Adipose-derived stem cells decolonize skin Staphylococcus aureus by enhancing phagocytic activity of peripheral blood mononuclear cells in the atopic rats

Staphylococcus aureus (S. aureus ) is known to induce apoptosis of host immune cells and impair phagocytic clearance, thereby being pivotal in the pathogenesis of atopic dermatitis (AD). Adipose-derived stem cells (ASCs) exert therapeutic effects against inflammatory and immune diseases. In the present study, we investigated whether systemic administration of ASCs restores the phagocytic activity of peripheral blood mononuclear cells (PBMCs) and decolonizes cutaneous S.aureus under AD conditions. AD was induced by injecting capsaicin into neonatal rat pups. ASCs were extracted from the subcutaneous adipose tissues of naïve rats and administered to AD rats once a week for a month. Systemic administration of ASCs ameliorated AD-like symptoms, such as dermatitis scores, serum IgE, IFN-γ+/IL-4+ cell ratio, and skin colonization by S. aureus in AD rats. Increased FasL mRNA and annexin V+/7-AAD+ cells in the PBMCs obtained from AD rats were drastically reversed when co-cultured with ASCs. In contrast, both PBMCs and CD163+ cells bearing fluorescent zymosan particles significantly increased in AD rats treated with ASCs. Additionally, the administration of ASCs led to an increase in the mRNA levels of antimicrobial peptides, such as cathelicidin and β-defensin, in the skin of AD rats. Our results demonstrate that systemic administration of ASCs led to decolonization of S. aureus by attenuating apoptosis of immune cells in addition to restoring phagocytic activity. This contributes to the improvement of skin conditions in AD rats. Therefore, administration of ASCs may be helpful in the treatment of patients with intractable AD.

Oxytocin produces thermal analgesia via vasopressin-1a receptor by modulating TRPV1 and potassium conductance in the dorsal root ganglion neurons

Recent studies have provided several lines of evidence that peripheral administration of oxytocin induces analgesia in human and rodents. However, the exact underlying mechanism of analgesia still remains elusive. In the present study, we aimed to identify which receptor could mediate the analgesic effect of intraperitoneal injection of oxytocin and its cellular mechanisms in thermal pain behavior. We found that oxytocin-induced analgesia could be reversed by d(CH₂)₅[Tyr(Me)²,Dab⁵] AVP, a vasopressin-1a (V1a) receptor antagonist, but not by desGly-NH₂-d(CH₂)₅[DTyr², Thr⁴]OVT, an oxytocin receptor antagonist. Single cell RT-PCR analysis revealed that V1a receptor, compared to oxytocin, vasopressin-1b and vasopressin-2 receptors, was more profoundly expressed in dorsal root ganglion (DRG) neurons and the expression of V1a receptor was predominant in transient receptor potential vanilloid 1 (TRPV1)-expressing DRG neurons. Fura-2 based calcium imaging experiments showed that capsaicin-induced calcium transient was significantly inhibited by oxytocin and that such inhibition was reversed by V1a receptor antagonist. Additionally, whole cell patch clamp recording demonstrated that oxytocin significantly increased potassium conductance via V1a receptor in DRG neurons. Taken together, our findings suggest that analgesic effects produced by peripheral administration of oxytocin were attributable to the activation of V1a receptor, resulting in reduction of TRPV1 activity and enhancement of potassium conductance in DRG neurons.

