Functional, morphological and immunohistochemical studies of the peripheral nervous system in the diabetic rip-i/hifn¿ transgenic mouse model, and additional studies of diabetic neuropathy in other murine models

Resumen

Diabetic neuropathy (DPN) is the most frequent secondary complication of Diabetes Mellitus (DM), and affects more than 60% of diabetic patients. The most important contributors to reduction in the quality of life of patients with DPN are neurological pain and foot ulcerations, which are responsible for more than 75% of non-traumatic amputations. The pathogenesis of DPN is multifactorial, complex, and poorly understood. Early manifestations of the disease mainly consist of functional sensory and motor abnormalities, which are associated to structural alterations in the peripheral nervous system (PNS) at chronic stages of the disease. Structural changes in peripheral nerves from diabetic patients include distal axonal degeneration, degenerative changes in sensory neurons and Schwann cells (SCs), demyelination, and vascular changes. Among possible causes, the progressive fiber loss has been attributed to impaired nerve fiber regeneration in diabetic patients. Unfortunately, despite numerous clinical trials, there is a lack of effective therapies against DPN and current treatment remains largely symptomatic, non-specific and not uniformly effective. Experimental animal models are useful biological tools to investigate the underlying mechanisms involved in DPN; however, none of them faithfully reproduce the human disease. The global aim of the present study was to investigate functional and structural abnormalities in the PNS of the diabetic RIP-I/hIFNß mice due to short duration of DM, as well as to detect possible alterations in peripheral nerve degenerative and regenerative responses in these animals at different time points after sciatic nerve crush. Our results in intact nerves indicated that, after 3 months of DM, diabetic RIP-I/hIFNß mice mimic early stages of DPN in human patients and other animal models. Early DPN in these mice is characterized by electrophysiological abnormalities in motor and sensory parameters and reduction of intraepidermal nerve fiber (IENF) density in foot pad. Four and 8 weeks after nerve crush, our diabetic mice showed delayed recovery of motor and sensory electrophysiological responses. Impaired functional recovery was consistent with altered nerve fiber maturation, with myelin thickness being the most affected parameter, and increased number of SCs associated to regenerating fibers, as noted by morpohmetrical and teased-nerve fiber studies in tibial nerves, respectively. Furthermore, our diabetic mice showed delayed small nerve fiber innervation in foot pads. At eight weeks after crush, these findings were accompanied by increased immunopositivity for all neurotrophins and their receptors in injured nerves, correlated in most cases to the same results in dorsal root ganglia (DRG) and spinal cord, and by cellular stress-related markers in DRG. Taken together, these results suggest that among other possible causes, SC dysfunction play an important role in the impaired nerve regeneration found in our diabetic mice. From these studies we conclude that the diabetic RIP-I/hIFNß transgenic mice should be considered a potential useful model for the study of DPN avoiding the possible neurotoxic effect of STZ. It will be also useful to investigate the underlying mechanisms contributing to impaired nerve regeneration in diabetic patients and to assess new therapeutic strategies against DPN. Additionally, previous studies carried by the group of Dra. Fàtima Bosch characterized the transgenic hIFN/IGFII as a good animal model of spontaneous type 1 DM. In order to determine whether these mice could serve for the study of DPN, functional and histopathological investigations were carried in this model, and the obtained results were compared with those from the same studies in db/db mouse, an spontaneous model of type 2 DM that has been already characterized as a good animal model of DPN. Our investigations in long-term diabetic models lead to the thought that the spontaneous type 1 diabetic transgenic hIFNß/IGF-II should not be considered as a good model of DPN since only mild electrophysiologial abnormalities in the sciatic/tibial nerves, without evidence of IENF loss in foot pads were found after 6 months of DM.