Yan Ping Zhang

Transgenic Inhibition of Glial NF-kappa B Reduces Pain Behavior and Inflammation after Peripheral Nerve Injury

&NA; The transcription factor nuclear factor kappa B (NF‐&kgr;B) is a key regulator of inflammatory processes in reactive glial cells. We utilized a transgenic mouse model (GFAP‐I&kgr;B&agr;‐dn) where the classical NF‐&kgr;B pathway is inactivated by overexpression of a dominant negative (dn) form of the inhibitor of kappa B (I&kgr;B&agr;) in glial fibrillary acidic protein (GFAP)‐expressing cells, which include astrocytes, Schwann cells, and satellite cells of the dorsal root ganglion (DRG) and sought to determine whether glial NF‐&kgr;B inhibition leads to a reduction in pain behavior and inflammation following chronic constriction injury (CCI) of the sciatic nerve. As expected, following CCI nuclear translocation, and hence activation, of NF‐&kgr;B was detected only in the sciatic nerve of wild type (WT) mice, and not in GFAP‐I&kgr;B&agr;‐dn mice, while upregulation of GFAP was observed in the sciatic nerve and DRGs of both WT and GFAP‐I&kgr;B&agr;‐dn mice, indicative of glial activation. Following CCI, mechanical and thermal hyperalgesia were reduced in GFAP‐I&kgr;B&agr;‐dn mice compared to those in WT, as well as gene and protein expression of CCL2, CCR2 and CXCL10 in the sciatic nerve. Additionally, gene expression of TNF, CCL2, and CCR2 was reduced in the DRGs of transgenic mice compared to those of WT after CCI. We can therefore conclude that transgenic inhibition of NF‐&kgr;B in GFAP‐expressing glial cells attenuated pain and inflammation after peripheral nerve injury. These findings suggest that targeting the inflammatory response in Schwann cells and satellite cells may be important in treating neuropathic pain.

Prophylactic and antinociceptive effects of coenzyme Q10 on diabetic neuropathic pain in a mouse model of type 1 diabetes.

Background:Oxidative stress is a key factor implicated in the development of diabetic neuropathy. This study evaluates the prophylactic and antinociceptive effects of the antioxidant coenzyme Q10 (CoQ10) on diabetes-induced neuropathic pain in a diabetic mouse model. Methods:Total 56 mice with type 1 diabetes induced by streptozotocin were used, 20 normal mice were used as control. Mechanical and thermal nociceptive behavioral assays were applied to evaluate diabetic neuropathic pain. Tissue lipid peroxidation, immunohistochemistry, reverse transcription, and polymerase chain reaction were used to evaluate the molecular mechanisms of CoQ10. Data are presented as mean ± SEM. Results:CoQ10 administration was associated with reduced loss of body weight compared with nontreated diabetic mice, without affecting blood glucose levels. Low dose and long-term administration of CoQ10 prevented the development of neuropathic pain. Treatment with CoQ10 produced a significant dose-dependent inhibition of mechanical allodynia and thermal hyperalgesia in diabetic mice. Dorsal root ganglia, sciatic nerve, and spinal cord tissues from diabetic mice demonstrated increased lipid peroxidation that was reduced by CoQ10 treatment. CoQ10 administration was also noted to reduce the proinflammatory factors in the peripheral and central nervous system. Conclusions:The results of this study support the hypothesis that hyperglycemia induced neuronal oxidative damage and reactive inflammation may be pathogenic in diabetic neuropathic pain. CoQ10 may be protective by inhibiting oxidative stress and reducing inflammation by down-regulating proinflammatory factors. These results suggest that CoQ10 administration may represent a low-risk, high-reward strategy for preventing or treating diabetic neuropathy.

