Augmented Buyang Huanwu Decoction facilitates axonal regeneration after peripheral nerve transection through the regulation of inflammatory cytokine production.

ETHNOPHARMACOLOGICAL RELEVANCE: Herbal formulation Buyang Huanwu Decoction (BYHWD) has been used to treat cardiovascular disorders including cerebral ischemia. Recent studies showed its effects on promoting axonal regeneration after nerve injury. However, compositional reformulation supplemented with herbal components that regulates inflammation may increase its efficacy for nerve repair. AIM OF THE STUDY: We prepared a new herbal decoction by adding selected herbal components to BYHWD (augmented BYHWD; ABHD) and investigated the effect of ABHD on the production of inflammatory cytokines and axonal regeneration using an animal model of nerve transection and coaptation (NTC). MATERIALS AND METHODS: A rat model of NTC was performed on the sciatic nerve. The sciatic nerve and dorsal root ganglion (DRG) were isolated and used for immunofluorescence staining and western blot analysis. DRG tissue was also used to prepare primary neuron culture and the length of neurites was analyzed. Sensorimotor nerve activities were assessed by rotarod and von Frey tests. RESULTS: Three herbal components that facilitated neurite outgrowth were chosen to formulate ABHD. ABHD administration into the sciatic nerve 1 week or 3 months after NTC facilitated axonal regeneration. Cell division cycle 2 (Cdc2) and brain-derived neurotrophic factor (BDNF) proteins were induced from the reconnected distal portion of the sciatic nerve and the levels were further elevated by in vivo administration of ABHD. Phospho-Erk1/2 level was increased by ABHD treatment as well, implying its role in mediating retrograde transport of BDNF signals into the neuronal cell body. Production of inflammatory cytokines IL-1ß and TNF-α was induced in the reconnected nerve but attenuated by ABHD treatment. Behavioral tests revealed that ABHD treatment improved functional recovery of sensorimotor activities. CONCLUSIONS: A newly formulated ABHD is effective at regulating the production of inflammatory cytokines and promoting axonal regeneration after nerve transection and may be considered to develop therapeutic strategies for peripheral nerve injury disorders.

Anti-Inflammatory Effects of Modified Buyang Huanwu Decoction.

METHODS: A cytotoxicity assay for BHD was performed using the MTT assay. Following treatment with BHD, mBHD-1, and mBHD-2 in the presence of lipopolysaccharide (LPS), nitric oxide (NO) secretion was detected in cell supernatants using a NO detection kit. The expression of proinflammatory mediators was detected using RT-PCR and western blotting. To verify the mechanism of mBHD, specific inhibitors of JNK (SP600125) or p38 (SB203580) were used for co-treatment with mBHD, and then the changes in NO and nitric oxide synthase (iNOS) were measured. RESULTS: Both mBHD-1 and mBHD-2 showed greater anti-inflammatory effects than BHD. Both mBHD-1 and mBHD-2 inhibited NO secretion and decreased the expression of IL-1ß, IL-6, TNF-α, and iNOS. Treatment with a p38 inhibitor and a JNK inhibitor in mBHD-1- and mBHD-2-treated cells resulted in inhibition of NO and iNOS. CONCLUSION: We provided the first experimental evidence that mBHD may be a more useful anti-inflammatory than BHD. High concentrations or long-term use of BHD may be harmful to inflammatory status. Therefore, the length of treatment and concentration should be considered depending on the targeted disease.

Acupuncture stimulation attenuates TNF-α production via vagal modulation in the concanavalin A model of hepatitis.

