Injection Of Nerve Growth Factor Research Articles

Nerve growth factor induces sensitization of nociceptors without evidence for increased intraepidermal nerve fiber density

Nerve growth factor (NGF) is involved in the long-term sensitization of nociceptive processing linked to chronic pain. Functional and structural (“sprouting”) changes can contribute. Thus, humans report long-lasting hyperalgesia to mechanical and electrical stimulation after intradermal NGF injection and NGF-induced sprouting has been reported to underlie cancer bone pain and visceral pain. Using a human-like animal model we investigated the relationship between the structure and function of unmyelinated porcine nociceptors 3weeks after intradermal NGF treatment. Axonal and sensory characteristics were studied by in vivo single-fiber electrophysiology and immunohistochemistry. C fibers recorded extracellularly were classified based on mechanical response and activity-dependent slowing (ADS) of conduction velocity. Intraepidermal nerve fiber (IENF) densities were assessed by immunohistochemistry in pigs and in human volunteers using the same NGF model. NGF increased conduction velocity and reduced ADS and propagation failure in mechano-insensitive nociceptors. The proportion of mechano-sensitive C nociceptors within NGF-treated skin areas increased from 45.1% (control) to 71% and their median mechanical thresholds decreased from 40 to 20mN. After NGF application, the mechanical receptive fields of nociceptors increased from 25 to 43mm2. At the structural level, however, IENF density was not increased by NGF. In conclusion, intradermal NGF induces long-lasting axonal and mechanical sensitization in porcine C nociceptors that corresponds to hyperalgesia observed in humans. Sensitization is not accompanied by increased IENF density, suggesting that NGF-induced hyperalgesia might not depend on changes in nerve fiber density but could be linked to the recruitment of previously silent nociceptors.

Repeated intramuscular injections of nerve growth factor induced progressive muscle hyperalgesia, facilitated temporal summation, and expanded pain areas

Intramuscular injection of nerve growth factor (NGF) is known to induce deep-tissue mechanical hyperalgesia. In this study it was hypothesised that daily intramuscular injections of NGF produce a progressive manifestation of soreness, mechanical hyperalgesia, and temporal summation of pain. In a double-blind placebo-controlled design, 12 healthy subjects were injected on 3days with NGF into the tibialis anterior muscle and with isotonic saline on the contralateral side. Assessments were performed before and after the injections on days 0, 1, and 2, and repeated on days 3, 6, and 10. The self-perceived muscle soreness was assessed on a Likert scale. Computer-controlled pressure algometry was used to assess the pressure pain thresholds (PPTs). Temporal summation of pain after repeated pressure stimulations was assessed by computer-controlled pressure algometry. The pain distribution following painful pressure stimulation was also recorded. Compared with baseline and isotonic saline, the NGF injections caused (P<0.05): (1) progressively increasing soreness scores from 3hours after the first injection until day 2, after which it remained increased; (2) decreased PPTs at days 1 to 3; (3) facilitated temporal summation of pressure pain at days 1 to 10; and (4) enlarged pressure-induced pain area after the injection on day 1 to day 6. The daily injections of NGF produced a progressive manifestation of muscle soreness, mechanical hyperalgesia, temporal summation of pressure pain, and pressure-induced pain distribution. These data illustrate that the prolonged NGF application affects peripheral and central mechanisms and may reflect process in musculoskeletal pain conditions.

Augmented pain behavioural responses to intra-articular injection of nerve growth factor in two animal models of osteoarthritis

ObjectivesNerve growth factor (NGF) is a promising analgesic target, particularly in osteoarthritis (OA) where existing therapies are inadequate. We hypothesised that pain responses to NGF are increased in OA joints....

