Model Of Diabetic Neuropathy Research Articles

Combination of Quercetin, Hirudin and Cinnamaldehyde Promotes Schwann Cell Differentiation and Myelination against High Glucose by Inhibiting ERK Signaling Pathway.

To investigate the therapeutic and synergistic effects of QHC (combination of quercetin (Q), hirudin (H) and cinnamaldehyd (C)) on Schwann cell differentiation and myelination against high glucose (HG) induced injury. Primary-culture Schwann cells exposed to HG (50 mmol/L) for 72 h and Schwann cell-dorsal root ganglion (DRG) neuron cocultures exposed to HG (50 mmol/L) for 7 days were employed as in vitro model of diabetic neuropathy. The cells were randomly divided into 10 groups: control (CON, 25 mmol/L glucose), HG (50 mmol/L glucose), HG plus 10 μmol/L quercetin (Q), HG plus 0.04 IU/mL hirudin (H), HG plus 100 nmol/L cinnamaldehyd (C), HG plus 10 μmol/L quercetin and 0.04 IU/mL hirudin (QH), HG plus 10 μmol/L quercetin and 50 nmol/L cinnamaldehyd (QC), HG plus 0.04 IU/mL hirudin and 50 nmol/L cinnamaldehyd (HC), HG plus 10 μmol/L quercetin, 0.04 IU/mL hirudin and 50 nmol/L cinnamaldehyd (QHC) or 10 μmol/L U0126. Cell differentiation was evaluated by periaxin immunofluorescence staining. The protein expression levels of myelin protein zero (P0), myelin basic protein (MBP), myelin-associated glycoprotein (MAG), extracellular signal-regulated kinase (ERK), p-ERK, p-c-Jun, c-Jun, notch intracellular domain (NICD) and the mRNA expression levels of P0, MBP, MAG, Krox-20, Notch1 and Jagged1 were detected by Western blotting and real-time quantitative PCR analysis. The secretion of ciliary neurotrophic factor (CNTF) was determined by enzyme-linked immunosorbent assay (ELISA). The number and length of the myelin segments were evaluated by MBP immunofluorescence staining. The expression and the location of p-ERK in cocultures were detected by MAG and p-ERK immunofluorescence double staining. Co-treatment with Q, C, H and their combination promoted Schwann cell differentiation, increased CNTF secretion, up-regulated the protein and mRNA expressions of myelin, and increased the number and length of the myelin segments (P<0.01 or P<0.05). In particular, the combination therapy of Q, H and C was superior to the respective monotherapy (P<0.01). Combination therapy of QHC exhibited higher inhibitory activities for ERK signaling related molecules than each monomer or the combination of the two monomers (P<0.01). QHC combination yielded synergy in promoting Schwann cell differentiation and myelination and the protective effect may involve in the inhibition of ERK signaling pathway, providing scientific evidence for better understanding of combination of Q, H and C in clinical applications.

Changes in expression of Kv7.5 and Kv7.2 channels in dorsal root ganglion neurons in the streptozotocin rat model of painful diabetic neuropathy

Diabetic peripheral neuropathic pain (DPNP), the most debilitating complication of diabetes mellitus, is resistant to current therapy. The pathogenesis of DPNP is still elusive, but several mechanisms have been proposed including abnormal hyperexcitability of dorsal root ganglion (DRG) neurons. The underlying molecular mechanisms of such aberrant hyperexcitability are incompletely understood. Using the streptozotocin (STZ) rat model of DPNP, we have recently provided evidence implicating neuronal Kv7 channels that normally exert a powerful stabilizing influence on neuronal excitability, in the abnormal hyperexcitability of DRG neurons and in pain hypersensitivity associated with DPNP. In the present immunohistochemical study, we sought to determine whether Kv7.2 and/or Kv7.5 channel expression is altered in DRG neurons in STZ rats. We found 35 days post-STZ: (1) a significant decrease in Kv7.5-immunoreactivity in small (<30 μm) DRG neurons (both IB4 positive and IB4 negative) and medium-sized (30−40 μm) neurons, and (2) a significant increase in Kv7.2-immunoreactivity in small (<30 μm) neurons, and a non-significant increase in medium/large neurons. The decrease in Kv7.5 channel expression in small and medium-sized DRG neurons in STZ rats is likely to contribute to the mechanisms of hyperexcitability of these neurons and thereby to the resulting pain hypersensitivity associated with DPNP. The upregulation of Kv7.2 subunit in small DRG neurons may be an activity dependent compensatory mechanism to limit STZ-induced hyperexcitability of DRG neurons and the associated pain hypersensitivity. The findings support the notion that Kv7 channels may represent a novel target for DPNP treatment.