Juvenile Obesity Aggravates Disease Severity in a Rat Model of Atopic Dermatitis

PURPOSE: There is increasing epidemiological evidence of an association between childhood obesity and atopic dermatitis, but little is known about the underlying mechanism(s). In the present study, we used a rat model of atopic dermatitis to assess whether juvenile obesity, induced by reduction of litter size, aggravated the signs of atopic dermatitis and, if so, whether this aggravation was associated with changes in plasma concentration of adipokines, such as leptin and adiponectin. METHODS: Dermatitis was induced by neonatal capsaicin treatment. Body weight, dermatitis score, serum IgE, skin nerve growth factor (NGF), serum leptin and adiponectin, and cytokine mRNA expression in the skin lesion were compared between small (SL, 5 pups) and large litters (LL, 15 pups). RESULTS: The body weight of juvenile rats up to 6 weeks of age was significantly heavier in the SL group, compared with those in the LL group. The SL group showed more robust development of dermatitis, and higher levels of serum IgE and skin NGF than the LL group. Additionally, the SL group demonstrated higher levels of leptin and pro-inflammatory cytokine mRNA but lower levels of adiponectin than the LL group. CONCLUSIONS: These results suggest a causal link between a decrease in immunological tolerance, induced by juvenile obesity, and aggravation of atopic dermatitis.

Are Spinal GABAergic Elements Related to the Manifestation of Neuropathic Pain in Rat?

Impairment in spinal inhibition caused by quantitative alteration of GABAergic elements following peripheral nerve injury has been postulated to mediate neuropathic pain. In the present study, we tested whether neuropathic pain could be induced or reversed by pharmacologically modulating spinal GABAergic activity, and whether quantitative alteration of spinal GABAergic elements after peripheral nerve injury was related to the impairment of GABAergic inhibition or neuropathic pain. To these aims, we first analyzed the pain behaviors following the spinal administration of GABA antagonists (1 microgram bicuculline/rat and 5 microgram phaclofen/rat), agonists (1 microgram muscimol/rat and 0.5 microgram baclofen/rat) or GABA transporter (GAT) inhibitors (20 microgram NNC-711/rat and 1 microgram SNAP-5114/rat) into naive or neuropathic animals. Then, using Western blotting, PCR or immunohistochemistry, we compared the quantities of spinal GABA, its synthesizing enzymes (GAD65, 67) and its receptors (GABAA and GABAB) and transporters (GAT-1, and -3) between two groups of rats with different severity of neuropathic pain following partial injury of tail-innervating nerves; the allodynic and non-allodynic groups. Intrathecal administration of GABA antagonists markedly lowered tail-withdrawal threshold in naive animals, and GABA agonists or GAT inhibitors significantly attenuated neuropathic pain in nerve-injured animals. However, any quantitative changes in spinal GABAergic elements were not observed in both the allodynic and non-allodynic groups. These results suggest that although the impairment in spinal GABAergic inhibition may play a role in mediation of neuropathic pain, it is not accomplished by the quantitative change in spinal elements for GABAergic inhibition and therefore these elements are not related to the generation of neuropathic pain following peripheral nerve injury.

Behavioral Manifestation of Neuropathic Pain Following Sciatic Nerve Section in Rats

The present study was performed to observe the time course of behavioral signs of painful sensations in sciatic neurectomy animal model and to test the effects of sympathectomy and saphenous nerve section on these behavioral signs. Sciatic nerve was ligated and cut at the mid-thigh level under gaseous anesthesia. The application of von Frey filaments to the medial plantar surface of foot revealed weak and long-lasting mechanical allodynia (until end of test period, 20 weeks PO). Acetone application to the plantar surface of foot was used ti measure the sensitivity to cold stimulation. Cold allodynia which is interpreted as increased response to acetone application developed fairly well and lasted the end of test period (20 weeks PO). The cumulative duration of foot lifts off neutral or cold plate was used to test spontaneous, ongoing pain and was increased until 16 weeks PO and 20 weeks PO respectively. These results suggest that sciatic neurectomy which has been widely used as chronic pain model shows behavioral signs suggsting painful sensations except autotomy, which has been used as index of pain in experimental animal. Surgical sympathectomy performed 1 week after sciatic neurectomy partially reduced the behavioral signs of mechanical allodynia and cold allodynia, suggesting behavioral changes developed following section of sciatic nerve was partially sympathetic dependent. Saphenous nerve section 1 week after sciatic neurectomy almost completely reduced mechanical allodynia and cold allodynia, but did not change spontaneous, ongoing pain. These results suggest that evoked responses such as mechanical and cold allodynia are mediated by saphenous nerve activity and activating and/or maintaining mechanisms of spontanous, ongoing pain and evoked pain may be different.