TRPV1 and TRPV4 Play Pivotal Roles in Delayed Onset Muscle Soreness

Unaccustomed strenuous exercise that includes lengthening contraction (LC) often causes tenderness and movement related pain after some delay (delayed-onset muscle soreness, DOMS). We previously demonstrated that nerve growth factor (NGF) and glial cell line-derived neurotrophic factor (GDNF) are up-regulated in exercised muscle through up-regulation of cyclooxygenase (COX)-2, and they sensitized nociceptors resulting in mechanical hyperalgesia. There is also a study showing that transient receptor potential (TRP) ion channels are involved in DOMS. Here we examined whether and how TRPV1 and/or TRPV4 are involved in DOMS. We firstly evaluated a method to measure the mechanical withdrawal threshold of the deep tissues in wild-type (WT) mice with a modified Randall-Selitto apparatus. WT, TRPV1−/− and TRPV4−/− mice were then subjected to LC. Another group of mice received injection of murine NGF-2.5S or GDNF to the lateral gastrocnemius (LGC) muscle. Before and after these treatments the mechanical withdrawal threshold of LGC was evaluated. The change in expression of NGF, GDNF and COX-2 mRNA in the muscle was examined using real-time RT-PCR. In WT mice, mechanical hyperalgesia was observed 6–24 h after LC and 1–24 h after NGF and GDNF injection. LC induced mechanical hyperalgesia neither in TRPV1−/− nor in TRPV4−/− mice. NGF injection induced mechanical hyperalgesia in WT and TRPV4−/− mice but not in TRPV1−/− mice. GDNF injection induced mechanical hyperalgesia in WT but neither in TRPV1−/− nor in TRPV4−/− mice. Expression of NGF and COX-2 mRNA was significantly increased 3 h after LC in all genotypes. However, GDNF mRNA did not increase in TRPV4−/− mice. These results suggest that TRPV1 contributes to DOMS downstream (possibly at nociceptors) of NGF and GDNF, while TRPV4 is located downstream of GDNF and possibly also in the process of GDNF up-regulation.

Effect of lowering intraocular pressure treatment on ocular hemodynamics in patients with nonarteritic anterior ischemic optic neuropathy

Objective To observe the effect of lowering intraocular pressure(IOP) treatment on ocular hemodynamics in patients with nonarteritic anterior ischemic optic neuropathy (NAION).Methods A total of 68 patients with NAION (68 eyes) were enrolled in this study.The patients were randomly divided into treatment group (38 eyes of 38 patients) and control group (30 eyes of 30 patients).All the patients were received methylprednisolone pulse therapy (200 mg,three days),vasodilator therapy with intravenous infusion of Xueshuantong solution (300 mg),optic nerve nutritional therapy with mouse nerve growth factor (30 μg) and acupoint injection in temporal with compound anisodine (2 ml).The total course was 10 days.The patients of treatment group received IOP lowering treatment to reduce the IOP to ≥ 8 mm Hg (1 mm Hg=0.133 kPa) or in a 30％ reduction.The patients of control group received no IOP lowering treatment.The peak systolic velocity (PSV),pulsatility index (PI) and resistance index (RI) of ophthalmic artery (OA),central retinal artery (CRA) and short posterior ciliary arteries (PCA) before and after treatment were comparatively analyzed by color doppler flow imaging.Results The differences of PSV (t=1.023,1.145,0.569),PI (t=0.679,0.956,1.634) and RI (t=0.816,1.657,0.998) of OA,CRA and PCA before treatment in treatment group and control group were not statistically significant (P＞ 0.05).Compared with before treatment,PSV (t=3.150,7.650,3.520) and PI (t=2.420,5.430,7.650) of OA,CRA and PCA increased obviously (P＜0.05),RI of OA,CRA and PCA decreased obviously (t=5.320,9.640,18.360; P＜0.05) after treatment in treatment group.In control group,the differences of PSV (t=2.090,-2.550,-2.100) and PI (t=-2.310,-2.230,-4.490) of OA,CRA and PCA between before and after treatment were not statistically significant (P＞0.05) ; but the differences of RI of OA,CRA and PCA between before and after treatment was statistically significant (t=2.970,2.160,2.690μ P＜0.05).Compared with control group,PSV (t=2.632,2.135,5.364) and PI (t=3.251,2.432,4.243) of OA,CRA and PCA increased obviously (P＜0.05),RI of OA,CRA and PCA decreased obviously (t=3.664,2.938,4.324; P＜0.05) after treatment in treatment group.Conclusion Lowering intraocular pressure treatment can improve the ocular hemodynamics in NAION patients. Key words: Optic neuropathy, ischemic/therapy; Ocular hypertension/drug therapy; Hemodynamics; Ultrasonography,doppler,color

Injection of nerve growth factor into a low back muscle induces long-lasting latent hypersensitivity in rat dorsal horn neurons