Gait analysis correlates mechanical hyperalgesia in a model of streptozotocin-induced diabetic neuropathy: A CatWalk dynamic motor function study

Peripheral neuropathy is a complication of diabetes commonly associated with pain and decline in motor compound action potential, leading to alterations in plantar pressure during gait. We identified motor impairments in streptozotocin (STZ)-induced diabetic neuropathic rats and correlated with mechanical withdrawal thresholds, establishing this correlation as a complementary method to investigate the development of chronic hyperalgesia in diabetic neuropathy. MethodsUNICAMP’s Ethics Committee (protocol number 3902-1) approved all experiments. Male Lewis rats (200-250 g) received a STZ-low-dose (25 mg/kg/day) (STZ group) or 0.1 M sodium citrate buffer (SCB, control group) once a day, during five consecutive days. Diabetic rats (250 mg/dL blood glucose) were submitted to electronic von Frey and CatWalk tests at 0, 7, 14, 21, and 28 days after treatment. ResultsSTZ, but not SCB, induced diabetes. After the 14th day (STZ)-induced diabetic rats showed mechanical hyperalgesia and a reduction in the hind limbs footprint intensities. At the 28th day, rats presented alterations in spatial parameters (Maximum Contact Area; Stride Length; Print Area), which showed a strong correlation with mechanical withdrawal thresholds (r2 = 0.97; 0.99, and 0.93, respectively). ConclusionsCorrelation between gait parameters and mechanical withdrawal thresholds enables a better experimental approach to evaluate the development of chronic hyperalgesia in the STZ-induced diabetes model. It allows a concise crosstalk of motor and sensorial functions, which are usually analyzed individually. CatWalk gait parameters can be used as a complementary tool to investigate the development of hyperalgesia in STZ-induced diabetic neuropathic rats.

Bergenin Reduces Experimental Painful Diabetic Neuropathy by Restoring Redox and Immune Homeostasis in the Nervous System.

Diabetic neuropathy is a frequent complication of diabetes. Symptoms include neuropathic pain and sensory alterations—no effective treatments are currently available. This work characterized the therapeutic effect of bergenin in a mouse (C57/BL6) model of streptozotocin-induced painful diabetic neuropathy. Nociceptive thresholds were assessed by the von Frey test. Cytokines, antioxidant genes, and oxidative stress markers were measured in nervous tissues by ELISA, RT-qPCR, and biochemical analyses. Single (3.125–25 mg/kg) or multiple (25 mg/kg; twice a day for 14 days) treatments with bergenin reduced the behavioral signs of diabetic neuropathy in mice. Bergenin reduced both nitric oxide (NO) production in vitro and malondialdehyde (MDA)/nitrite amounts in vivo. These antioxidant properties can be attributed to the modulation of gene expression by the downregulation of inducible nitric oxide synthase (iNOS) and upregulation of glutathione peroxidase and Nrf2 in the nervous system. Bergenin also modulated the pro- and anti-inflammatory cytokines production in neuropathic mice. The long-lasting antinociceptive effect induced by bergenin in neuropathic mice, was associated with a shift of the cytokine balance toward anti-inflammatory predominance and upregulation of antioxidant pathways, favoring the reestablishment of redox and immune homeostasis in the nervous system. These results point to the therapeutic potential of bergenin in the treatment of painful diabetic neuropathy.

Blockade of CCR4 Diminishes Hypersensitivity and Enhances Opioid Analgesia – Evidence from a Mouse Model of Diabetic Neuropathy

Chemokine signaling has been implicated in the pathogenesis of diabetic neuropathy; however, the role of chemokine CC motif receptor 4 (CCR4) remains unknown. The goal was to examine the function of CCR4 in hypersensitivity development and opioid effectiveness in diabetic neuropathy. Streptozotocin (STZ; 200 mg/kg, intraperitoneally administered)-induced mouse model of diabetic neuropathy were used. An analysis of the mRNA/protein expression of CCR4 and its ligands was performed by qRT-PCR, microarray and/or Western blot methods. C021 (CCR4 antagonist), morphine and buprenorphine were injected intrathecally or intraperitoneally, and pain-related behavior was evaluated by the von Frey, cold plate and rotarod tests. We observed that on day 7 after STZ administration, the blood glucose level was increased, and as a consequence, hypersensitivity to tactile and thermal stimuli developed. In addition, we observed an increase in the mRNA level of CCL2 but not CCL17/CCL22. The microarray technique showed that the CCL2 protein level was also upregulated. In naive mice, the pronociceptive effect of intrathecally injected CCL2 was blocked by C021, suggesting that this chemokine acts through CCR4. Importantly, our results provide the first evidence that in a mouse model of diabetic neuropathy, single intrathecal and intraperitoneal injections of C021 diminished neuropathic pain-related behavior in a dose-dependent manner and improved motor functions. Moreover, both single intrathecal and intraperitoneal injections of C021 enhanced morphine and buprenorphine effectiveness. These results reveal that pharmacological modulation of CCR4 may be a good potential therapeutic target for the treatment of diabetic neuropathy and may enhance the effectiveness of opioids.