BACKGROUND: A growing body of evidence shows that neuronal activity is involved in modulating the efficacy of acupuncture therapy. However, it has been seldom investigated whether neuronal activity following acupuncture stimulation is effective at regulating hepatic inflammation. OBJECTIVE: Using the concanavalin A (ConA) model of hepatitis, we investigated the regulation of inflammatory cytokine tumor necrosis factor (TNF)-α in the liver tissue and the blood after acupuncture stimulation at ST36. METHODS: Mice were subjected to ConA injection, acupuncture stimulation at ST36 by manual acupuncture (MA) or electroacupuncture (EA) procedures, and vagotomy (VNX). Liver tissue and blood were collected for TNF-α analysis. TNF-α mRNA was analyzed by real-time polymerase chain reaction (PCR), and TNF-α, CD11b, CD68, and Erk1/2 proteins were analyzed by Western blotting, immunofluorescence staining, and enzyme-linked immunosorbent assay. RESULTS: TNF-α mRNA and protein were induced in CD11b-positive hepatic cells and the plasma at 6-24 h after ConA injection. The application of MA or EA was very effective at attenuating the production of TNF-α. Anti-inflammatory effects of acupuncture were greatly suppressed by VNX in ConA-injected animals, suggesting the requirement of vagus nerve activity in acupuncture-mediated anti-inflammatory responses. Electrical stimulation of the sciatic nerve (SNS) resulted in an anti-inflammatory effect similar to acupuncture stimulation. In parallel with TNF-α, production of phospho-Erk1/2, which was induced in the liver tissue, was downregulated by MA and EA in liver cells. CONCLUSION: The regulatory effects of acupuncture stimulation on inflammatory responses in the liver may be modulated through the activation of the vagus nerve pathway.

Effects of Acupuncture on Chronic Stress-Induced Depression-Like Behavior and Its Central Neural Mechanism.

Depression is a serious psychiatric disorder with an enormous socioeconomic burden, and it is commonly comorbid with pain, chronic fatigue, or other inflammatory diseases. Recent studies have shown that acupuncture is an effective therapeutic method for reducing depressive symptoms; however, the underlying mechanism remains unknown. In this study, we investigated the effects of acupuncture on chronic stress-induced depression-like behavior and its central neural mechanisms in the brain. We induced chronic restraint stress (CRS) in male C57BL/6 mice for 14 or 28 consecutive days. Acupuncture treatment was performed at KI10·LR8·LU8·LR4 or control points for 7 or 14 days. Depression-like behavior was assessed with the open field test. Then, brain neural activity involving c-Fos and serotonin-related mechanisms via the 5-HT1A and 5-HT1B receptors were investigated. Acupuncture treatment at KI10·LR8·LU8·LR4 points rescued the depressive-like behavior, while control points (LU8·LR4·HT8·LR2) and non-acupoints on the hips did not. Brain neural activity was changed in the hippocampus, cingulate cortex, motor cortex, insular cortex, thalamus, and the hypothalamus after acupuncture treatment. Acupuncture treatment increased expression of 5-HT1A receptor in the cortex, hippocampus, thalamus, and the hypothalamus, and of 5-HT1B in the cortex and thalamus. In conclusion, acupuncture treatment at KI10·LR8·LU8·LR4 was effective in alleviating the depressive-like behavior in mice, and this therapeutic effect was produced through central brain neural activity and serotonin receptor modulation.

Electroacupuncture therapy in inflammation regulation: current perspectives.

Although acupuncture therapy is increasingly used to treat diverse symptoms and disorders in humans, its underlying mechanism is not known well. Only recently have experimental studies begun to provide insights into how acupuncture stimulation generates and relates to pathophysiological responsiveness. Acupuncture intervention is frequently used to control pathologic symptoms in several visceral organs, and a growing number of studies using experimental animal models suggest that acupuncture stimulation may be involved in inducing anti-inflammatory responses. The vagus nerve, a principal parasympathetic nerve connecting neurons in the central nervous system to cardiovascular systems and a majority of visceral organs, is known to modulate neuroimmune communication and anti-inflammatory responses in target organs. Here, we review a broad range of experimental studies demonstrating anti-inflammatory effects of electroacupuncture in pathologic animal models of cardiovascular and visceral organs and also ischemic brains. Then, we provide recent progress on the role of autonomic nerve activity in anti-inflammation mediated by electroacupuncture. We also discuss a perspective on the role of sensory signals generated by acupuncture stimulation, which may induce a neural code unique to acupuncture in the central nervous system.

