Roles of TRPV1 and neuropeptidergic receptors in dorsal root reflex-mediated neurogenic inflammation induced by intradermal injection of capsaicin

Background

Acute cutaneous neurogenic inflammation initiated by activation of transient receptor potential vanilloid-1 (TRPV₁) receptors following intradermal injection of capsaicin is mediated mainly by dorsal root reflexes (DRRs). Inflammatory neuropeptides are suggested to be released from primary afferent nociceptors participating in inflammation. However, no direct evidence demonstrates that the release of inflammatory substances is due to the triggering of DRRs and how activation of TRPV₁ receptors initiates neurogenic inflammation via triggering DRRs.

Results

Here we used pharmacological manipulations to analyze the roles of TRPV₁ and neuropeptidergic receptors in the DRR-mediated neurogenic inflammation induced by intradermal injection of capsaicin. The degree of cutaneous inflammation in the hindpaw that followed capsaicin injection was assessed by measurements of local blood flow (vasodilation) and paw-thickness (edema) of the foot skin in anesthetized rats. Local injection of capsaicin, calcitonin gene-related peptide (CGRP) or substance P (SP) resulted in cutaneous vasodilation and edema. Removal of DRRs by either spinal dorsal rhizotomy or intrathecal administration of the GABA_A receptor antagonist, bicuculline, reduced dramatically the capsaicin-induced vasodilation and edema. In contrast, CGRP- or SP-induced inflammation was not significantly affected after DRR removal. Dose-response analysis of the antagonistic effect of the TRPV₁ receptor antagonist, capsazepine administered peripherally, shows that the capsaicin-evoked inflammation was inhibited in a dose-dependent manner, and nearly completely abolished by capsazepine at doses between 30–150 μg. In contrast, pretreatment of the periphery with different doses of CGRP_8–37 (a CGRP receptor antagonist) or spantide I (a neurokinin 1 receptor antagonist) only reduced the inflammation. If both CGRP and NK₁ receptors were blocked by co-administration of CGRP_8–37 and spantide I, a stronger reduction in the capsaicin-initiated inflammation was produced.

Conclusion

Our data suggest that 1) the generation of DRRs is critical for driving the release of neuropeptides antidromically from primary afferent nociceptors; 2) activation of TRPV₁ receptors in primary afferent nociceptors following intradermal capsaicin injection initiates this process; 3) the released CGRP and SP participate in neurogenic inflammation.     

Decreased nitric oxide- and axon reflex-mediated cutaneous vasodilation with age during local heating.

Cutaneous vasodilation is reduced in healthy older vs. young subjects; however, the mechanisms that underlie these age-related changes are unclear. Our goal in the present study was to determine the role of nitric oxide (NO) and the axon reflexes in the skin blood flow (SkBF) response to local heating with advanced age. We placed two microdialysis fibers in the forearm skin of 10 young (Y; 22 +/- 2 yr) and 10 older (O; 77 +/- 5 yr) men and women. SkBF over each site was measured by laser-Doppler flowmetry (LDF; Moor DRT4). Both sites were heated to 42 degrees C for ~60 min while 10 mM N(G)-nitro-L-arginine methyl ester (L-NAME) was infused throughout the protocol to inhibit NO synthase (NOS) in one site and 10 mM L-NAME was infused after 40 min of local heating in the second site. Data were expressed as a percentage of maximal vasodilation (%CVC(max); 28 mM nitroprusside infusion). Local heating before L-NAME infusion resulted in a significantly reduced initial peak (Y: 61 +/- 2%CVC(max) vs. O: 46 +/- 4%CVC(max)) and plateau (Y: 93 +/- 2%CVC(max) vs. O: 82 +/- 5%CVC(max)) CVC values in older subjects (P < 0.05). When NOS was inhibited after 40 min of heating, CVC declined to the same value in the young and older groups. Thus the overall contribution of NO to the plateau phase of the SkBF response to local heating was less in the older subjects. The initial peak response was significantly lower in the older subjects in both microdialysis sites (Y: 52 +/- 4%CVC(max) vs. O: 38 +/- 5%CVCmax; P < 0.05). These data suggest that age-related changes in both axon reflex-mediated and NO-mediated vasodilation contribute to attenuated cutaneous vasodilator responses in the elderly.     