The Preventive Effect of IL-1beta Antagonist on Diabetic Peripheral Neuropathy.

To investigate the relationship between Interleukin-1beta (IL-1beta) and diabetic peripheral neuropathy (DPN) using animal models. The rat model of diabetic neuropathy was induced by intraperitoneal injection of a single dose of streptozotocin (STZ) at 65mg/kg. Diabetic rats were randomly divided into two groups (10 each), one treated with 0.9% saline (DMS group) and the other with interleukin-1 receptor antagonist (IL-1RA) at 50mg/kg (DMI group) twice a day for 5 weeks. Ten normal rats matched for weight, age and sex served as normal controls (Con group) and were treated with saline. Morphologic studies of sciatic nerves were achieved using light and transmission electron microscopy. Transmission electron microscopy of the sciatic nerve showed the ultrastructure of myelin and the axon in the IL-1RA group was highly protected compared to diabetic controls. High levels of circulating IL-1beta may be associated with the risk of DPN and anti-IL-1 treatment may provide a potential strategy for the prevention of diabetic neuropathy.

Cutaneous Aβ-Non-nociceptive, but Not C-Nociceptive, Dorsal Root Ganglion Neurons Exhibit Spontaneous Activity in the Streptozotocin Rat Model of Painful Diabetic Neuropathy in vivo.

Diabetic peripheral neuropathic pain (DPNP) is the most devastating complication of diabetes mellitus. Unfortunately, successful therapy for DPNP remains a challenge because its pathogenesis is still elusive. However, DPNP is believed to be due partly to abnormal hyperexcitability of dorsal root ganglion (DRG) neurons, but the relative contributions of specific functional subtypes remain largely unknown. Here, using the strepotozotocin (STZ) rat model of DPNP induced by a STZ injection (60 mg/kg, i.p), and intracellular recordings of action potentials (APs) from DRG neurons in anesthetized rats, we examined electrophysiological changes in C-and Aβ-nociceptive and Aβ-low threshold mechanoreceptive (LTM) neurons that may contribute to DPNP. Compared with control, we found in STZ-rats with established pain hypersensitivity (5 weeks post-STZ) several significant changes including: (a) A 23% increase in the incidence of spontaneous activity (SA) in Aβ-LTMs (but not C-mechanosensitive nociceptors) that may cause dysesthesias/paresthesia suffered by DPNP patients, (b) membrane hyperpolarization and a ∼85% reduction in SA rate in Aβ-LTMs by Kv7 channel activation with retigabine (6 mg/kg, i.v.) suggesting that Kv7/M channels may be involved in mechanisms of SA generation in Aβ-LTMs, (c) decreases in AP duration and in duration and amplitude of afterhyperpolarization (AHP) in C-and/or Aβ-nociceptors. These faster AP and AHP kinetics may lead to repetitive firing and an increase in afferent input to the CNS and thereby contribute to DPNP development, and (d) a decrease in the electrical thresholds of Aβ-nociceptors that may contribute to their sensitization, and thus to the resulting hypersensitivity associated with DPNP.

Antinociceptive effect of chrysin in diabetic neuropathy and formalin-induced pain models

ABSTRACT Chrysin, a natural flavonoid, is the main ingredient of many medicinal plants, which shows potent pharmacological properties. In the present study, the antinociceptive effects of chrysin were examined in ICR mice. Chrysin orally administered at the doses of from 10 to 100 mg/kg exerted the reductions of formalin-induced pain behaviors observed during the second phase in the formalin test in a dose-dependent manner. In addition, the antinociceptive effect of chrysin was further characterized in streptozotocin-induced diabetic neuropathy model. Oral administration chrysin caused reversals of decreased pain threshold observed in diabetic-induced peripheral neuropathy model. Intraperitoneally (i.p.) pretreatment with naloxone (a classic opioid receptor antagonist), but not yohimbine (an antagonist of α2-adrenergic receptors) or methysergide (an antagonist of serotonergic receptors), effectively reversed chrysin-induced antinociceptive effect in the formalin test. Moreover, chrysin caused a reduction of formalin-induced up-regulated spinal p-CREB level, which was also reversed by i.t. pretreated naloxone. Finally, chrysin also suppressed the increase of the spinal p-CREB level induced by diabetic neuropathy. Our results suggest that chrysin shows an antinociceptive property in formalin-induced pain and diabetic neuropathy models. In addition, spinal opioid receptors and CREB protein appear to mediate chrysin-induced antinociception in the formalin-induced pain model.

