Knockout Animal Models

Siarim, M. and Krishnamurthy, H. 2001. The role of follicle stimulating hormone in spermatogenesis lessons from knockout animal models. Archives of Medical Research 32(6), 601-608. 

Rosenberg MP, Bortner D. Why transgenic and knockout animal models should be used (for drug efficacy studies in cancer). Cancer Metastasis Rev 1998 17 295-9. 

A. Inui, Cytokines and sickness behaviour implications from knockout animal models, Trends in Immunology, 22, 2001, 469-73. 

The evidence for the role of Toll-like receptors 4 or 2 as receptors for OxPLs is based on impaired responses to OxPLs or mmLDL in knockout animal models or after knockdown of TLRs in cultured cells [49-54], OxPLs do not demonstrate canonical activation of TLRs in cells naturally expressing these receptors or in reporter cell lines [55-57]. It can be hypothesized that this discrepancy is explained by the need of additional soluble or membrane-associated proteins that present OxPLs to TLRs [51,52]. The role of TLRs as candidate receptors mediating proinflammatory action of OxPLs is discussed in more detail in Chapter 11 in this book. 

Knockout mice have been reported for several FATPs [1]. As insulin desensitization has been closely linked to excessive fatty acid uptake and intracellular diacylgly-cerol and TG accumulation, these animal models were particularly evaluated in the context of protection from diet-induced type 2 diabetes ( Type 2 Diabetes Mellitus (T2DM)). In addition, studies on human subjects have also established genetic links between polymorphisms in FATP genes and metabolic alterations [1]. 

Although many animal models for iron overload exist, some mimicking certain aspects of HH, the 32-microglobulin knockout mouse is of special interest as it revealed for the first time crucial aspects of the pathogenesis of human HH in an animal model, and also because it underlines the important links between iron metabolism and the immune system. Hepatic iron overload in 32-microglobulin ( 32m)-deficient mice appeared to be similar to that found in HH, with pathological iron depositions occurring predominantly in liver parenchymal cells (de Sousa et ah,... 

Likelihood of developing dependence to nicotine will involve specific functional changes in the brain. Examining the detailed genetic basis for these functional changes is difficult in humans, so animal models are needed. Three approaches have been taken to examining genetic influences of the effects of nicotine in rodents namely inbred lines, selectively bred lines and knockout mice. 

During product development, many of the initial non-clinical studies were undertaken in GAA knockout mice (i.e. mice devoid of a functional GAA gene), which serves as an animal model for Pompe s disease. The mice proved useful in assessing the pharmacodynamic effect of Myozyme on glycogen depletion and helped establish appropriate dosage regimens. The mice were also used to evaluate pharmacokinetics and biodistribution of GAA following its administration at clinically relevant doses. 

This methodology provides resolution of approximately 1 mm. Thus, not only in vitro but in vivo data from genetic knockouts and other animal models can be obtained with PET and more recently SPECT imaging [65]. 

In the case of spontaneous autoimmune diseases mice are the most frequently used animal model. With the advent of transgenic and genetically modified (knockout, KO) mice, the number of genetically predisposed autoimmune models has substantially increased. Other species that have been useful include rats, monkeys, cats, dogs, rabbits, and chickens for some specific forms of autoimmune diseases [4, 5]. 

Inhibitors of Tpl-2 (MKKK), MEK1/2 (MKK1/2) and ERK1/2 have been reported. Macrophages from Tpl-2 knockout mice lack ERK1/2 activation leading to loss of TNFa expression and are insensitive to LPS-induced endotoxin shock [9]. Inhibition of Tpl-2 and MEK1/2 produces potent anti-inflammatory effects in cellular and animal models. 

For in vivo studies, animal models are set up and how the target is involved in the disease is analyzed. One such model is the use of knockout or transgenic mice (Exhibit 2.8). It should be borne in mind, however, that there are differences between humans and animals in terms of gene expression, functional characteristics, and biochemical reactions. Nevertheless, animal models are important for the evaluation of drug-target interactions in a living system. 

Ti-ansgenic and knockout animals are used as models of human disease. 

In the case of immunoglobulin, microarray data indicated the opposite effect that the Fc-receptor is elevated in chronic MS but not in acute lesions. Using Fcy-receptor knockout mice, the disease was found to be absent. Inter-venous immrmoglobulin therapy in the EAE mouse model was reported (see Lock, 2002, Reference 29). In summary. Lock et al. were able to apply the results of microarray-based gene expression clustering of a human disease pathological state (acute vs. chronic MS) to successfully identify fherapeutic targets for an animal model (EAE) potentially applicable to the human condition. 

The best evidences are studies from preclinical animal models [86, 87, 105], or knockout animals lacking appropriate anti-oxidative pathways [106]. For example, Balb/c mice administered a variety of anti-oxidants in their chow were protected from acetaminophen hepatotoxicity [107]. Rats fed with the anti-oxidant melatonin were protected from cholesterol mediated oxidative liver damage [108]. The best clinical evidence that oxidative stress is a key player in a variety of liver injury diseases is the beneficial application of silymarin in these disease indications [109]. Silymarin is a polyphenolic plant fiavonoid (a mixture of flavonoid isomers such as silibinin, isosilibinin, silidianin and silichristin) derived from Silymarin maria-num that has antioxidative, antilipid peroxidative, antifibrotic and anti-inflammatory effects [109, 110]. 

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