Bogijetong Decoction and Its Selected Formulation Are Involved in Alleviating Neuropathic Pain in a Rat Model of Chronic Constrictive Injury.

Bogijetong decoction (BGJTD) is a formulation that is used for the treatment of neuropathic pain caused by cancer therapy, diabetes, and peripheral nerve injury. In the previous study, we selected four herbal constituents from BGJTD, formulated new decoction (BeD), and demonstrated its efficacy on the neuroprotection of peripheral sciatic nerve in streptozotocin-induced diabetic animals. Here, we report attenuating effects of BGJTD and BeD on neuropathic pain. Neuropathic pain was induced by ligation of the sciatic nerve to generate chronic constrictive injury (CCI). BeD was more effective than BGJTD in alleviating neuropathic pain lasting 3 - 4 weeks after CCI. In vivo administration of BeD did not alter the levels of brain-derived neurotrophic factor (BDNF) which were strongly induced by CCI in the sciatic nerve but downregulated TrkB production in the sciatic nerve. Downregulation of TrkB signals by BeD was confirmed in cultured DRG neurons. BGJTD was more effective in attenuating TNF-α production in the sciatic nerve than BeD, whereas BeD increased IL-6 more efficiently than BGJTD. Furthermore, phopsho-Erk1/2 was increased in the sciatic nerve and dorsal root ganglia (DRG) after BeD treatment. Neurite outgrowth of primary DRG neurons prepared from rats which had undergone CCI for 7 days was significantly increased in BeD-treated group of animals compared to the control and BGJTD-treated groups. Compositional comparison of BeD revealed that the neurite outgrowth was facilitated by the treatments of Panax ginseng and Paeonia lactiflora. Together, these data suggest that BeD induces unique molecular response at the injury site and may trigger retrograde signaling into the neuronal cell body to modulate pain responses.

Facilitating effects of Buyang Huanwu decoction on axonal regeneration after peripheral nerve transection.

ETHNOPHARMACOLOGICAL RELEVANCE: In traditional Asian medicine, Buyang Huanwu decoction (BYHWD) has been used for the treatment of cardiovascular and neurological disorders. Recent experimental studies have begun to provide evidence on the protective effects of BYHWD on injured peripheral nerves. AIM OF THE STUDY: To examine whether BYHWD was effective in inducing axonal regeneration after peripheral nerve transection, and if so, how it acted on the nerve. MATERIALS AND METHODS: The sciatic nerve in rats was transected and resutured 0, 1, or 4 weeks later. BYHWD was orally administered daily into the animals with nerve transection and coaptation (NTC). Axonal regeneration was measured by immunofluorescence staining of NF-200 and superior cervical ganglion 10 (SCG10) and by retrograde tracing method. Changes of protein levels in the sciatic nerve were analyzed by western blot analysis. Effects of BYHWD and its constituents on neurite outgrowth were analyzed in cultured dorsal root ganglion (DRG) neurons. Hot plate and treadmill training tests were performed to assess the levels of functional recovery after nerve injury. RESULTS: The rate of axonal regeneration was attenuated by delayed coaptation after transection, but improved by BYHWD treatment. Levels of phospho-Erk1/2 and Cdc2 phosphorylation of vimentin, measured as indicators of the activation of regenerating axons and supportive Schwann cells, were increased in the sciatic nerve of NTC animals, and their distribution in the proximal and distal nerves were affected by BYHWD treatment. Treatment of BYHWD during the period of chronic denervation significantly increased axonal regeneration when analyzed by immunofluorescence staining and retrograde tracing methods. Neurite outgrowth of DRG neurons cocultured with Schwann cells from the chronically transected sciatic nerves was enhanced by BYHWD treatment. Radix Paeoniae Rubra induced neurite outgrowth most efficiently among all herbal constituents of BYHWD. Finally, hot plate and treadmill training tests demonstrated that BYHWD administration significantly improved the sensorimotor nerve function in NTC animals. CONCLUSIONS: Our data suggest that BYHWD treatment may contribute to the timely interaction between regenerating axons and distal Schwann cells in the transected nerve and facilitate axonal regeneration.