Neuropathic Pain Behaviors and the Change of Spinal Neuropeptides following Peripheral Nerve Injury in Neonatal Rats

OBJECTIVE: It has been suggested that the occurrence of persistent pain signal during the early postnatal period may alter an individual's response to pain later in life. The aim of this study is to assess whether neonatal nerve injury resulted in long-lasting consequences on nociceptive system in the rat. METHODS: We examined whether neuropathic pain behaviors and the changes of spinal neuropeptides (SP, CGRP, VIP and VIP) induced by peripheral nerve injury within 1 day after birth (Neonate group) were different from those at 8 weeks after birth (Mature group). RESULTS: The Neonate group showed more robust and long-lasting pain behaviors than the Mature group. Immunohistochemical findings demonstrated that spinal SP- & CGRP-immunoreactivities(ir) of the ipsilateral to the contralateral side increased in the Neonate group, whereas those decreased in the Mature group. In addition, increase in spinal VIP- & NPY-ir of the ipsilateral to the contralateral side was more robust in the Mature group than in the Neonate group. CONCLUSION: These results suggest that peripheral nerve injury in the early postnatal period may result in long-lasting and potentially detrimental alterations in nociceptive pathways.

Does HSP70 Induced by Amphetamine Prevent Renal Ischemia/Reperfusion Injury in Rat? / 대한신장학회잡지

BACKGROUND: Heat shock proteins (HSPs) induced by variable kinds of stress produce tolerance to a variety of adverse conditions. However, the protective effect of HSP on ischemia/reperfusion injury (I/R injury) of kidney in vivo remains unclear. The present study was designed to evaluate whether HSP70 induced by hyperthermic preconditioning had renoprotective effect on I/R injury of the kidney in vivo. METHODS: 82 Sprague-Dawley male rats were used. Animals in control group (n=24) were subjected to bilateral occlusion of renal pedicles for 30 or 60 minutes followed by 24-hour reperfusion. In amphetamine (Amp, n=18) and quercetin (Q, n=16) group, amphetamine sulfate, a sympathomimetic drug which can elevate the body temperature as a result of enhanced endogenous lipolysis, and quercetin, a biflavonoid which inhibit the expression of HSP, were injected 4 hours prior to renal ischemia, respectively. In quercetin-amphetamine (QAmp, n=7) group, quercetin was injected 1 hour before administration of amphetamine. AA (n=8) or QQ (n=9) group was identical to Amp or Q group except that sham operation was performed instead of ischemic insult. In all groups, animals were sacrificed prior to or 24 hours after I/R injury. HSP70 induction was confirmed by immunohistochemistry. To assess the I/R injury of kidney, BUN, Cr, histopathologic change of tubular cell and HSP70 expression were evaluated. RESULTS: In Amp group, an increase of BUN and Cr were significantly lower than other groups and less severe renal tubular injury was also observed. In addition, HSP70 was strongly expressed in Amp group, whereas HSP70 was weakly expressed in control group and not expressed in QAmp and Q group. There were no differences in the functional and histologic injuries of kidney after I/R injury between AA, QQ and control group. CONCLUSION: These data demonstrate that the renoprotective effect by amphetamine preconditioning to I/R injury is linked with the expression of HSP70.

Effects of Clonidine and Yohimbine on Neuropathic Pain in a Rat Model

OBJECTIVE: In a rat model of peripheral neuropathy, to determine whether neuropathic pain is related to the alpha-2 adrenergic receptor. METHOD: The neuropathic pain was produced by unilateral transection of the superior caudal trunk between the S3 and S4 spinal nerves. These animals showed the behavioral signs of neuropathic pain in the tail. Two weeks after the neuropathic surgery, tail withdrawal responses to the mechanical stimuli with von Frey hair (2.0 g) were examined 1, 2 and 24 hrs following the administration of clonidine, alpha-2 receptor agonist. One week after the clonidine test, the same behavioral test was done after the administration of clonidine along with yohimbine, alpha-2 receptor antagonist. RESULTS: Clonidine significantly reduced the frequency of tail response and yohimbine reversed the clonidine-induced anti-allodynic effect. CONCLUSION: These results suggest that neuropathic pain is related to the sympathetic nervous system via alpha-2 adrenergic receptor.