Diabetic cardiomyopathy: recent evidence from mouse models of type 1 and type 2 diabetes

Diabetic cardiomyopathy is defined as ventricular dysfunction of the diabetic heart in the absence of coronary artery disease. With the use of both in vivo and ex vivo techniques to assess cardiac phenotype, reduced contractile performance can be observed in experiments with mouse models of both type 1 (insulin-deficient) and type 2 (insulin-resistant) diabetes. Both systolic dysfunction (reduced left ventricular pressures and decreased cardiac output) and diastolic dysfunction (impaired relaxation) is observed in diabetic hearts, along with enhanced susceptibility to ischemic injury. Metabolism is also altered in diabetic mouse hearts: glucose utilization is reduced and fatty acid utilization is increased. The use of genetically engineered mice has provided a powerful experimental approach to test mechanisms that may be responsible for the deleterious effects of diabetes on cardiac function.     

An association between fibrinogen gene polymorphisms and diabetic peripheral neuropathy in young patients with type 1 diabetes

Molecular Biology Reports - In complex etiopathogenesis of diabetic peripheral neuropathy (DPN), hemostatic dysfunction and subclinical inflammation play a possible role. Fibrinogen is involved in...     

Diabetic state affects the innervation of gut in an animal model of human type 1 diabetes

Gastrointestinal symptoms in diabetic patients are commonplace, and are believed to be due, at least partly, to neuropathy of the gut. In the present study, therefore, some important neurotransmitters in the myenteric plexus were investigated in non-obese diabetic mice, an animal model of human type 1 diabetes. For this purpose, immunocytochemistry was applied on sections from antrum, duodenum and colon, subsequently quantified by computerized image analysis. Whereas the number of vasoactive intestinal peptide (VIP)-positive neurons was increased in antral myenteric ganglia of diabetic mice, there was a decreased density of nerve fibres in muscularis propria. No difference was seen in the VIP of duodenum and colon. Acetylcholine-containing nerve fibres showed an increased volume density in muscularis propria of antrum and duodenum, but a decreased density in colon of diabetic mice, as compared with controls. There was a decreased number of neurons containing nitric oxide synthase (NOS) in myenteric ganglia of antrum and duodenum. No difference was seen in density of NOS-containing nerve fibres in muscularis propria. There was no difference regarding neuropeptide Y (NPY) and galanin between diabetic and control mice; nor was there any difference between pre-diabetic NOD mice and controls regarding all bioactive substances investigated. It is concluded that the diabetic state affects the innervation of gut in this animal model. The present findings may be of some relevance to the gastrointestinal symptoms seen in patients with diabetes.     

'Sensing' autoimmunity in type 1 diabetes

Type 1 diabetes (T1D) results from autoimmune-mediated loss of insulin-producing beta-cells. Recent findings suggest that the events controlling T1D development are not only immunological, but also neuronal in nature. In the non-obese diabetic (NOD) mouse model of T1D, a mutant sensory neuron channel, TRPV1, initiates chronic, progressive beta-cell stress, inducing islet cell inflammation. This novel mechanism of organ-specific damage requires a permissive, autoimmune-prone host, but ascribes tissue specificity to the local secretory dysfunction of sensory afferent neurons. In NOD mice, normalizing this neuronal function by administration of the neurotransmitter substance P clears islet cell inflammation, reduces insulin resistance and restores normoglycemia. Here, we discuss this neuro-immuno-endocrine model, its implications and the involvement of sensory neurons in other autoimmune disorders. These developments might provide novel neuronal-based therapeutic interventions, particularly in diabetes.     

Multifactorial inheritance in type 1 diabetes

To date, twelve separate chromosome regions have been implicated in the development of human type 1 (insulin-dependent) diabetes mellitus. The major disease locus, IDDM1 in the major histocompatibility complex (MHC) on chromosome 6p21, accounts for about 35% of the observed familial clustering and its contribution to disease susceptibility is likely to involve polymorphic residues of class II molecules in T-cell-mediated autoimmunity. IDDM2 is encoded by a minisatellite locus embedded in the 5 regulatory region of the insulin gene. Familial clustering of disease can be explained by the sharing of alleles of at least 10 loci. IDDM1 and IDDM2 interact epistatically. For a multifactorial disease, such as type 1 diabetes, important information concerning the pathways and mechanisms involved can be gained from examining such interactions between loci, using methods that simultaneously take account of the joint effects of the various underlying genetic components.     