Little is known about the central mechanisms underlying the transition from local or regional to widespread pain in low back pain patients. The aim of the study was to find out if muscle input induced by injection of nerve growth factor (NGF) can be used as an animal model for studying spinal mechanisms involved in widespread myofascial low back pain. Electrophysiological recordings from rat dorsal horn neurons were made in vivo to study alterations in their responsiveness caused by 2 injections of NGF into the multifidus muscle at an interval of 5days. NGF is known to be closely associated with many painful muscle disorders. The results demonstrate that the 2 NGF injections—but not a single one—caused a significant hyperexcitability of spinal neurons. Five days after the first NGF injection, the neurons were not significantly sensitized but were easier to sensitize by a second injection. The state of the neurons resembles nociceptive priming. Important findings were that the proportion of neurons having multiple receptive fields (RFs) in various tissues was significantly higher after 2 NGF injections, and new RFs appeared on the distal hind limb. The new RFs were located not in the skin but in deep tissues (muscles, thoracolumbar fascia). If similar changes occur in patients, the data might explain the diffuse nature and spread of myofascial low back pain.

Local nociceptor sensitization with NGF: Mechanical or heat hyperalgesia á la carte?

In this issue, you will find a paper by C. Mills et al., (2013) entitled ‘Characterization of nerve growth factor induced mechanical and thermal hypersensitivity in the rat’. Nerve growth factor (NGF) has been found to sensitize pain processing already in the 1990s (Lewin et al.,1992; Woolf et al.,1994; Bennett et al.,1998; Koltzenburg et al.,1999; Kerr et al.,2001). Successful pain treatment using NGF antibodies (Lane et al.,2010; Katz et al.,2011; Schnitzer et al.,2011) has renewed the interest in this mechanism also inciting studies on human pain models applying NGF intramuscularly (Andersen et al.,2008; Svensson et al.,2008), in the muscle fascia (Deising et al.,2012) and in the skin of human volunteers (Rukwied et al.,2010; Weinkauf et al.,2012). Upon injection in the skin, NGF-induced mechanical and thermal hyperalgesia is found in animal models (Andreev et al.,1995). There are however, major differences in the time course of the hyperalgesia between tissues and between species. While the duration of hyperalgesia upon injection in the muscle is only a few days (Andersen et al.,2008; Svensson et al.,2008), intracutaneous injection in humans causes mechanical hyperalgesia lasting more than a month (Rukwied et al.,2010; Weinkauf et al.,2012). In rodents, heat and mechanical hyperalgesia have been reported to start hours after the injection and last hours or up to a couple of days. Mills and colleagues in this issue of European Journal of Pain report on a differential time course of heat and mechanical hyperalgesia in a rat model of NGF-induced hypersensitivity. Upon dermal injection of NGF, heat hyperalgesia was maximum after 3 h and had returned to baseline at 48 h. In contrast, mechanical hyperalgesia as measured with paw withdrawal threshold had developed 1 h after the injection but was significant for 1 week even for low NGF doses (0.3 μg) and for about 2 weeks at 3 μg. Interestingly, hypersensitivity to pin prick stimulation was significant only for the time point 1 h after the injection. The basis for the mismatch between pin prick hyperalgesia and withdrawal thresholds is unclear. However, one might suggest that the phasic pin prick tests are assessing behavioural pain threshold whereas the tonic nature of the paw pressure tests would allow more for suprathreshold testing. Along these lines, it is conceivable that the NGF-induced long-lasting mechanical hyperalgesia is based on sensitization of suprathreshold encoding as has similarly been found in the human NGF model (Rukwied et al.,2010), but also in the rat ultraviolet irradiation model (Bishop et al.,2010) in which high threshold mechanosensitive C-nociceptors were found to increase their firing upon suprathreshold mechanical stimulation, with their activation threshold being unchanged. The differential time course of mechanical and heat hypersensitivity resembles the delayed mechanical hyperalgesia observed in human volunteers and suggests different underlying mechanisms. While heat hyperalgesia might be based on sensitization and translocation of TRPV1, the prolonged mechanical sensitization might be linked to expression changes of axonal or sensory transduction proteins. The key implication of these results in respect to pharmacological testing is obviously the use of early and late hyperalgesia for separate testing of analgesic effects. The centrally acting analgesics morphine and gabapentin did not show a clear separation of their analgesic potency when comparing early and late phase or heat versus mechanical hyperalgesia. In contrast, duloxetin appeared to be more effective against the late mechanical hyperalgesia whereas the anti-inflammatory drugs Diclofenac and Celecoxib might be more effective in the early phase. In summary, the prolonged mechanical hyperalgesia as compared to the more transient heat hyperalgesia in the rat NGF model might allow for a better characterization of analgesic effects. Conflict of interest None declared.

