Staff:

Fernando Tecles Vicente, Ld Veterinaria, Becario.
Jose Joaquín Cerón Madrigal, Email: jjceron@fcu.um.es, Dr. Veterinaria, Profesor Titular.
Cándido Gutierrez Panizo, Email: cguti@cu.um.es, Dr Veterinaria, Catedrático del Departamento de Patología Animal.

Total staff involved in alternative methods is 3 people.

Activities / aims: This university department is mainly involved in basic research; it performs toxicology (ecotoxicity) testing on a routine basis. Pesticides and environmental pollutants are routinely evaluated.

The main use of alternative methods is for complementary studies.

Model systems: The model systems used are conventional animal models and in vitro methods -- primary culture of dispersed cells (chick embryo neuroblasts).

The endpoints employed are in vivo haematology, organic metabolites, blood enzyme activities and in vitro metabolic activity (cholinesterase).

Experimental systems (examples):

1 chick embryo neuroblasts culture, cholinesterase inhibition, by spectrophotometry, toxicity of pesticides, complementary test

Work lines: Haematological and biochemical effects of environmental contaminants in animals. Analytical techniques applied in environmental biomonitoring.


Type of Institution: University

Staff:

Antonio Juan García Fernández, Email: ajgf@fcu.um.es, Dr Veterinaria, Profesor Titular de Toxicología, 50 % time devoted to altern. meth.
Diego Romero García, Dr Veterinaria, Contrato de Investigador, 100 % time devoted to altern. meth
Maximiliano Gómez Zapata, Dr Medicina y Cirugía, Profesor Titular, 100 % time devoted to altern. meth.
Miguel Motas Guzmán, Email: mmotas@fcu.um.es, Ldo Veterinaria, Becario FPI, 75 % time devoted to altern. meth.
Pedro José Mojica, Dr Veterinaria, Investigador, 75 % time devoted to altern. meth.
Isabel Maria Navas Ruiz, Lda Veterinaria, Investigadora, 50 % time devoted to altern. meth.

Total staff involved in alternative methods is 6 people.

Activities / aims: This university department is mainly involved in basic research, non-regulated applied research, method development and alternatives to animals in education; it performs toxicology (mechanisms of toxicity, basal cytotoxicity, nephrotoxicity) and monitoring --chemical, biological- studies on a routine basis, and toxicology (ocular cytotoxicity, dermal irritation and corrosivity, reproductive cytotoxicity, immunotoxicity, sensitisation, endocrine disruption, chronic toxicity) studies at a research level. Pesticides, environmental pollutants and wastes are routinely evaluated; cosmetics also being occasionally studied.

The main use of alternative methods is for complementary and replacement studies.

Model systems: The model systems used are conventional animal models (refinement) and in vitro methods -- cell lines culture (BF-2, RTG-2, CHO, VERO, PK-15, BGM, EPO).

The endpoints employed in vivo are lipid peroxidation, glutathione and related enzymes, ALA-D, LDH and metallothioneins.

Experimental systems (examples):

1 kidney cell lines, neutral red uptake, MTT reduction and LDH, toxicity evaluation, replacement test
2 kidney cell lines, electronic microscopy, risk assessment, complementary test

Quality assurance / Validation programmes: The institution applies Good Laboratory Practices rules according to the EU. The team are available to participate in EU validation programmes.

Type of Institution: University

Staff:

Adela López de Cerain Salsamendi, Email: acerain@unav.es, Dra Biología, Profesora Adjunta, 10 % time devoted to altern. meth., duties related to altern. meth.: Miembro del Comité de Etica para la Experimentación animal de la UNAV
Olga Ezpeleta Echevarri, Email: oezpeleta@unav.es, Ld Biología, Ayudante, 35 % time devoted to altern. meth.

Total staff involved in alternative methods is 2 people.

Activities / aims: This university department is mainly involved in basic research; it performs toxicology (mechanisms of toxicity, basal cytotoxicity, genotoxicity / mutagenicity) testing on a routine basis. Toxins are routinely evaluated; pharmaceuticals, cosmetics and medical devices also being occasionally studied.

The main use of alternative methods is for screening and complementary studies.

Model systems: The model systems used are conventional animal models and in vitro methods -- primary culture of dispersed cells (rat hepatocytes, human lymphocytes), cell lines culture (V79, CHO, MDCK, HT-29, Vero, EVIT-6, MDA-468, etc.

The endpoints employed in vitro are cell viability (MTT reduction, neutral red uptake), cytoskeleton /membranes/enzyme release (LDH leakage), cellular proliferation (clonogenicity, total protein content), metabolic activity and biotransformation systems (S9 fractions from tissues induced for cytochromes).

Quality assurance / Validation programmes: A Quality Assurance Unit is operating inside the facility. The institution applies Good Laboratory Practices rules.

Fuente / Source: Guillermo Repetto, Ana del Peso, Manuel Salguero, Manuel Repetto (1999) Inventory of the Spanish Institutions and Scientists Involved in Alternatives to the use of Laboratory Animals (Refinement, Reduction or Replacement) Revista de Toxicología 16: 50-127.