Immunogenetics of type 1 diabetes

The T-cell mediated autoimmune process that destroys pancreatic beta cells in type 1 diabetes (T1D) is a complex phenotype influenced by multiple genetic and environmental factors. Human leukocyte antigen (HLA) accounts for about half of the genetic susceptibility, through a large variety of protective and predisposing haplotypes. Other important loci associated with T1D, with much smaller effects than HLA, include the insulin variable number of tandem repeats, PTPN22, and CTLA-4. Detecting the association and confirming it beyond doubt is only the first step. Identifying the functional variant from among a block of polymorphisms in tight linkage disequilibrium and determining its biological consequences can be an even more challenging task. It is hoped that the identification of additional loci and functional analysis of known ones, no matter how small each individual effect is, will provide: (1) pathophysiological insights necessary for the development of preventive interventions; (2) risk prediction to identify individuals that can benefit from them, and (3) potentially, identification of distinct subgenotypes, with different immune dysregulation pathways leading to the common disease phenotype that may respond to different preventive interventions.     

Distinguishing type 2 diabetes from type 1 diabetes in African American and Hispanic American pediatric patients

Objective

To test the hypothesis that clinical observations made at patient presentation can distinguish type 2 diabetes (T2D) from type 1 diabetes (T1D) in pediatric patients aged 2 to 18.

Subjects and Methods

Medical records of 227 African American and 112 Hispanic American pediatric patients diagnosed as T1D or T2D were examined to compare parameters in the two diseases. Age at presentation, BMI z-score, and gender were the variables used in logistic regression analysis to create models for T2D prediction.

Results

The regression-based model created from African American data had a sensitivity of 92% and a specificity of 89%; testing of a replication cohort showed 91% sensitivity and 93% specificity. A model based on the Hispanic American data showed 92% sensitivity and 90% specificity. Similarities between African American and Hispanic American patients include: (1) age at onset for both T1D and T2D decreased from the 1980s to the 2000s; (2) risk of T2D increased markedly with obesity. Racial/ethnic-specific observations included: (1) in African American patients, the proportion of females was significantly higher than that of males for T2D compared to T1D (p<0.0001); (2) in Hispanic Americans, the level of glycated hemoglobin (HbA1c) was significantly higher in T1D than in T2D (p<0.002) at presentation; (3) the strongest contributor to T2D risk was female gender in African Americans, while the strongest contributor to T2D risk was BMI z-score in Hispanic Americans.

Conclusions

Distinction of T2D from T1D at patient presentation was possible with good sensitivity and specificity using only three easily-assessed variables: age, gender, and BMI z-score. In African American pediatric diabetes patients, gender was the strongest predictor of T2D, while in Hispanic patients, BMI z-score was the strongest predictor. This suggests that race/ethnic specific models may be useful to optimize distinction of T1D from T2D at presentation.     

Quantifying macrophage defects in type 1 diabetes

Macrophages from animals prone to autoimmune (type 1) diabetes differ from those of diabetes-resistant animals in processing and clearing apoptotic cells. Using in vitro time-course assays of the number of engulfed apoptotic cells observed within macrophages, we quantified these differences in non-obese diabetic (NOD) versus Balb/c mice. Simple models lead to several elementary parameter estimation techniques. We used these to compute approximate rates of macrophage engulfment and digestion of apoptotic cells from basic features of the data (such as initial rise-times, phagocytic index and percent phagocytosis). Combining these estimates with full fitting of a sequence of model variants to the data, we find that macrophages from normal (Balb/c) mice engulf apoptotic cells up to four times faster than macrophages from the diabetes-prone (NOD) mice. Further, Balb/c macrophages appear to undergo an activation step before achieving their high engulfment rate. In NOD macrophages, we did not see evidence for this activation step. Rates of digestion of engulfed apoptotic cells by macrophages are similar in both types. Since macrophage clearance is an important mechanism of disposal of self-antigen, these macrophage defects could potentially be a factor in predisposition to type 1 diabetes.     

The insulin gene in type 1 diabetes

Insulin is a key autoantigen in the autoimmune process leading to the development of type 1 diabetes. Recent studies in both humans and mice have shown that variation in the expression of the insulin gene, in the thymus rather than the pancreas, contributes to disease susceptibility by affecting self-tolerance to insulin. These findings have brought about a paradigm-shift in our understanding of self-tolerance and autoimmunity to molecules with tissue-restricted expression, which are often the target of autoimmune disease.