NGF rescues hippocampal cholinergic neuronal markers, restores neurogenesis, and improves the spatial working memory in a mouse model of Huntington's Disease.

Recent studies in Huntington's disease (HD) mouse models and patients suggest that hippocampal neurons and their cholinergic afferents are involved in the cognitive deficits seen in the disease. Nerve growth factor (NGF) is an essential regulator of cholinergic neuronal survival and neurotransmission. We asked whether NGF might be involved in HD and if intra-cerebroventricular infusion of NGF can rescue hippocampal cholinergic neuronal markers, restore neurogenesis, and improve the spatial working memory in R6/1 mouse model of HD. We quantified NGF protein level by enzyme-linked immunosorbent assay (ELISA), intracerebroventricularly infused NGF, assessed cholinergic neuronal markers by Western blotting and quantitative RT-PCR, evaluated neurogenesis by immunohistochemistry, and studied spatial working memory using radial maze. By quantifying NGF protein in the hippocampus of the R6/1 mice at different ages, we found progressive decreases in NGF protein levels. We then increased NGF levels in the R6/1 mice through intra-cerebroventricular infusion. We observed elevations of the cholinergic neurochemical markers vesicular acetylcholine transporter (VAChT) and choline acetyltransferase (ChAT) in the hippocampus and in the septal region, which contain the cell bodies of basal forebrain cholinergic neurons (BFCNs), but not in the striatum that harbors cholinergic interneurons. Finally, we found that NGF infusion also restored hippocampal neurogenesis and improved spatial working memory. Our results suggest that intracerebral injections of NGF might be a valuable therapy against cognitive symptoms in HD and should be further studied in HD animal models and patients.

Exogenous nerve growth factor supplementation elevates myocardial immunoreactivity and attenuates cardiac remodeling in pressure-overload rats

It is postulated that supplementation of exogenous nerve growth factor (NGF) might mediate improvement of the cardiac sympathetic nerve function in heart failure (HF). Local intramuscular injection of NGF near the cardiac sympathetic ganglia could influence the innervation pattern, norepinephrine transporter (NET) gene expression, and improve the cardiac remodeling in experimental HF animals. In this study, we injected NGF into the scalenus medius muscles of Sprague-Dawley rats with abdominal aortic constriction (AC). The nerve innervated pattern, left ventricular morphology, and function following injection in rats with AC were investigated respectively by immunohistochemistry and echocardiography. Levels of mRNA expression of NET, growth associated protein 43 (GAP 43), NGF and its receptors TrkA and p75(NTR), and brain natriuretic peptide (BNP) were measured by real-time polymerase chain reaction. The results showed that myocardial NGF mRNA levels were comparable in rats with AC. Short-term supplementation of exogenous NGF raised the myocardial NGF immunoreactivity, but did not cause hyperinnervation and NET mRNA upregulation in the AC rats. Furthermore, myocardial TrkA mRNA was found to be remarkably decreased and p75(NTR) mRNA was increased. Myocardial TrkA downregulation may play a beneficial effect for avoiding the hyperinnervation, and it is reasonable to postulate that p75(NTR) can function as an NGF receptor in the absence of TrkA. Interestingly, local NGF administration into the neck muscles near the ganglia could attenuate cardiac remodeling and downregulate BNP mRNA. These results suggest that exogenous NGF can reach the target tissue along the axons anterogradely, and improve the cardiac remodeling.

Characterization of nerve growth factor‐induced mechanical and thermal hypersensitivity in rats