Effect of heat shock protein 72 on the generation of reperfusion arrhythmias

The causal relationship between heat shock protein (HSP) and second window of cardioprotective effect is still undetermined. In the present study, we assessed whether HSP-producing substances, amphetamine and ketamine, afforded protection against reperfusion-induced ventricular fibrillation (VF) and these protective effect remained after the inhibition of HSP72 production by quercetin, a mitochondrial ATPase inhibitor. Adult mongreal male cats (n=60, 2.5 ~ 4 kg) were used in this study. Experimental animals were divided into five groups; control group (n=15), amphetamine ('A', n=11) group, ketamine ('K', n=9) group, amphetamine-ketamine ('AK', n=16) group and amphetamine-ketamine-quercetin ('AKQ', n=9) group. Twenty-four hours after the drug treatment, an episode of 20-min coronary artery occlusion was followed by 10-min reperfusion. The incidence of reperfusion-induced VF in the AK and AKQ groups was significantly lower than that in control group (p<0.01). After the ischemia/reperfusion procedure, western blot analysis of HSP72 expression in the myocardial tissues resected from each group was performed. HSP72 production in the AK group was marked, whereas HSP72 was not detected in the AKQ and control groups. These results suggest that the suppressive effect against reperfusion-induced VF induced by amphetamine and ketamine is not mediated by myocardial HSP72 production but by other mechanisms.

Effects of Exercise on Neuropathic Pain in an Experimental Model of Peripheral Neuropathy

OBJECTIVE: The purpose of this study was to evaluate the short- and long-term effects of exercise on neuropathic pain. METHOD: Pain responses between rats in the exercise and control groups were compared to evaluate the effects of exercise in neuropathic pain. Materials consisted of 30 male Sprague-Dawley rats (8 weeks old, 180~200 g), which were divided into an exercise group (n=15) and a control group (n=15). Neuropathic pain was produced by partially injuring the nerve innervating the tail. Running exercise was given on a Rota-rod treadmill exercise machine for 3 weeks (3.1 Km/day, 6 cycle of 9 minutes exercise and 1 minute rest). Behavioral reactions to mechanical allodynia were checked using a von Frey hairs of 2.0 g (19.6 mN) bending force at 10 minutes, 1 hour and 24 hours post-exercise to evaluate the short term effects of exercise. Behavioral reactions to mechanical and thermal allodynia with 4 degrees C or 40 degrees C were evaluated 7, 14, 21 and 28 days following exercise. RESULT: The exercise group exhibited less tail-flick frequencies to mechanical stimulation from 58.8+/-6.8% to 41.1+/-5.4%, 37.6+/-13.2% at 1 and 24 hours post-exercise compared to the control group, but there was no significant difference between the groups at weeks 1 through 4. In the exercise group, the decrease of tail-flick frequencies were blocked by naloxone (2 mg/kg i.p.). It is suggested that long-lasting muscle exercise (e.g. running) which influences central endorphin mechanisms giving analgetic effects. CONCLUSION: The results of this study support the hypothesis that the exercise can reduce neuropathic pain in the acute stage.