Agomelatine potentiates anti-nociceptive effects of morphine in a mice model for diabetic neuropathy: involvement of NMDA receptor subtype NR1 within the raphe nucleus and periaqueductal grey

ABSTRACT Objectives Opioid analgesics have been used for a long time in the treatment of acute and chronic pain. However, they have many side effects including tolerance development to a significant extent. Agomelatine, an atypical antidepressant, has been demonstrated to be effective in experimental studies on pain. However, the effect of agomelatine on morphine tolerance development and its mechanism of action are unknown. The antinociceptive effects of agomelatine, morphine and their combination were assessed in a mice model for painful diabetic neuropathy. The roles of glutamate ionotropic receptor N-methyl-D-aspartate (NMDA) type subunit-1 (GluN1) in raphe nucleus and periaqueductal gray (PAG) in the effect of agomelatine on neuropathic pain were also investigated in diabetic mice. Methods Agomelatine (10 mg/kg), morphine (10 mg/kg) and agomelatine + morphine were administered intraperitoneally for 15 consecutive days (twice per day), and the analgesic responses were assessed at days 1, 3, 6, 9, 12 and 15 in healthy and diabetic mice. Real time polymerase chain reaction (RT-PCR) was used to determine the changes in GluN1 expression. Results The tolerance development for morphine was evident, started at 6th day and remained thereafter, but not for agomelatine. GluN1 in raphe nucleus and PAG was upregulated in morphine treated but not in agomelatine-treated groups. Discussion The combination of agomelatine with morphine alone causes outlasting analgesic effects of repeated treatment, which can be interpreted as attenuated tolerance. Moreover, we also pointed out for the first time the modulatory effects of agomelatine on GluN1 expression in raphe nucleus and PAG after chronic morphine treatment. Abbreviations Ca2+: Calcium; D2DR: Dopamine D2 receptor; GAPDH: Glyceraldehyde-3-phosphate dehydrogenase; GluN1: Glutamate ionotropic receptor N-methyl-D-aspartate type subunit-1; 5-HT: 5-hydroxytryptamine; i.p.: intraperitoneal injection; MPE: Maximal possible effect; MT: Melatonin; NMDA: N-methyl-D-aspartate; NMDAR1: NMDA receptors-1; PAG: Periaqueductal grey; PKCγ: Protein kinase C gamma; RT-PCR: Real time polymerase chain reaction; STZ: Streptozotocin

Neuroprotective effect of diosgenin in a mouse model of diabetic peripheral neuropathy involves the Nrf2/HO-1 pathway

BackgroundDiabetic peripheral neuropathy (DPN) is one of the most common chronic complications of diabetes. Diosgenin is a natural steroidal saponin with a variety of beneficial effects, including antidiabetic effects, and is a raw material for the synthesis of carrier hormones. In our study, we aimed to assess the antioxidant effects of diosgenin in diabetic mice.MethodsMale C57 mice were fed a high-fat diet for 8 weeks and intraperitoneally injected with streptozotocin (STZ) at a dose of 100 mg/kg for 2 consecutive days. Eligible mice were divided into the normal control group (CON), diabetic group (DM), low-dose diosgenin (50 mg/kg) group (DIO50) and high-dose diosgenin (100 mg/kg) group (DIO100). Treatment was started 6 weeks after the induction of diabetes by STZ and continued for 8 weeks. Blood sugar and body weight were monitored dynamically. The behavioural effects of diosgenin were detected by a hot tail immersion test and paw tactile responses. HE staining was used to evaluate edema and degeneration of the sciatic nerve. The levels of SOD, MDA and GPx were tested according to the instructions of the respective kits. The levels of Nrf2, HO-1 and NQO1 were detected by immunofluorescence and Western blotting. Statistical analysis was performed using SPSS, and P < 0.05 was considered statistically significant.ResultsDiosgenin decreased the blood glucose levels and increased the body weight of diabetic mice. There was a significant increase in the tail withdrawal latency of diabetic animals, and mechanical hyperalgesia was significantly alleviated after diosgenin treatment. Histopathological micrographs of HE-stained sciatic nerves showed improvement after diosgenin treatment. Diosgenin attenuated the level of MDA but increased the activities of SOD and GPx. Diosgenin increased the expression of Nrf2, HO-1 and NQO1.ConclusionsOur results demonstrate that diosgenin can ameliorate behavioural and morphological changes in DPN by reducing oxidative stress. The Nrf2/HO-1 signalling pathway was involved in its neuroprotective effects.