Injection of nerve growth factor (NGF) produces mechanical and thermal hypersensitivity in rodents and humans. Treatment with sequestering antibodies demonstrates the importance of NGF in various pain states, with efficacy seen in a number of animal pain models and in painful human conditions. However, these phenomena have not been evaluated in the context of using NGF-induced hypersensitivities as a model of pain. NGF-induced behaviours were characterized using von Frey filament, pinprick and thermal endpoints and then pharmacologically evaluated with known reference agents. Intraplantar NGF injection produced a dose-dependent increase in thermal sensitivity that lasted through 24 h post-injection and an immediate long-lasting (2 week) increase in mechanical sensitivity at the injection site, with no effects detected at secondary sites. NGF-induced mechanical sensitivity was pharmacologically characterized at 4 h and 1 week post-NGF injection. The nonsteroidal anti-inflammatory drugs (NSAIDs), celecoxib and diclofenac, were minimally effective against both thermal and mechanical endpoints. Gabapenitn and duloxetine were only moderately effective against thermal and mechanical hypersensitivity. Morphine was effective against thermal and mechanical endpoints at every time point examined. Treatment with the transient receptor potential vanilloid 1 (TRPV1) antagonist A-784168 partially attenuated NGF-induced thermal and mechanical sensitivity at all time points examined. The results reported here suggest that effects of NGF on thermal and mechanical sensitivity in rats are similar to those reported in human and are partially driven by TRPV1. The rat NGF model may serve as a potential translational model for exploring the effects of novel analgesic agents.

Transient receptor potential vanilloid 1 mediates nerve growth factor‐induced bladder hyperactivity and noxious input

To explore the role of transient receptor potential vanilloid 1 (TRPV1) in the excitatory effects of chronic administration of nerve growth factor (NGF) on bladder-generated sensory input and reflex activity. To explore new therapeutic targets for bladder dysfunction. Wild-type (WT) and TRPV1 knockout (KO) mice received daily intraperitoneal injections of NGF (1 µg/10 g) or saline for a period of 4 days, during which time thermal sensitivity was evaluated daily. On the 5th day, mice were anaesthetized and cystometries were performed. The frequency, amplitude and area under the curve (AUC) of bladder reflex contractions were determined. c-Fos expression was evaluated on L6 spinal cord sections of WT and TRPV1 KO mice treated with saline or chronic NGF by immunohistochemistry. TrkA receptor staining intensity was determined in L6 spinal cord sections and respective dorsal root ganglia of WT and TRPV1 KO mice. Repeated administration of NGF induced thermal hypersensitivity in WT but not in TRPV1 KO mice. The frequency of bladder contractions of saline-treated WT and TRPV1 KO mice was similar, the values respectively being 0.45 ± 0.12/min and 0.46 ± 0.16/min. Treatment with NGF enhanced bladder reflex activity in WT mice to 1.23 ± 0.41/min (P < 0.05). In NGF-treated KO mice, the frequency of bladder contractions was 0.60 ± 0.05/min. Irrespective of treatment, no differences were observed in the amplitude of bladder contractions of WT and TRPV1 KO mice. The AUC was significantly increased in NGF-treated WT-mice, when compared with saline-treated WT-mice. No changes were found in AUC of saline-treated and NGF-treated TRPV1 KO mice. Chronic administration of NGF resulted in a significant increase of spinal c-Fos expression in WT mice (P < 0.05 vs KO animals), but not in TRPV1 KO animals. TrkA expression was similar in WT and TRPV1 KO mice. NGF-induced bladder overactivity and noxious input depend on the interaction of NGF with TRPV1. The lack of bladder overactivity in TRPV1 KO mice treated with NGF does not represent loss of TrkA expression. TRPV1 is essential for NGF-driven bladder dysfunction and represents a bottleneck target in bladder pathologies associated with NGF up-regulation.

Nerve growth factor regulates axial rotation during early stages of chick embryo development

Nerve growth factor (NGF) was discovered because of its neurotrophic actions on sympathetic and sensory neurons in the developing chicken embryo. NGF was subsequently found to influence and regulate the function of many neuronal and non neuronal cells in adult organisms. Little is known, however, about the possible actions of NGF during early embryonic stages. However, mRNAs encoding for NGF and its receptors TrkA and p75(NTR) are expressed at very early stages of avian embryo development, before the nervous system is formed. The question, therefore, arises as to what might be the functions of NGF in early chicken embryo development, before its well-established actions on the developing sympathetic and sensory neurons. To investigate possible roles of NGF in the earliest stages of development, stage HH 11-12 chicken embryos were injected with an anti-NGF antibody (mAb αD11) that binds mature NGF with high affinity. Treatment with anti-NGF, but not with a control antibody, led to a dose-dependent inversion of the direction of axial rotation. This effect of altered rotation after anti NGF injection was associated with an increased cell death in somites. Concurrently, a microarray mRNA expression analysis revealed that NGF neutralization affects the expression of genes linked to the regulation of development or cell proliferation. These results reveal a role for NGF in early chicken embryo development and, in particular, in the regulation of somite survival and axial rotation, a crucial developmental process linked to left-right asymmetry specification.