Transgenic glial nuclear factor-kappa B inhibition decreases formalin pain in mice

In this work, we studied transgenic glial fibrillary acidic protein-IκB&agr;-dn mice that selectively inactivate the classical nuclear factor κB pathway by overexpressing the inhibitory protein of κB&agr; in astrocytes, under the control of glial fibrillary acidic protein promoter. We sought to determine if glial nuclear factor κB inhibition decreases formalin pain. Formalin testing was carried out on 25–35 g littermate adult male wild-type and transgenic C57Bl/6 mice. Formalin increased spinal cord c-Fos expression and glial fibrillary acidic protein immunostaining in both wild-type and transgenic mice. Transgenic glial fibrillary acidic protein-inhibitory protein of κB&agr;-dn mice had lower duration of formalin-induced paw-licking behavior. These data support a role of glial nuclear factor κB inhibition in reducing pain after peripheral nerve inflammation.

Diabetic neuropathic pain development in type 2 diabetic mouse model and the prophylactic and therapeutic effects of coenzyme Q10

UNLABELLED The early onset of type 2 diabetes mellitus (DM), driven by increasing obesity, is associated with peripheral neuropathy. Here, we characterize diabetic neuropathic pain in New Zealand obese diabetic mice (NZO/HILtJ) as a polygenic model of obesity with type 2 diabetes and investigate the role of coenzyme Q10 (CoQ10) in the prevention and treatment of diabetic neuropathic pain. Since the overexpression of mitogen-activated protein kinase (MAPK), nuclear factor-κB proteins (NF-Kb), toll-like receptor 4 (TLR4) and downstream cytokines (such as CCL2, CXCL10) are considered important factors contributing to the development of neuropathic pain, the expression of these factors and the inhibitory effects of CoQ10 were evaluated. NZO/HILtJ mice spontaneously developed type 2 DM and increased body mass with diabetic neuropathic pain. CoQ10 treatment decreased pain hypersensitivity and long-term supplementation prevented the development of diabetic neuropathic pain but did not attenuate diabetes. Spinal cord, blood serum, liver tissue, and dorsal root ganglia (DRG) from diabetic mice demonstrated increased lipid peroxidation, which was decreased by CoQ10 treatment. The percentage of positive neurons of p65 (the activated marker of NF-KB) and MAPK in DRG were significantly higher in DM mice compared to controls. However, CoQ10 treatment significantly decreased p65 and MAPK positive neurons in the DRG of DM mice. RT-PCR demonstrated that elevated levels of mRNA of CCL2, CXCL10 or TLR4 in the spinal cord of DM mice decreased significantly when DM mice were treated with CoQ10. CONCLUSION This model may be useful in understanding the mechanisms of neuropathic pain in type 2 DM induced neuropathic pain and may facilitate preclinical testing of therapies. CoQ10 may decrease oxidative stress in the central and peripheral nervous system by acting as an anti-oxidant and free-radical scavenger. These results suggest that CoQ10 might be a reasonable preventative strategy for long-term use and using CoQ10 treatment may be a safe and effective long-term approach in the treatment of diabetic neuropathy.

Glial NF-kappa B inhibition alters neuropeptide expression after sciatic nerve injury in mice

We utilized a transgenic mouse model where nuclear factor kappa B (NF-κB) is selectively inhibited in glial fibrillary acidic protein (GFAP) expressing cells. The transgene, GFAP-IκBα-dn, overexpresses a dominant negative form of the inhibitor of NF-κB (IκBα) under the control of the GFAP promoter. In the present work, we sought to understand the impact of glial NF-κB inhibition on the expression of pain mediating sensory neuropeptides galanin and calcitonin gene related peptide (CGRP) in a model of neuropathic pain in mice. Chronic constriction injury (CCI) of the left sciatic nerve was performed on wild type (WT) and GFAP-IκBα-dn transgenic mice. RT-PCR and immunohistological staining were performed in sciatic nerve and/or L4-L5 DRG tissue for galanin, CGRP and macrophage marker CD11b. GFAP-IκBα-dn mice had less mechanical and thermal hyperalgesia compared to WT mice post-CCI. After CCI, we observed galanin upregulation in DRG and sciatic nerve, which was less in GFAP-IκBα-dn mice. CGRP gene expression in the DRG increased transiently on day 1 post-CCI in WT but not in GFAP-IκBα-dn mice, and no evidence of CGRP upregulation in sciatic nerve post-CCI was found. After CCI, upregulation of CD11b in sciatic nerve was less in GFAP-IκBα-dn mice compared to WT mice, indicative of less macrophage infiltration. Our results showed that glial NF-κB inhibition reduces galanin and CGRP expression, which are neuropeptides that correlate with pain behavior and inflammation after peripheral nerve injury.