Polarization of Adipose‐Derived Mesenchymal Stem/Stromal Cells via Toll‐Like Receptor Priming and Potential Synergy with Interleukin‐27 Delivery

Mesenchymal stem/stromal cells (MSCs) are a multipotent stromal cell population of interest for clinical applications in different diseases due to their ability to home to injury, differentiate into non‐hematopoietic cell lineages, and secrete paracrine factors with roles in tissue repair and immunomodulation. Although the manner in which MSCs modulate their many functions are incompletely understood, it has been shown that modulating toll‐like receptor (TLR) signaling can alter MSC activity. Promoting desired effector properties of MSCs represents a potential strategy for the use of these cells in different diseases. It has been reported in the past that MSCs derived from human bone marrow (hBM‐MSCs) can be “polarized”, by TLR priming, into two opposite immunomodulatory phenotypes. TLR4 priming leads to an immune activating MCS1 phenotype with roles in tumor repression, whereas TLR3 priming results in a strong immune suppressive MSC2 phenotype associated with support of tumor progression, and reduction of inflammation in a model of painful diabetic peripheral neuropathy. An alternative subpopulation of MSCs for therapeutic development are human adipose‐derived MSCs (hASCs), with the advantages of a relatively ease of isolation and higher abundance in comparison to their bone marrow counterpart. Although the MSC1 and MSC2 polarization have not been reported in hASCs, we hypothesize that hASCs can also undergo MSC1/MSC2 polarization upon short term and low level TLR stimulation in a similar way to hBM‐MSCs. To test our hypothesis, we primed hASCs with TLR3 or TLR4 agonists and evaluated gene expression changes expected to occur, based on the previously described hBM‐MSCs polarization phenotypes. Thus far, we have identified some similarities and differences in the gene expression signatures of the two populations upon TLR priming. Upregulation of IL‐8 was observed for TLR4 stimulated hASCs. CCL5, IDO and IP10 were upregulated in TLR3 stimulated hASCs. These results suggest that hASC polarization may occur in a similar way as in BM‐MSCs. However, no changes were observed in the expression of TGFβ or IL‐6, suggesting cell lineage differences in TLR stimulation response. In addition, some differences have been observed across different donor cells and priming conditions, which could contribute to the immunomodulatory inconsistencies reported in the literature. Current and future experiments aim to further characterize the gene expression signature changes on hASCs upon polarization as well as the key signaling pathways and molecules associated with these changes, and to determine if these polarized phenotypes affect the therapeutic potential of hASCs. Additional work aims to introduce episomal vectors expressing therapeutic genes such as interleukin (IL)‐27 in certain populations of polarized hASC for testing whether polarized hASC can act in synergy with IL‐27 in therapeutic applications such as prostate cancer and rheumatoid arthritis in vivo models of disease.Support or Funding InformationWe acknowledge support from the Purdue Research Foundation 19–20 PRF Research Grant 60000025 (MLF), and Purdue Doctoral Fellowship (CRC).

Simultaneous Target-Mediated Drug Disposition Model for Two Small-Molecule Compounds Competing for Their Pharmacological Target: Soluble Epoxide Hydrolase.

1-(1-propanoylpiperidin-4-yl)-3-[4-(trifluoromethoxy)phenyl]urea (TPPU) and 1-(4-trifluoro-methoxy-phenyl)-3-(1-cyclopropanecarbonyl-piperidin-4-yl)-urea (TCPU) are potent inhibitors of soluble epoxide hydrolase (sEH) that have much better efficacy in relieving nociceptive response than the Food and Drug Administration-approved drug gabapentin in a rodent model of diabetic neuropathy. Experiments conducted in sEH knockout mice or with coadministration of a potent sEH displacer demonstrated that the pharmacokinetics of TPPU and TCPU were influenced by the specific binding to their pharmacologic target sEH, a phenomenon known as target-mediated drug disposition (TMDD). To quantitatively characterize the complex pharmacokinetics of TPPU and TCPU and gain better understanding on their target occupancy, population pharmacokinetics analysis using a nonlinear mixed-effect modeling approach was performed in the current study. The final model was a novel simultaneous TMDD interaction model, in which TPPU and TCPU compete for sEH, with TCPU binding to an additional unknown target pool with larger capacity that we refer to as a refractory pool. The total amount of sEH enzyme in mice was predicted to be 16.2 nmol, which is consistent with the experimental value of 10 nmol. The dissociate rate constants of TPPU and TCPU were predicted to be 2.24 and 2.67 hours-1, respectively, which is close to the values obtained from in vitro experiments. Our simulation result predicted that 90% of the sEH will be occupied shortly after a low dose of 0.3 mg/kg TPPU administration, with ≥40% of sEH remaining to be bound with TPPU for at least 7 days. Further efficacy experiments are warranted to confirm the predicted target occupancy. SIGNIFICANCE STATEMENT: Although target-mediated drug disposition (TMDD) models have been well documented, most of them were established in a single compound scenario. Our novel model represents the first TMDD interaction model for two small-molecule compounds competing for the same pharmacological target.