Local injection of nerve growth factor via a hydrogel enhances bone formation during mandibular distraction osteogenesis

The objective of this study was to evaluate the effects of the injectable NGF-carrying collagen/nano-hydroxyapatite/alginate hydrogel on the bone formation in a rabbit mandibular distraction osteogenesis model. Thirty-five New Zealand white rabbits underwent bilateral madibular distraction osteogenesis at a rate of 0.75 mm/12 h for 6 days. The rabbits were divided into 4 groups: group 1 received injections of collagen/nano-hydroxyapatite/alginate hydrogel containing hNGFβ; groups 2, 3, and 4 received injections of hNGFβ, Col/nHA/Alg hydrogel, and saline, respectively. The injections were performed on both sides of the mandible at the end of the lengthening phase. All the animals were killed at a consolidation time of 14 days. No difference in regenerate bone dimensions was observed among the 4 groups. Bone mineral density, the maximum load, and the bone volume/total volume of the new bone in the distraction gap in group 1 was significantly greater (P < .05) than in the other 3 groups. Application of the Col/nHA/Alg hydrogel as an NGF delivery during the consolidation phase of distraction osteogenesis increased regeneration and new bone formation.

Tissue repair driven by two different mechanisms of growth factor plasmids VEGF and NGF in mice auricular cartilage: regeneration mediated by administering growth factor plasmids

The focus of this study was to compare the role of nerve growth factor (NGF) and vascular endothelial growth factor (VEGF) in the regeneration of experimental skin and cartilage trauma. The role of VEGF in this process is known since decade; the NGF participation on this process has been first discussed within the spinal cord injury repair. We hypothesized that both VEGF and NGF induce angiogenesis and take part on the repair process. The angiogenesis response and the cartilage regeneration after phVEGF(165) plasmid and rat pcNGF plasmid administration were investigated using BALB/c mice. PhVEGF(165) and pcNFG were injected into the right mice ear and plain vector injection into the left ear the day before trauma. The next day, all mice were ear-punched, resulting in 2-mm diameter puncture through the center of both pinnae. In BALB/c mouse strain, a significantly faster cartilage repair was observed after phVEGF(165) and pcNGF injection into punched ear area in comparison to the control group. It has been shown that the healing process is after VEGF and NGF injection driven differentially. In case of VEGF is the cartilage wound repaired by induction of new chondrocytes differentiation. In the case of NGF, the regeneration is supported by immature leukocytes attracted into the punched area. The leukocytes induct angiogenesis so far indirectly by inflammation. The NGF-induced inflammation environment may be a part of mosaic creating the complete picture of regeneration.

Nerve growth factor selectively decreases activity-dependent conduction slowing in mechano-insensitive C-nociceptors

Nerve growth factor (NGF) induces acute sensitization of nociceptive sensory endings and long-lasting hyperalgesia. NGF modulation of sodium channel expression might contribute to neurotrophin-induced hyperalgesia. Here, we investigated NGF-evoked changes of the activity-dependent slowing of conduction in porcine C-fibers. Animals received intradermal injections of NGF (2 μg or 8 μg) or saline in both hind limbs. Extracellular recordings from the saphenous nerves were performed 1 week later. Based on sensory thresholds and electrically induced activity-dependent slowing (ADS) of axonal conduction, C-fibers were classified as mechano-sensitive afferents, mechano-insensitive afferents, cold nociceptors, and sympathetic efferents. NGF (2 μg) increased conduction velocity in C-fibers from 1.0 ± 0.05 m/s to 1.2 ± 0.07 m/s. In mechano-insensitive afferents, NGF (8 μg) reduced activity-dependent slowing of conduction, from 5.3 ± 0.2% to 3.2 ± 0.5% (0.125–0.5 Hz stimulation) and from 28.5 ± 1.3% to 20.9 ± 1.9% (2 Hz stimulation), such that ADS no longer differentiated between mechano-sensitive and mechano-insensitive fibers. Accordingly, the number of fibers with pronounced ADS decreased but more units with pronounced ADS were mechano-sensitive. Spontaneously active C-fibers were increased above the control level (1%) by NGF 8 μg (8%). The results demonstrate that NGF changes the functional axonal characteristics of mechano-insensitive C-fibers and enhances spontaneous activity thereby possibly contributing to hyperalgesia.