Carbonic anhydrase-8 regulates inflammatory pain by inhibiting the ITPR1-cytosolic free calcium pathway

Calcium dysregulation is causally linked with various forms of neuropathology including seizure disorders, multiple sclerosis, Huntington’s disease, Alzheimer’s, spinal cerebellar ataxia (SCA) and chronic pain. Carbonic anhydrase-8 (Car8) is an allosteric inhibitor of inositol trisphosphate receptor-1 (ITPR1), which regulates intracellular calcium release fundamental to critical cellular functions including neuronal excitability, neurite outgrowth, neurotransmitter release, mitochondrial energy production and cell fate. In this report we test the hypothesis that Car8 regulation of ITPR1 and cytoplasmic free calcium release is critical to nociception and pain behaviors. We show Car8 null mutant mice (MT) exhibit mechanical allodynia and thermal hyperalgesia. Dorsal root ganglia (DRG) from MT also demonstrate increased steady-state ITPR1 phosphorylation (pITPR1) and cytoplasmic free calcium release. Overexpression of Car8 wildtype protein in MT nociceptors complements Car8 deficiency, down regulates pITPR1 and abolishes thermal and mechanical hypersensitivity. We also show that Car8 nociceptor overexpression alleviates chronic inflammatory pain. Finally, inflammation results in downregulation of DRG Car8 that is associated with increased pITPR1 expression relative to ITPR1, suggesting a possible mechanism of acute hypersensitivity. Our findings indicate Car8 regulates the ITPR1-cytosolic free calcium pathway that is critical to nociception, inflammatory pain and possibly other neuropathological states. Car8 and ITPR1 represent new therapeutic targets for chronic pain.

Carbonic anhydrase-8 gene therapy inhibits the ITPR1-cytosolic free calcium pathway producing analgesia and anti-hyperalgesia

Carbonic anhydrase-8 gene therapy inhibits the ITPR1-cytosolic free calcium pathway producing analgesia and anti-hyperalgesia Roy C Levitt, Gerald Z Zhuang, Benjamin Keeler, Jeff Grant, Laura Bianchi, Eugene S Fu, Yan Ping Zhang, Diana M Erasso, Jian Guo Cui, Tim Wiltshire, Qiongzhen Li, Shuanglin Hao, Konstantinos D Sarantopoulos, Keith Candiotti, Shad B Smith, William Maixner, Luda Diatchenko, Eden R Martin

Acute Hypoglycemia Induces Painful Neuropathy and the Treatment of Coenzyme Q10

Diabetic neuropathic pain is reduced with tight glycemic control. However, strict control increases the risk of hypoglycemic episodes, which are themselves linked to painful neuropathy. This study explored the effects of hypoglycemia-related painful neuropathy. Pretreatment with coenzyme Q10 (CoQ10) was performed to explore the preventive effect of CoQ10 on hypoglycemia-related acute neuropathic pain. Two strains of mice were used and 1 unit/kg of insulin was given to induce hypoglycemia. Mechanical sensitivity of hindpaw withdrawal thresholds was measured using von Frey filaments. Blood glucose levels were clamped at normal levels by joint insulin and glucose injection to test whether insulin itself induced hypersensitivity. Results suggest that the increased mechanical sensitivity after insulin injection is related to decreased blood glucose levels. When blood glucose levels remained at a normal level by the linked administration of insulin and glucose, mice demonstrated no significant change in mechanical sensitivity. Pretreatment with CoQ10 prevented neuropathic pain and the expression of the stress factor c-Fos. These results support the concept that pain in the diabetic scenario can be the result of hypoglycemia and not insulin itself. Additionally, pretreatment with CoQ10 may be a potent preventive method for the development of neuropathic pain.