Dorsal Root Ganglion Stimulation in Experimental Painful Diabetic Polyneuropathy: Delayed Wash-Out of Pain Relief After Low-Frequency (1Hz) Stimulation

Up until now there is little data about the pain relieving effect of different frequency settings in DRGS. The aim of this study was to compare the pain relieving effect of DRGS at low-, mid-, and high-frequencies and Sham-DRGS in an animal model of painful diabetic neuropathy (PDPN). Diabetes mellitus was induced by an intraperitoneal injection of streptozotocin in 8-week-old female Sprague-Dawley rats (n = 24; glucose ≥15 mmol/L: n = 20; mechanical hypersensitivity: n = 15). Five weeks later, a DRGS device was implanted at the L5 DRG. Ten animals were included for stimulation, alternating 30 minutes of low (1 Hz)-, mid (20 Hz)-, and high (1000 Hz)-frequencies and Sham-DRGS during four days, with a pulse width of 0.2 msec (average amplitude: 0.19 ± 0.01 mA), using a randomized cross-over design. The effect on mechanical hypersensitivity of the hind paw to von Frey filaments was evaluated. All DRGS frequencies resulted in a complete reversal of mechanical hypersensitivity and "a clinically relevant reduction" was achieved in 70-80% of animals. No significant differences in maximal pain relieving effect were found between the different frequency treatments (p = 0.24). Animals stimulated at 1000 and 20 Hz returned to baseline mechanical hypersensitivity values 15 and 30 min after stimulation cessation, respectively, while animals stimulated at 1 Hz did not. These results show that DRGS is equally effective when applied at low-, mid-, and high-frequency in an animal model of PDPN. However, low-frequency-(1 Hz)-DRGS resulted in a delayed wash-out effect, which suggests that this is the most optimal frequency for pain therapy in PDPN as compared to mid- and high-frequency.

Nonlinear Relation Between Burst Dorsal Root Ganglion Stimulation Amplitude and Behavioral Outcome in an Experimental Model of Painful Diabetic Peripheral Neuropathy

Background and objectiveDorsal root ganglion stimulation (DRGS) has recently emerged as a neuromodulation modality in the treatment of chronic neuropathic pain. The objective of this study was to compare the efficacy of different Burst‐DRGS amplitudes in an experimental model of painful diabetic peripheral neuropathy (PDPN).MethodsDiabetes mellitus was induced in female Sprague–Dawley rats by intraperitoneal injection of streptozotocin (STZ, n = 28). Animals were tested for mechanical hypersensitivity (von Frey paw withdrawal test) before, and four weeks after STZ injection. PDPN rats (n = 13) were implanted with a unilateral bipolar electrode at the L5 DRG. Animals received Burst‐DRGS at 0%, 10%, 33%, 50%, 66%, and 80% of motor threshold (MT) in a randomized crossover design on post‐implantation days 2–7 (n = 9). Mechanical hypersensitivity was assessed before stimulation onset, 15 and 30 min during stimulation, and 15 and 30 min after stimulation.ResultsBurst‐DRGS at amplitudes of 33%, 50%, 66%, and 80% MT resulted in significant attenuation of STZ‐induced mechanical hypersensitivity at 15 and 30 min during stimulation, as well as 15 min after cessation of stimulation. No effect on mechanical hypersensitivity was observed for Burst‐DRGS at 0% MT and 10% MT. Optimal pain relief and highest responder rates were achieved with Burst‐DRGS at 50–66% MT, with an estimated optimum at 52% MT.ConclusionOur findings indicate a nonlinear relationship between Burst‐DRGS amplitude and behavioral outcome, with an estimated optimal amplitude of 52% MT. Further optimization and analysis of DRGS driven by insights into the underlying mechanisms related to the various stimulation paradigms is warranted.