Effects of Clonidine and Epinephrine on Neuropathic Pain in an Experimental Animal Model for Peripheral Neuropathy

OBJECTIVE: To evaluate the effect of clonidine on the experimental neuropathic pain model and to observe whether neuropathic pain is related to the sympathetic nervous system in this model by reversal of allodynia with administration of epinephrine. METHOD: The neuropathic pain was produced by unilateral transection of the superior caudal trunk innervating the rat's tail. Tail withdrawal responses based on mechanical (withdrawal frequency to bending force of von Frey hair 2.0 g) and the thermal (withdrawal latency to tail immersion in a 4degrees C or 40degrees C water with a cut-off time of 15 seconds) stimuli were used. Experiments were performed two weeks after surgery when neuropathic pain had fully been developed. Experimental group by administration of clonidine was examined by tail withdrawal responses at Day 1, Day 3 and Day 5. After one week of wash-out period, reversal of allodynia by administration of epinephrine was examined by the same test. RESULTS: Clonidine significantly decreased the frequency of withdrawal with the mechanical stimuli compared with control (P<0.01), but did not significantly decrease with the cold or warm stimuli. Epinephrine tended to aggravate the mechanical allodynia, but it was not significant compared with the control. CONCLUSION: Clonidine may relieve mechanical allodynia in neuropathic pain, but the mechanism of neuropathic pain that is related to the sympathetic nervous system in this experimental model may be unreliable.

Expression of protein kinase C-gamma mRNA on the spinal cord in a rat model of neuropathic pain / 대한해부학회지

Partial peripheral nerve injury occasionally results in neuropathic pain, including spontaneous burning pain and increased sensitivity to sensory stimuli such as hyperalgesia and allodynia. The pathophysiological mechanisms underlying this disease are poorly understood and the available treatments unsatisfactory. Presently, the neuropathic pain is believed to result from an increase in the excitability of the dorsal horn neurons (central sensitization), which is induced by abnormal signals from injured afferents. PKC-gamma is known to play a pivotal role in central sensitization following peripheral nerve injury. In the present study, we examine the expression of PKC-gamma mRNA of the spinal dorsal horn after neuropathic injury. There was no significant difference of PKC-gamma mRNA between lesion and control sides. These results suggest that PKC-gamma mRNA is not a key factor for the generation of neuropathic pain.

Role of Protein Kinase C in Normal and Preconditioned Myocardium

No abstract available.

Hypothermic preconditioning lowers the incidence of hypothermic arrest in neonatal rat

This study was performed to examine 1) Whether hypothermic cardiac arrest produces myocardial HSP72 expression; 2) And if, whether it serves to protect the heart against the subsequent hypothermic arrest. In the present study, neonatal rats were placed in an icebath to induce hypothermia. To determine whether hypothermic cardiac arrest produces myocardial HSP72, experimental animals were subjected to 10-min hypothermic insult before the extraction of the heart. The intervals between the insult and extraction were 1 (1 HR), 4 (4 HR), 8 (8 HR), 24 (24 HR) or 72 (72HR) hours. A minimal amount of HSP72 was detected in control, 1 HR and 72 HR groups. In contrast, 8 HR and 24 HR groups showed a significant level of HSP72 expressions. To assess the cardioprotective effect of HSP72 against hypothermic cardiac arrest, we compared the proportion of recovery from the arrest between control and preconditioned (PREC) animals. Control animals were subjected to 20-min hypothermic insult, while PREC group was preconditioned by 10-min hypothermic insult 8 hours before the 20-min test hypothermic insult. Resuscitation rate from cardiac arrest induced by the 20-min hypothermic insult in PREC group was significantly higher than that in controls. These results suggest that the cardioprotective effect of hypothermic preconditioning is associated with an increase in HSP72 expression.