Protective Effects of Bogijetong Decoction and Its Selected Formula on Neuropathic Insults in Streptozotocin-Induced Diabetic Animals.

Bogijetong decoction (BGJTD) is a mixture of herbal formulation which is used in the traditional Korean medicine for the treatment of neuropathic pain caused by diabetes. Here, we investigated the regulatory effects of BGJTD and its reconstituted decoction subgroups on the neuropathic responses in streptozotocin- (STZ-) induced diabetic animals. Be decoction (BeD) was formulated by selecting individual herbal components that induced neurite outgrowth most efficiently in each subgroup. BeD induced the neurite outgrowth in DRG neurons most efficiently among decoction subgroups and downregulated the production of TNF-α from the sciatic nerves in STZ-diabetic animals. While the levels of phospho-Erk1/2 were elevated in the sciatic nerves of STZ-diabetic animals by BGJTD and BeD treatments, p38 level was downregulated by BGJTD and BeD. A single herbal component of BeD induced neurite outgrowth comparable to BeD and was involved in the regulation of Erk1/2 activation and TNF-α production in DRG neurons. Oral administration of BGJTD and BeD in STZ-diabetic animals reduced the latency time responding to thermal stimulation. Our results suggest that the reconstituted formulation is as effective as conventional BGJTD in inducing biochemical and behavioral recoveries from the neuropathy in peripheral nerves and thus the experimental reductionism may be applied to develop the methodology for compositional analysis of herbal decoctions.

Modulation of hippocampal neuronal activity by So-ochim-tang-gamibang in mice subjected to chronic restraint stress.

BACKGROUND: So-ochim-tang-gamibang (SOCG) is a decoction formula which has been used to improve mental activity in traditional Korean medicine. The present study was performed to evaluate whether the treatment of SOCG was involved in activating hippocampal neurons in mice which were subjected to chronic restraint stress (CRS). METHODS: Mice were subjected to CRS for 2 weeks to induce depressive-like behaviors. SOCG was orally administered for the same period. mRNA expression in the hippocampus was analyzed by RT-PCR. Levels of serotonin receptor 5-HT1AR in the hippocampus were determined by western blotting and by immunofluorescence staining in coronal brain sections. Cultured neurons were prepared from the dorsal root ganglia (DRG) in mice to examine the effects of CRS and SOCG treatment on neurite outgrowth. Depressive-like behaviors of experimental animals were measured by open field test (OFT) and forced swimming test (FST). RESULTS: mRNA levels of serotonin 1A and 1B receptors (5-HT1AR and 5-HT1BR) were decreased in the hippocampus of CRS animals and increased by SOCG treatment. Signals of 5-HT1AR protein in CA3 pyramidal cells were decreased by CRS but elevated back to levels in control animals after SOCG treatment. Phospho-Erk1/2 protein in CA3 cells showed similar pattern of changes as in 5-HT1AR, suggesting coordinated regulation after SOCG treatment in CRS animals. Axonal growth-associated protein GAP-43 levels were also decreased by CRS and then increased by SOCG treatment. In vivo administration of SOCG improved neurite outgrowth of primary DRG neurons from CRS animals and also increased 5-HT1AR protein signals. Behavioral tests of open field and forced swimming showed that immobility time periods were significantly decreased by SOCG treatment. CONCLUSIONS: Our data suggest that SOCG treatment may increase synaptic responsiveness to serotonergic neuronal inputs by upregulating 5-HT1AR in the hippocampal neurons.

Induction of α6 and ß1 integrins by acupuncture stimulation in rats.