Diet-Induced Rodent Models of Diabetic Peripheral Neuropathy, Retinopathy and Nephropathy.

Unhealthy dietary habits are major modifiable risk factors for the development of type 2 diabetes mellitus, a metabolic disease with increasing prevalence and serious consequences. Microvascular complications of diabetes, namely diabetic peripheral neuropathy (DPN), retinopathy (DR), and nephropathy (DN), are associated with high morbidity rates and a heavy social and economic burden. Currently, available therapeutic options to counter the evolution of diabetic microvascular complications are clearly insufficient, which strongly recommends further research. Animal models are essential tools to dissect the molecular mechanisms underlying disease progression, to unravel new therapeutic targets, as well as to evaluate the efficacy of new drugs and/or novel therapeutic approaches. However, choosing the best animal model is challenging due to the large number of factors that need to be considered. This is particularly relevant for models induced by dietary modifications, which vary markedly in terms of macronutrient composition. In this article, we revisit the rodent models of diet-induced DPN, DR, and DN, critically comparing the main features of these microvascular complications in humans and the criteria for their diagnosis with the parameters that have been used in preclinical research using rodent models, considering the possible need for factors which can accelerate or aggravate these conditions.

N-Acetylcysteine causes analgesia in a mouse model of painful diabetic neuropathy.

N-Acetylcysteine, one of the most prescribed antioxidant drugs, enhances pain threshold in rodents and humans by activating mGlu2 metabotropic glutamate receptors. Here, we assessed the analgesic activity of N-acetylcysteine in the streptozotocin model of painful diabetic neuropathy and examined the effect of N-acetylcysteine on proteins that are involved in mechanisms of nociceptive sensitization. Mice with blood glucose levels ≥250 mg/dl in response to a single intraperitoneal (i.p.) injection of streptozotocin (200 mg/kg) were used for the assessment of mechanical pain thresholds. Systemic treatment with N-acetylcysteine (100 mg/kg, i.p., either single injection or daily injections for seven days) caused analgesia in diabetic mice. N-acetylcysteine-induced analgesia was abrogated by the inhibitors, sulfasalazine (8 mg/kg, i.p.), erastin (30 mg/kg, i.p.), and sorafenib (10 mg/kg, i.p.), or by the mGlu2/3 receptor antagonist, LY341495 (1 mg/kg, i.p.). Repeated administrations of N-acetylcysteine in diabetic mice reduced ERK1/2 phosphorylation in the dorsal region of the lumbar spinal cord. The analgesic activity of N-acetylcysteine was occluded by the MEK inhibitor, PD0325901 (25 mg/kg, i.p.), the TRPV1 channel blocker, capsazepine (40 mg/kg, i.p.), or by a cocktail of NMDA and mGlu5 metabotropic glutamate receptor antagonists (memantine, 25 mg/kg, plus MTEP, 5 mg/kg, both i.p.). These findings offer the first demonstration that N-acetylcysteine relieves pain associated with diabetic neuropathy and holds promise for the use of N-acetylcysteine as an add-on drug in diabetic patients.

Roles of TRPA1 in Painful Dysesthesia

Dysesthesia is an unpleasant abnormal sensation, often accompanied by pain, paresthesia (abnormal sensation), and numbness (decrease or loss of sensation). Dysesthesia has been associated with various conditions, although its underlying mechanisms are largely unknown. This study assessed the roles of transient receptor potential ankyrin 1 (TRPA1) in dysesthesia by utilizing three animal models of dysesthesia characterized by reductions in blood flow to the skin: a transient hindlimb ischemia/reperfusion model, characterized by spontaneous licking and tactile hypoesthesia of the ischemic hindpaw; a streptozotocin-induced diabetic neuropathy model in mice, characterized by cold hypersensitivity, which is likely parallel to the reduced skin blood flow of the hindpaw; and a hindlimb ischemia model. TRPA1 inhibition or deficiency blocked spontaneous licking in the transient hindlimb ischemia/reperfusion model and cold hypersensitivity in the diabetic mouse model mice. Consistent with these results, the nocifensive behaviors induced by intraplantar injection of a TRPA1 agonist were enhanced in the diabetic neuropathy and hindlimb ischemia models. Hypoxia enhanced H2O2-induced TRPA1 responses in human TRPA1-expressing cells and cultured mouse dorsal root ganglion neurons, with this hypoxia-induced TRPA1 sensitization to H2O2 being associated with hypoxia-induced inhibition of the hydroxylation of prolyl hydroxylases. These results suggest that dysesthesia following blood flow reduction is caused by the activation of TRPA1 sensitized by hypoxia and that hypoxia-induced TRPA1 sensitization plays a pivotal role in painful dysesthesia induced by peripheral blood flow reduction.

