Inactivation of TNF-α ameliorates diabetic neuropathy in mice

Abstract

Tumor necrosis factor (TNF)-α is a potent proinflammatory cytokine involved in the pathogenesis of diabetic neuropathy. We inactivated TNF-α to determine if it is a valid therapeutic target for the treatment of diabetic neuropathy. We effected the inactivation in diabetic neuropathy using two approaches: by genetic inactivation of TNF-α (TNF-α(-/-) mice) or by neutralization of TNF-α protein using the monoclonal antibody infliximab. We induced diabetes using streptozotocin in wild-type and TNF-α(-/-) mice. We measured serum TNF-α concentration and the level of TNF-α mRNA in the dorsal root ganglion (DRG) and evaluated nerve function by a combination of motor (MNCV) and sensory (SNCV) nerve conduction velocities and tail flick test, as well as cytological analysis of intraepidermal nerve fiber density (IENFD) and immunostaining of DRG for NF-κB p65 serine-276 phosphorylated and cleaved caspase-3. Compared with nondiabetic mice, TNF-α(+/+) diabetic mice displayed significant impairments of MNCV, SNCV, tail flick test, and IENFD as well as increased expression of NF-κB p65 and cleaved caspase-3 in their DRG. In contrast, although nondiabetic TNF-α(-/-) mice showed mild abnormalities of IENFD under basal conditions, diabetic TNF-α(-/-) mice showed no evidence of abnormal nerve function tests compared with nondiabetic mice. A single injection of infliximab in diabetic TNF-α(+/+) mice led to suppression of the increased serum TNF-α and amelioration of the electrophysiological and biochemical deficits for at least 4 wk. Moreover, the increased TNF-α mRNA expression in diabetic DRG was also attenuated by infliximab, suggesting infliximab's effects may involve the local suppression of TNF-α. Infliximab, an agent currently in clinical use, is effective in targeting TNF-α action and expression and amelioration of diabetic neuropathy in mice.

Ablation of a small subpopulation of diabetes-specific bone marrow-derived cells in mice protects against diabetic neuropathy

“…Before diabetes induction, none of the mouse DRGs displayed detectable PI or TNF␣ immunostaining. In agreement with previous observations (8,18,22,27), we noted a large number of immunoreactive PI-positive and TNF␣-positive cells in the DRGs of the diabetic (DM) iDTRf¡WT control mice. Importantly, we found a significantly reduced number of PI-and TNF␣-positive cells in the DRGs of DM DT-ablated iDTR-f/RIP-Cre¡WT mice (Fig.…”

“…In contrast, however, the mRNA expression level of these mRNAs was not significantly different in the DRGs of non-DM or DM iDTR-f/RIP-Cre¡WT mice. Therefore, DT ablation of PI-BMDCs substantially reduced the level of PIBMDCs (marked biochemically by Ins1 and Ins2 expression) in the DRGs of iDTR-f/RIP-Cre¡WT mice; ablation also substantially suppressed the level of Tnf mRNA expression, a gene product known to occur in PI-BMDCs that had also been shown to play a pathogenic role in DPN (8,18,22,27).…”

“…We subsequently reported that the diabetes-specific PI-BMDCs play a key role in the pathogenesis of DPN by directly damaging dorsal root ganglion (DRG) neurons in diabetic rodents (19). Most of these abnormal PIBMDCs coexpress TNF-␣; the TNF␣-expressing PI-BMDCs fuse with DRG neurons, leading to their malfunction and premature apoptosis, culminating in DPN (8,18,22,27).Studies to date strongly suggest that PI-BMDCs are heavily involved in the pathogenesis of DPN. In this investigation, we sought to determine whether PI-BMDCs play an essential role in DPN development and whether this cell type could be a target for therapeutic intervention.…”

“…We subsequently reported that the diabetes-specific PI-BMDCs play a key role in the pathogenesis of DPN by directly damaging dorsal root ganglion (DRG) neurons in diabetic rodents (19). Most of these abnormal PIBMDCs coexpress TNF-␣; the TNF␣-expressing PI-BMDCs fuse with DRG neurons, leading to their malfunction and premature apoptosis, culminating in DPN (8,18,22,27).…”

Animal Models of Peripheral Neuropathies

“…Use of intraepidermal nerve fiber density as a true morphological correlate of the degree of sensory axonal loss is likely to improve the usefulness of rodent models of DPN, both in mechanistic studies and in preclinical efficacy studies of potential therapeutic drugs. Recent examples of drugs demonstrating efficacy in reversing loss of epidermal innervation in diabetic rats and mice include drugs aimed at different molecular mechanisms of DPN [35][36][37][38][39][40]. It remains to be seen whether these drugs fare better in clinical trials compared to previous drugs that relied on changes in nerve conduction velocity as the primary outcome measures in preclinical studies.…”