NGF enhances electrically induced pain, but not axon reflex sweating

High-affinity receptors for nerve growth factor (NGF) are found on nociceptors and sympathetic efferents. NGF is known to sensitize nociceptors, increase innervation density, and fire frequency of sympathetic fibers. We explored axonal sensitization of afferent and efferent fibers following intracutaneous injection of NGF in human and pig skin. In humans, frequency-dependent (5, 20, 100Hz) electrically induced pain was assessed 1, 3, 7, 21, and 49days post injection. Sweat output was recorded in parallel using the quantitative sudomotor axon reflex test (QSART). Electrically induced pain ratings (7.5mA for 30s) significantly increased at the NGF sites for 5Hz (numeric rating scale [NRS] 6±0.5 vs 3.7±0.4), 20Hz (NRS 7.2±0.4 vs 5±0.5), and 100Hz stimulation (NRS 6.9±0.4 vs 5.4±0.3) at day 21, and also for 5Hz at day 49 (NRS 5.4±0.4 vs 3.8±0.3). Electrically evoked QSART increased frequency dependent, but was not altered by NGF throughout the entire observation period (average QSART at 5Hz: 3mL/h/m2, 20Hz: 9mL/h/m2, 100Hz: 10mL/h/m2). Similarly, NGF did not change the activity-dependent slowing of conduction of sympathetic efferents (6±2% vs 5.1±1.5%, for 3minutes, 2Hz) in pig single-fiber recordings. In parallel to the increased pain ratings recorded in humans, activity-dependent slowing of mechano-insensitive nociceptors was reduced by NGF (18.1±2% vs 29±1.4%). In summary, axonal sensitization of nociceptors by NGF could underlie the hyperalgesia to electrical stimulation. Enhanced responses were limited to nociceptors, as no sensitization was found in sympathetic efferent neurons.NGF administration increased electrically induced pain, decreased activity-dependent slowing of mechano-insensitive nociceptors, but did not affect sympathetic efferents. Axonal sensitization could underlie the NGF-evoked hyperalgesia.

Synergistic effects of ultrashortwave electrical stimulation and nerve growth factor in treating sciatic nerve injury

Objective To investigate the effects of ultrashortwave (USW) diathermy and electrical stimulation (ES) used in combination with nerve growth factor (NGF) in the treatment of experimental sciatic nerve injury.Methods Sixty adult Wistar rats were randomly divided into a normal control group, a model group, an NGF group,a physiotherapy group and a combined treatment group. A model of sciatic nerve injury was established in the latter four groups. Beginning on the 2nd day after the operation, no treatment was given in the model group, NGF was injected in the NGF group, diathermy and ES were administrated to rats in the physiotherapy group, and the combined treatment group was treated with USW diathermy, EW and NGF. Function, electrophysiology and morphology were evaluated at the 2nd, 7th, 14th and 30th days after the operation Results The average sciatic nerve function index (SFI), nerve conduction velocity (NCV) and nerve regeneration in the NGF, physiotherapy and combined treatment groups were significantly better than in the model group, with those in the combined treatment group improved to the greatest extent. At the 30th day there was no significant difference between the combined treatment group and the normal control group in terms of SFI, NCV, axon regeneration or myelin sheath thickness. The number of myelinated nerve fibers and the average axon diameter in the combined treatment group and normal control group were significantly higher than those in the model, NGF or physiotherapy group. Conclusions With NGF injection, additional application of USW diathermy and ES may significantly enhance the regeneration of the sciatic nerve and aid functional recovery after injury. Key words: Sciatic nerve injury; Ultrashortwave diathermy; Electrical stimulation; Nerve growth factor

Local Pain and Spreading Hyperalgesia Induced by Intramuscular Injection of Nerve Growth Factor Are Not Reduced by Local Anesthesia of the Muscle