Experimental Diabetic Neuropathy: A Newer Insight to Diet Induced Models in Animals

Type 2 diabetes mellitus is the most prevalent form of diabetes and accounts for approximately 95% of diabetic cases. One of the most common and debilitating complications associated with diabetes is diabetic neuropathy (DN) which affects about 10% of newly diagnosed diabetes patients and more than 50% of patients with longstanding diabetes. Animal models are widely used in the research of diabetes and its complications, where rats and mice are most commonly used animals for these experimental models. The present review summarizes the current understanding of the metabolic profile and pathology involved in the development of diabetic neuropathy. Different models of diabetic neuropathy in an experimental setup are described and finally an insight of diet induced models of DN is highlighted. Recent studies have described that diet induced models are beneficial over chemical-mediated and genetically-mediated models in the development of type 2 diabetic neuropathy. The High Fat Diet (HFD) combined with low doses of streptozotocin is the most successful and well established model of diet induced neuropathy, which mimics natural progress of diabetes development as well as metabolic features in human type 2 diabetes HFD causes obesity and insulin resistance in peripheral tissues due to lipotoxicity, while, low dose of streptozotocin induces mild defect in insulin secretion.

Effect of Four-week Honey-treatment on Blood Glucose and Lipid Profile in an Experimental Model of Diabetic Neuropathy

Background and Aim: Honey is a natural substance with various medicinal properties which include antibacterial, antihypertensive, hepatoprotective, hypoglycemic and antioxidant effects. However, the role of honey in the management of neuropathic morbidities in diabetic neuropathy has not been studied. Especially, the effect of honey on fasting blood glucose and lipid profile in diabetic neuropathy has not been assessed. Therefore, in the present study we have assessed the effect of honey treatment on blood glucose and lipid profile in animal diabetic neuropathy model. Methods: Twenty-four healthy male Wistar albino rats of 10-12 weeks age, weighing 200±30 g were obtained from JIPMER central animal house. After one week of habituation, rats were divided into three groups randomly. After developing diabetic neuropathy, Fasting Blood Glucose (FBG) and lipid profile, Aspartate Transaminase (AST) and Alanine transaminase (ALT) were measured in blood sample in the rats before and after honey treatment. Results: Significant increase in FBG, AST, ALT and lipid profile except HDL cholesterol was seen in diabetic neuropathy when compared with normal healthy rats. There was a significant reduction in all these parameters except HDL-cholesterol after four-week honey treatment in comparison with diabetic neuropathy rats without treatment. Conclusion: Honey, given at a dose of 0.5 gm/kg BW for four weeks is effective in reducing blood glucose, atherogenic index and lipid profile and improving liver functions in albino rats.

Effects of Citrullus colocynthis L. in a rat model of diabetic neuropathy

ObjectiveThis study investigated the biochemical, histopathological and physiological effects of Citrullus colocynthis on peripheral neuropathy in rats with streptozotocin (STZ)-induced diabetes. MethodsSeventy adult male Sprague-Dawley rats were included in the present study. Diabetes was induced in 60 rats, with a single intraperitoneal injection of STZ (65 mg/kg). After 4 weeks, the diabetic rats were assessed for neuropathy. Then, the diabetic rats with neuropathy were randomly divided into 6 groups for a 4-week treatment with gabapentin, oral administration of C. colocynthis fruit pulp powder (100 and 300 mg/kg per day), topical preparations as oil-based solution and ointment, or placebo. Changes in metabolic, physiological, biochemical and histological parameters were considered as treatment outcomes. ResultsMetabolic outcomes (body weight and blood glucose level) were improved in the C. colocynthis-treated groups as compared to placebo. Tail-flick and hot-plate tests also had lower latency in the C. colocynthis-treated groups. Measurement of oxidative stress markers (malondialdehyde, superoxide dismutase and catalase) showed the antioxidant effect of C. colocynthis. Histological evaluation of the sciatic nerve showed that C. colocynthis decreased the number of demyelinated and degenerated nerve fibers. Among the C. colocynthis-treated groups, the one receiving 100 mg/kg power per day orally had the best treatment outcomes. ConclusionThe present study showed that C. colocynthis fruit, through its antioxidant and hypoglycemic activities, has a positive effect in the treatment of diabetic neuropathy.