This doctoral thesis describes the expression and roles of Metallothioneins I and II (MT-I+II) during CNS pathology. MT-I+II expression increases significantly in reactive astrocytes and microglia/macrophages after traumatic brain injury (cryogenic lesion in cerebral cortex) and brain stem toxicity induced by the gliotoxin

6-aminonicotinamide (6-AN). Interleukin-6 (IL-6) is a key inducer of brain MT-I+II, as judged by using genetic IL-6 deficient mice (IL-6 knock-out (IL-6KO) mice) and IL-6 overexpressing (GFAP-IL6) mice subjected to brain injury. These studies also indicate that IL-6 and/or IL-6-induced MT-I+II exert neuroprotective functions.

To study MT-I+II roles in CNS, genetically modified mice with MT-I+II deficiency (MT-I+II knock-out (MT-KO) mice) or transgenic MT-I overexpression (TgMT mice) were applied along wildtype controls; as well as wildtype, and MT-KO mice receiving exogenous MT-I or MT-II treatment were examined. Both MT-KO and TgMT genotypes and mice receiving exogenous MT were studied after cryogenic brain injury and 6-AN-induced toxicity in brain stem. These experiments overall show that MT-I+II have significant immunomodulating, antioxidant and neuroprotective roles. Accordingly, MT-I+II reduce CNS activation of macrophages and lymphocytes including expression of proinflammatory cytokines IL-1b, IL-6, IL-12 and TNF-a. Moreover, MT-I+II are antioxidant and antiapoptotic factors counteracting reactive oxygen species (ROS)/oxidative stress, neurodegeneration and apoptotic cell death, whereby the delayed (secondary) tissue damage was inhibited after brain injury and 6-AN-toxicity. MT-I+II also diminish the primary CNS toxicity caused directly by 6-AN and the clinical outcome (mortality). Additionally, MT-I+II stimulate astrogliosis; expression of growth factors, their receptors and neurotrophins (TGFb, TGFb-Receptor, bFGF, bFGF-Receptor, VEGF, NT-3, NT-4/5, NGF); angiogenesis; and growth cone formation. Hence, MT-I+II enhance CNS tissue repair as seen clearly after the cryogenic injury, after which MT-I+II promote substitution of the necrotic lesion cavity with a glial scar tissue including revascularization. In the surrounding tissue, MT-I+II stimulate growth cone formation and outgrowths of surviving neurons situated outside the lesioned area; while inside the lesion, normal neurons or neuronal cell bodies were never seen.

The angiogenic actions of MT-I+II were supported by comparing various genotypes like GFAP-IL6 mice; MT-I+II deficient GFAP-IL6 (GFAP-IL6/MT-KO) mice; MT-I+II heterozygous GFAP-IL6 mice (GFAP-IL6-MT+/- mice); double-transgenic IL-6 and MT-I overexpressors (GFAP-IL6/TgMT mice); MT-KO mice; and TgMT mice. The IL-6 overexpressors are suitable for this as they spontaneously develop proliferative angiopathy/angiogenesis, which is inhibited by MT-I+II deficiency and enhanced by MT-I overexpression. In fact, angiogenesis was highest in the MT-I overexpressing TgMT and GFAP-IL6/TgMT mice, even though the TgMT mice show quite low IL-6 levels as MT-I+II inhibit brain IL-6 mRNA and protein.

Moreover, GFAP-IL6/TgMT and TgMT and GFAP-IL6 genotypes were lesioned along wildtype controls in order to identify the roles of IL-6 versus MT-I+II. Although IL-6 induces CNS protection, this could be due to IL-6-induced MT-I+II, as neuroprotection and recovery are associated with MT-I+II levels only and not with IL-6 or inflammation: The more MT-I+II, the less neuropathology.

The MT-I+II actions seen after brain injury and 6-AN-toxicity are induced by both endogenous MT-I overexpression and exogenous MT-II treatment. As presented here for the first time, analogous therapeutic effects are obtained after brain injury by using native or recombinant MT-I or MT-II derived from diverse non-mammalian and mammalian species like drosophila, mouse, rabbit, horse and human. Treatment with these MT-I and MT-II proteins significantly reduce inflammation, oxidative stress, neurodegeneration and apoptotic cell death after brain injury, while astroglia is stimulated. This indicates that MT-I+II function independently of species of origin.

Previously, we showed that MT-I+II also ameliorate autoimmune, excitotoxic and inflammatory CNS disorders, and independent groups have confirmed this and have in fact added that MT-I+II are also major neuroprotective factors during ischemia and motor neuron disease. As emerging data validate these functions in human tissue and neurological patients, MT-I+II seem to demonstrate potentials as therapeutic targets for the treatment of CNS disorders.