Acupuncture therapy is performed by applying the needle insertion at discrete cutaneous locations and used for the treatments of diverse symptoms and disorders. In order to elucidate mechanistic basis on how acupuncture stimulation (AS) produces therapeutic effects, it is primarily important to understand tissue responses locally at the acupuncture site (acupoint). Here, we investigated integrin protein as molecular target responding to and integrating AS. Signals of α6 and ß1 integrins were clearly induced at zusanli acupoint 24 h after AS in areas of nuclear clusters around the needle track. Induction levels of integrin were largely reduced by needle insertion at non-acupuncture point or without needle rotation. Phospho-Erk1/2 was initially decreased below the basal level after AS but increased 24 h later. Induction pattern of phospho-Erk1/2 was as similar as that of α6 integrin in its selectivity to needling procedure and tissue distribution. We further found that mRNA expression of P2X3 purinergic receptor was upregulated in the dorsal root ganglion (DRG) after AS, but decreased by the inhibition of Erk1/2 activity at the acupuncture area. Moreover, AS-mediated integrin activation was required for Erk1/2 activation at the acupuncture site and regulation of pain sensitivity in the hind paw. The present results provide a new evidence on acupuncture-specific tissue response in terms of integrin induction, and further suggest that integrin activation may be involved in transmitting mechanosensory signals from the acupoint to afferent nerve fiber.

Bogijetong decoction and its active herbal components protect the peripheral nerve from damage caused by taxol or nerve crush.

BACKGROUND: Bogijetong decoction (BGJTD) is a herbal drug formulation used in the traditional Asian medicine to treat neuropathic insults associated with diabetes and anticancer therapy. To understand the biological basis of BGJTD on protective effects against neuropathy, we investigated physiological and biochemical responses of the sciatic nerves deranged by taxol injection or crush injury in the rats. METHODS: Dissociated Schwann cells and neurons were prepared from the sciatic nerve and dorsal root ganglia (DRG) respectively and were treated with taxol and BGJTD. The sciatic nerve in the rat was injected with taxol or given crush injury. Animals were then administered orally with BGJTD. Effects of BGJTD treatment on cultured cells and in vivo sciatic nerves and DRG tissues were examined by immunofluorescence staining and western blot analysis. Sciatic nerve regeneration was assessed by histological observation using retrograde tracing technique and by behavioral hot plate test. Eighteen different herbal components of BGJTD were divided into 4 subgroups and were used to select herbal drugs that enhanced neurite outgrowth in cultured neurons. RESULTS: Morphological abnormalities in the sciatic nerve axons and DRG tissue caused by taxol injection were largely improved by BGJTD treatment. BGJTD treatment enhanced neurite outgrowth in cultured DRG neurons and improved Schwann cell survival. Phospho-Erk1/2 levels were elevated by BGJTD administration in the injured- or taxol-injected sciatic nerves. Vimentin phosphorylation catalyzed by cell division cycle 2 (Cdc2) kinase was induced from Schwann cells in the sciatic nerves after taxol injection and crush injury, and phospho-vimentin levels were further upregulated by BGJTD treatment. Retrograde tracing of DiI-labeled DRG sensory neurons revealed growth-promoting activity of BGJTD on axonal regeneration. A drug group (Be) composed of 4 active herbal components which were selected by neurite growth-enhancing activity was as effective as BGJDT for the recovery of thermal sensitivity of the hind paws which had been suppressed by taxol administration. CONCLUSIONS: These data suggest that BGJTD and its active herbal components may protects the peripheral nerve from damage caused by taxol injection and nerve crush.

Enhanced axonal regeneration of the injured sciatic nerve by administration of Buyang Huanwu decoction.