Injections with local anesthesia for therapeutic and diagnostic purposes are common clinical practice. This double-blind placebo controlled study explores the rational of local anesthetic blocks for the detection of muscle pain as the primary generator in spreading hyperalgesic conditions. Experimental muscle pain was induced by injections of nerve growth factor (NGF) in the left and right supraspinatus muscle of 11 healthy volunteers. One day later ropivacaine and saline were administered with ultrasound guidance in the left or right supraspinatus muscle in a randomized double-blind manner. Assessments before NGF, day 1 (before and after ropivacaine/saline), and day 7 included: visual analog scale (VAS) pain scores at rest and during shoulder shrugging, time until exercise interruption owing to pain, cutaneous pain sensitivity, pressure pain thresholds, and pain VAS during tonic pressure stimulation. Cutaneous and pressure pain sensitivity were assessed over the supraspinatus, infraspinatus, and deltoideus muscles, and at the web space between first and second finger. Increased VAS pain scores and increased pressure pain sensitivity of the supraspinatus and infraspinatus muscles were found in both sides one day after the NGF injection compared with baseline. One day after NGF injections, the time until participants stopped exercising was reduced compared with baseline. The increased muscle pain sensitivity was not normalized by intramuscular ropivacaine. Saline caused increased VAS pain scores compared with ropivacaine. Muscle pain and spreading hyperalgesia induced by NGF is maintained despite anesthesia of the primary nociceptive locus. This indicates that intramuscular injection of local anesthetics may not be a valid diagnostic method for primary muscle pain.

Effect of nerve growth factor on the early phase of osseointegration around oral implants

To observe the early bone integration of oral implants after injection of exogenous nerve growth factor (NGF) and investigate the effects of NGF on peri-implant osseointegration. Twelve New Zealand white rabbits were used in this study to establish bi-mandible implant model. Then local injection of 1 µg NGF was given on the right side of the mandible as experimental group and normal saline only was injected on the left side as control group once a day for seven days. The rabbits were respectively sacrificed at 2, 4 and 8 weeks after surgery. The implant-bone grinding samples were prepared and stained by toluidine blue for general observation, X-ray, histology and bone histomorphometry analysis. The density of the new bone around implants at 2 and 4 weeks was lower than normal bone. Compared with the control group, the quantity of new bone and bone-implant contact ratio significantly increased in the experimental group. At 8 weeks, the new bone density in both groups was similar to the normal bone. In the experimental group, the haversian system was observed. Bone contact ratio was significantly different between experimental and control group at 2 and 4 weeks, but similar at 8 weeks.[control group at 2 weeks (26.67 ± 3.88)%, 4 weeks (52.59 ± 5.07)% and 8 weeks (97.33 ± 6.75)%, experimental group at 2 weeks (42.24 ± 6.67)%, 4 weeks (72.25 ± 6.30)% and 8 weeks (99.15 ± 4.68)%]. Applying exogenous NGF in the early phase could accelerate the formation and maturation of trabecular bone around the implants and shorten the period of osseointegration. Nerve growth factor could promote osseointegration in the early stage of oral implantation.

Peripheral nerve regeneration using composite poly(lactic acid‐caprolactone)/nerve growth factor conduits prepared by coaxial electrospinning

Many neurotrophic factors have been shown to promote neurite outgrowth by improving the microenvironment that is required for nerve regeneration. However, the delivery of these bioactive agents to the nerve injury site, as well as effective and local release, remains a challenging problem. We have developed a novel composite nerve conduit comprised of poly(lactic acid-caprolactone) (P(LLA-CL)) and nerve growth factor (NGF). This was developed from core-shell structured biodegradable nanofibers, which were fabricated by coaxial electrospinning of P(LLA-CL) for the shell and bovine serum albumin (BSA) or BSA/NGF for the core. In rats, gaps of 10-mm long sciatic nerves were bridged using an autograft, an empty P(LLA-CL) conduit, a NGF injection P(LLA-CL) conduit, a P(LLA-CL)/NGF composite conduit, respectively. Regenerated nerve fibers were harvested and morphological and functional evaluation of nerve regeneration was performed at 12 weeks postsurgery. Although partial biodegradation and small cracks in the conduits were observed, the conduit outlines remained intact for 12 weeks after surgery. Based on functional and histological observations, the number and arrangement of regenerated nerve fibers, myelination, and nerve function reconstruction was similar in the P(LLA-CL)/NGF conduit group to that of the nerve autograft group (p > 0.05), but was significantly greater to the empty P(LLA-CL) and injection NGF P(LLA-CL) conduit groups (both p < 0.05). Therefore, the composite P(LLA-CL)/NGF conduit, which exhibited favorable mechanical properties and biocompatibility, could effectively promote sciatic nerve regeneration in rats.