Gradual reperfusion lowers the incidence of reperfusion-induced ventricular fibrillation in a cat model of regional ischemia

Blood flow restoration to ischemic zone of the heart is essential to salvage of ischemic tissue. However, there is a large body of evidence documenting that the reperfusion can induce reperfusion injury like reperfusion-induced malignant arrhythmias. In the present study, employing a cat model of regional cardiac ischemia, we examined if reperfusion rendered in a gradual fashion could lower the incidence of reperfusion-induced ventricular fibrillation (VF), which usually precipitated within a few to several tens of seconds after abrupt reperfusion. The experiments were conducted with male mongrel cats (n=46, 2.5-5 kg). The animals in the control and 30 MIN groups were subjected to an episode of 20- and 30-min left anterior descending coronary artery occlusion, respectively, followed by abrupt reperfusion. The animals in 5 G and 10 G groups received gradual reperfusion over a 5- and 10-min period, respectively, following a 20-min occlusion. The proportion of animals that exhibited VF during the reperfusion phase was 11/15 in the control, 7/10 in the 30 MIN, 5/10 in the 5 G and 2/11 in the 10 G groups. The incidence of VF in the 10 G group was significantly lower than that in the control or 30 MIN group subjected to abrupt reperfusion. These results suggest that the gradual reperfusion is a useful procedure against reperfusion-induced VF.

The effect of the occlusion of the left bronchial artery on the production of HSP70 cat lung / 결핵

Several stresses are known to induce synthesis of heat shock protein. The present study was performed to see whether pulmonary ischemia, induced by the bronchial artery occlusion, produced HSP70 in cat lung. To this aim we compared experimental and control groups of cats with respect to the HSP70 production in the lung. Experimental animals were subjected to 10-min bronchial artery occlusion followed by reperfusion. The interval between the end of the occlusion and the end of the reperfusion was 1 hour, 4 hours and 8 hours, whereas control animal was not subjected to any manipulation except anesthesia. According to the interval differences, experimental animals were divided into 1HR, 4HRs and 8HRs groups. To determine the induction of HSP70 in each group, total proteins of lung tissues were extracted and separated by PAGE electrophoresis. Immunoblotting with a mouse monoclonal anti-HSP70 IgG antibody revealed that HSP70 was not detected in the pulmonary tissues resected from control, 1HR or 4HRs groups. In contrast, HSP70 expression in 8HRs group was marked. These results suggest that pulmonary ischemia by the bronchial artery occlusion produces HSP70 in a delayed

The Effect of Hypoxic-Preconditioning on the Reperfusion-Induced Arrhythmias in the Cat Hearts

BACKGROUND: Ischemic preconditioning(a prior short period of coronary artery occlusion) has been known to have protective effects on ischemia-induced myocardial injury. The purpose of this study was to investigate the effects of hypoxic preconditioning or ischemic preconditioning on the

Determinant role of the severity of hypoxia in the induction of reoxygenation injury in cat lung

Although reoxygenation is the best way to salvage hypoxic tissues, reduced oxygen species (ROS) generated during reoxygenation are known to cause further tissue injuries and the induction of heat shock proteins (HSPs). The present study was undertaken to determine any causal relationship between the severity of hypoxia and the opposite outcomes, either beneficial or detrimental, of the subsequent reoxygenation by measuring the HSP72. To this aim, one group (6 male cats, 2.5 ~ 3.5 kg) was subjected to a 5-min episode of hypoventilation (H, ventilation rate: 5/min) for the induction of slight hypoxia and the other group (6 male cats, 2.4-3.7 kg) was subjected to a 5-min episode of apnea (A) for severe hypoxia. Each 3 animals from both groups received a 10-min episode of ventilation with 95% O2 (O), whereas the remainder did not. After these procedures, all animals were allowed to be ventilated within physiological range for 1, 4, or 8 hours (1H, 1HO, 4H, 4HO, 8H, 8HO, 1A, 1AO, 4A, 4AO, 8A and 8AO groups). Control animals did not receive any manipulation. The arterial blood pCO2 was significantly higher just after apnea than hypoventilation, while pO2 and pH were significantly lower just after apnea than hypoventilation. Western blot analysis revealed that the magnitude of HSP72 synthesis is larger in 1H, 4H and 8H groups than in 1HO, 4H and 8HO groups, respectively. In contrast, 1AO, 4AO and 8AO groups more induced HSP72 than 1A, 4A and 8A groups, respectively. These results suggest that the reoxygenation is beneficial after slight hypoxia but detrimental after severe hypoxia.