ETHNOPHARMACOLOGICAL RELEVANCE: Buyang Huanwu decoction (BYHWD) has been used in the traditional Chinese medicine for the treatment of cardiovascular and neurological symptoms, and recent experimental studies have begun to provide evidence showing its protective effects on neural cells. Yet, its function for the regenerative responses of axons in the peripheral nerve after injury is not known. AIM OF THE STUDY: The primary objective of the present study was to explore that BYHWD is involved in growth-promoting activity of the peripheral nerve axons after injury. We further examined whether the effect of BYHWD exerted directly on regrowing axons or Schwann cells. MATERIALS AND METHODS: Sciatic nerves in rats were given crush injury, and BYHWD was injected by oral administration. Sciatic nerves or DRG tissues were prepared for immunofluorescence staining and western blot analysis. Levels of axonal regeneration were quantified by retrograde tracing technique. Cultured DRG sensory neurons and Schwann cells were prepared from rats and used to examine the effects of BYHWD on the neurite outgrowth. Behavioral analysis on functional recovery after nerve injury was assessed in mice by pin prick test, adhesive removal test, and toe-spreading reflex. RESULTS: Immunofluorescence and retrograde tracing analyses showed that the distal extension of the sciatic nerve axons was significantly improved by BYHWD treatment. Levels of axonal growth-associated protein GAP-43 were upregulated by BYHWD treatment in the sciatic nerve after injury and in the neurites of cultured DRG neurons. In vivo administration of BYHWD in rats upregulated the induction level of cell division cycle 2 (Cdc2) and its phosphorylation of vimentin in Schwann cells from injured sciatic nerve. Coculture of DRG neurons with Schwann cells prepared from preinjured sciatic nerves in animals administered with BYHWD led to the enhancement in neurite outgrowth. Behavioral tests in mice given sciatic nerve injury showed a significant improvement in sensorimotor activity by BYHWD administration. CONCLUSIONS: Our results suggest that BYHWD administration into animals given sciatic nerve injury facilitates axonal regeneration by acting on both the axons undergoing regeneration and neighboring Schwann cells and improves functional recovery.

Anti-Inflammatory Effects of Acupuncture Stimulation via the Vagus Nerve.

Although acupuncture therapy is widely used in traditional Asian medicine for the treatment of diverse internal organ disorders, its underlying biological mechanisms are largely unknown. Here, we investigated the functional involvement of acupuncture stimulation (AS) in the regulation of inflammatory responses. TNF-α production in mouse serum, which was induced by lipopolysaccharide (LPS) administration, was decreased by manual acupuncture (MAC) at the zusanli acupoint (stomach36, ST36). In the spleen, TNF-α mRNA and protein levels were also downregulated by MAC and were recovered by using a splenic neurectomy and a vagotomy. c-Fos, which was induced in the nucleus tractus solitarius (NTS) and dorsal motor nucleus of the vagus nerve (DMV) by LPS and electroacupuncture (EAC), was further increased by focal administration of the AMPA receptor blocker CNQX and the purinergic receptor antagonist PPADS. TNF-α levels in the spleen were decreased by CNQX and PPADS treatments, implying the involvement of inhibitory neuronal activity in the DVC. In unanesthetized animals, both MAC and EAC generated c-Fos induction in the DVC neurons. However, MAC, but not EAC, was effective in decreasing splenic TNF-α production. These results suggest that the therapeutic effects of acupuncture may be mediated through vagal modulation of inflammatory responses in internal organs.

Effects of inhalable microparticle of flower of Lonicera japonica in a mouse model of COPD.

ETHNOPHARMACOLOGICAL RELEVANCE: Flower of Lonicera japonica (FLJ) is a traditional herbal medicine widely used in East Asia as an anti-inflammatory and anti-oxidative agent. The purpose of this study is to develop an inhalable powder formulation of FLJ and to evaluate its biological effects in a mouse model of chronic obstructive pulmonary disease (COPD). METHODS: Inhalable dry powder containing FLJ was produced by spray-drying with leucine as an excipient. Its aerodynamic properties and anti-inflammatory activities were evaluated using the Anderson cascade impactor (ACI) and a mouse model of COPD, respectively. RESULTS: FLJ microparticle (FLJmp) had a hollow spherical shape in electron microscopy and showed aerodynamic properties suitable for inhalation (fine particle fraction of 54.0 ± 4.68% and mass median aerodynamic diameter of 4.6 ± 0.34µm). FLJmp decreased TNF-α and IL-6 expression in RAW264.7 cells activated by lipopolysaccharide (LPS). In mice challenged with LPS and cigarette smoke solution (CSS) to develop COPD, FLJmp decreased the levels of TNF-α and IL-6 in bronchoalveolar fluidas well as the number of inflammatory cells including neutrophils in peripheral blood. In addition, FLJmp induced recovery of elastin and collagen distribution, reduction of caspase-3 expression in lung tissues of COPD mice. CONCLUSIONS: Inhalational delivery of FLJ using a microparticle system is a promising strategy for the treatment of COPD.

Induction of regenerative responses of injured sciatic nerve by pharmacopuncture therapy in rats.

Although recent studies report that combined treatment of herbal drugs with acupuncture can improve clinical efficacy in traditional oriental medicine, experimental evidence that supports this pharmacopuncture therapy is rare thus far. Here, we investigated the effects of the herbal drug recipe Sciatica 5 (SCTA5) and acupuncture stimulation on gall bladder 30 (GB30) on regenerative responses of injured sciatic nerve in rats. Treatment of cultured dorsal root ganglion (DRG) neurons with SCTA5 improved neurite outgrowth. In vivo regenerative responses, in terms of distal extension of regenerating axons and retrogradely-labeled DRG neurons, were improved by either injury site application of SCTA5 or GB30 acupuncture stimulation and further increased by SCTA5 pharmacopuncture on GB30 acupoint. Moreover, combined treatment of SCTA5 and GB30 was more effective than singular treatments in inducing Cdc2 kinase and accompanying vimentin phosphorylation in Schwann cells of the injured nerve. These results suggest that SCTA5 and GB30 therapies may be cooperative in facilitating axonal regeneration in the injured peripheral nerves.

Acupuncture-stimulated activation of sensory neurons.

Acupuncture is one of the key therapeutics in clinical oriental medicine, and recent studies using experimental animals have begun to provide the pathophysiological basis for the efficacy of acupuncture. Here, we investigated neuronal responses in rodent models given acupuncture stimulation. In both mice and rats, acupuncture stimulation at zusanli (ST36) generated an increased expression of axonal growth-associated protein (GAP-43) in the sensory neurons of the dorsal root ganglion (DRG). Electroacupuncture stimulation at ST36 in rats induced GAP-43 mRNA and protein expression in DRG neurons at the levels of lumbar 4 and 5. Stimulation on a non-acupuncture site as a sham control induced GAP-43 expression as well, but the induction level was lower than it was with acupuncture. We further found that acupuncture stimulation upregulated phospho-Erk1/2 signals in DRG neurons. Electroacupuncture stimulation induced c-Fos expression in the neurons of the dorsal motor nucleus of the vagus nerve (DMV), which was identified by retrograde tracing. These data suggest that acupuncture stimulation may generate physiological effects on the autonomic nervous system via the activation of a somatosensory pathway.

Neuroprotective Activity of Sibjeondaebo-tang on Aß Peptide-Induced Damages.

Background. Sibjeondaebo-tang (SJDBT) has been used to treat diverse disorders including neuropsychiatric disabilities in traditional Korean medicine. Objective. The present study aims to investigate the potential effects of SJDBT on neuroprotection against Aß peptide-induced damage using in vitro culture and in vivo rat brain systems. Materials and Methods. PC12 cell viability was analyzed by MTT assay, and neurite arborizations and caspase 3 protein signals in cultured PC12 cells and in vivo cortical neurons were analyzed by immunofluorescence staining. Phospho-Erk1/2 protein was analyzed by immunofluorescence staining and western blot analysis. Results. In PC12 cells, atrophied cell body and reduced neurite extension by Aß treatment were recovered by SJDBT treatment. Caspase 3 protein signals were increased in Aß-treated PC12 cells, but SJDBT treatment decreased apoptotic cell death. Caspase 3 activation in cortical neurons, which was induced similarly by Aß treatment, was reduced by SJDBT treatment. Furthermore, phospho-Erk1/2 protein levels, which had been decreased by Aß treatment, were elevated in the cortical neurons by SJDBT treatment. Conclusion. These data show that SJDBT may play a role in protecting from damages induced by Aß in neuronal tissue and further suggest that SJDBT can be explored as the potential therapeutic target for AD treatments in human.

Regenerative effects of moxibustion on skeletal muscle in collagen-induced arthritic mice.

In this study, we demonstrate that the direct application of moxibustion significantly enhances muscle regeneration in mice with collagen-induced arthritis (CIA). Twelve Dilute Brown Non-Agouti (DBA)/1 J male mice were randomly divided into the following groups: intact control (n=4), CIA (n=4), and CIA with moxibustion treatment (CIA+moxi, n=4). Mice in the CIA and CIA+moxi groups were immunized twice via intradermal injections of bovine type II collagen (C II) at 3-week intervals. After the second injection, moxibustion was applied to the mouse equivalent of the BL24 and ST36 acupoints with a moxa cone five times/day, every other day (except Sundays), for 3 weeks (a total of 9 treatments were administered). Phospho-Erk1/2, myostatin, TFG-B1, and IGF-1 were analyzed using ELISA. Protein levels in skeletal muscle tissues of the hind limb were analyzed by Western blotting and immunofluorescent staining. Treatment with direct moxibustion led to a marked improvement in CIA and atrophy of individual muscle fibers. Collagen protein signaling in the muscle of the CIA group was stronger than the control and CIA+moxi groups. Myostatin protein expression, as determined by Western blotting and immunofluorescent staining, were stronger in the CIA group compared with the control and CIA+moxi groups. Immunofluorescent staining confirmed that the CIA group had the strongest TGF-B1 protein signals among the three groups. However, in serum analysis the intact control group showed the strongest TGF-B1 protein signaling. RT-PCR analysis of the muscle tissues of the CIA+moxi group showed significant IGF-1 mRNA expression, and the most intense phospho-Erk1/2 protein signaling was detected in the muscle tissues of the CIA group via Western blotting and immunofluorescent staining. These results confirm that the direct administration of moxibustion at Shenshu (BL 23) and Zusanli (ST 36) influences muscle regeneration in the CIA mouse model. Our results suggest that the establishment of the moxibustion mechanism will encourage the clinical application of moxi.

Shengmai-san-mediated enhancement of regenerative responses of spinal cord axons after injury in rats.

Shengmai-san (SMS) is a traditional Chinese medicine used to treat diverse symptoms including cardiovascular and neurological disorders. Here we investigated the effects of SMS on regenerative responses of spinal cord axons in rats that were given contusion injury at the lower thoracic level. The injury cavity was confined to a restricted area by SMS treatment, and the signals of glial scar protein chondroitin sulphate proteoglycan (CSPG) and inflammatory cell marker protein CD11beta were heavily observed within the injury cavity in SMS-treated animals. Anterograde tracing of DiI-labeled corticospinal tract (CST) axons revealed increases in collateral arborization around and within the injury cavity and caudal elongation by SMS treatment. Furthermore, SMS treatment facilitated neurite elongation of dorsal root ganglion (DRG) sensory neurons that were co-cultured with non-neuronal cells prepared from injured spinal cord. Phospho-Erk1/2 was strongly induced in both spinal cord and motor cortical areas after spinal cord injury (SCI), and it was further unregulated in the motor cortex by SMS treatment. In contrast, upregulation of cell division cycle 2 (Cdc2) production by SMS treatment was limited to a local, SCI area. These data suggest that SMS may play an active role in regenerative responses and facilitate axonal regrowth after SCI.

Growth-promoting activity of Sanyak (Dioscoreae rhizoma) extract on injured sciatic nerve in rats.

The present study evaluates the potential effects of Sanyak (Dioscoreae rhizome) on the regenerative capacity of the peripheral sciatic nerve after crush injury in rats. Focal application of Sanyak extract at the injury site increased GAP-43 and Cdc2 protein levels in the distal portion of the injured nerve. Immunohistochemical analysis showed that the signals of phospho-vimentin as Cdc2 substrate protein were almost colocalized with Cdc2. Retrograde DiI-tracing revealed enhancement in distal elongation of regenerating axons. Furthermore, the number of non-neuronal cells was higher in Sanyak-treated animals than saline controls. Thus, these data suggest that Sanyak extract is effective for promoting regenerative responses in injured peripheral neurons.