Analgesic action of suspended moxibustion in rats with chronic visceral hyperalgesia correlates with enkephalins in the spinal cord

Rats that modeled chronic visceral hyperalgesia received suspended moxibustion at bilateral Tianshu （ST25） and Shangjuxu （ST37） once daily over a period of 7 days. Results show that suspended moxibustion significantly depressed abdominal withdrawal reflex scores and increased enkephalin concentration in the spinal cord. The experimental findings suggest that spinal enkephalins contributed to the analgesic effect of suspended moxibustion in rats with chronic visceral hyperalgesia.

THE INTERACTION BETWEEN PERIAQUEDUCTAL GRAY AND NUCLEUS ACCUMBENS IN ACCELER-ATING THE RELEASE OF ENKEPHALINS AND β-ENDORPHIN

There is a growing body of evidence showing that periaqueductal gray (PAG) and nucleus accumbens (N. accumbens) play important roles in the modulation of nociception. In our previous study of the mechanisms of morphine analgesia we found that opioid antagonist naloxone injected into either of the two nuclei, PAG or N. accumbens, could

Opioid growth factor and the treatment of human pancreatic cancer:A review

Opioid growth factor (OGF), chemically termed [Met5]-enkephalin, and its receptor, OGF receptor (OGFr), form a biological axis that tonically regulates cell proliferation by delaying the G1/S interface of the cell cycle under homeostatic conditions or in neoplasia. Modulation of the OGF-OGFr pathway mediates the course of pancreatic cancer, with exogenous OGF or upregulation of OGFr repressing growth of human pancreatic cancer cells in culture and in nude mice. OGF therapy alone or in combination with standard chemotherapies such as gemcitabine and 5-fluorouracil results in enhanced inhibition of DNA synthesis and tumor growth. Molecular manipulation of OGFr confirms that the receptor is specific for OGF’s inhibitory action. Preclinical studies have warranted Phase I and Phase II clinical trials using OGF infusions as a treatment for patients with advanced, unresectable pancreatic cancers. OGF, an endogenous neuropeptide, is a safe, non-toxic, and effective biotherapy that utilizes the OGF-OGFr axis to mediate pancreatic tumor progression.

Effects of electric field stimulation on endogenous enkephalin content in artery wall

After 20min electric field stimulation,L-enkephalin （L-ENK） content in the rabbitear artery declined from the control value of 38.99±17.29 to 12.55±1.96pg/mg of wet tissue（n=12）,and M-enkephalin （M-ENK） content from 134.67±8.11 to 73.68±17.96pg/mg of wet tis-sue （n=6）,both being significantly decreased.This effect was not influenced after blockade ofadrenergic receptor by yohimbine.Contractions of artery strips induced by electric field stimulationwere enhanced distinctly after pretreatment of the artery strips with naloxone,an opioid receptorblocker,and with pronase,but not after pretreatment with trypsin.The results suggested that the re-lease of endogenous enkephalins in the ear artery could be induced by electric field stimulation andx2-receptor might not participate in the release course,The release amount of ENK from the ar-terv strips increased when the stimulus freduency increased within the range of 2 Hz to 8 Hz.

Serotonin样,Substance P样,Enkephalin样反应阳性神经元胞体在大鼠中缝核簇内的分布

本文用PAP法研究了Serotonin样,Substance P样,Leucine—Enkephalin样和Methionine—Enkephalin样反应阳性神经元胞体在大鼠中缝核簇内的分布。证明Serotonin存在于中缝苍白核、中缝隐核、中缝大核、中缝桥核、中央上核和中缝背核等处的某些神经元胞体内。以中、小细胞为主,在中缝大核内有少数大细胞。细胞形态为梭形、圆形、卵圆形以及多极细胞。Substance P样反应阳性胞体在上述中缝核内也被发现,其胞体的大小、形态和分布与serotonin样反应阳性胞体相似,但数目稍少。Leucine—Enkephalin样反应阳性胞体存在于中缝苍白核、中缝隐核、中缝大核、中缝桥核、中缝背核和中央上核内,但Methionine—Enkephalin样反应阳性细胞体仅在中缝苍白核和中缝隐核的吻侧部以及中缝大核内发现。后两者均以大,中型多极细胞为主。

Plasma L-ENK, AVP, ANP and serum gastrin in patients with syndrome of Liver-Qi-stagnation

AIM To investigate the pathophysiologic basis of syndrome of Liver Qi stagnation and parameters for clinical differentiation. METHODS Plasma L ENK, AVP, ANP and serum gastrin were determined by RIA in 84 patients with neurasthenia, mastodynia, chronic gastritis, and chronic cholecystitis presenting the same syndrome of Liver Qi stagnation in traditional Chinese medicine (TCM). Healthy subjects served as controls in comparison with patients having the same syndrome but with different diseases. RESULTS Among the patients with Liver Qi stagnation, the plasma L ENK, ANP and gastrin levels were 38 83ng/L ± 6 32ng/L , 104 11ng/L ± 29 01ng/L and 32 20ng/L ± 6 68ng/L , being significantly lower than those in the healthy controls ( t =3 34, 6 17, 4 48; P <0 01). The plasma AVP of the patient group ( 52 82ng/L ± 19 09ng/L ) was significantly higher than that of the healthy controls ( t =5 79, P <0 01). The above changes in patients having the same symptom complex but different diseases entities showed no significant differences, P >0 05. CONCLUSION The syndrome of Liver Qi stagnation is closely related to the emotional modulatory abnormality of the brain, with decrease of plasma L ENK, ANP and gastrin, and increase of plasma AVP as the important pathophysiologic basis.

Effects of altitude hypoxia on contents of TXB_2, 6-keto-PGF_(1α) and LENK and their correlations with lung injury induced by pyosepticemia

Objective: To explore the effects of altitude hypoxia on septic pulmonary injury (3080 m above sea level). Methods : A model of pyosepticemia was established by cecal ligation and puncture. Thromboxane A2 (TXA2),prostacyclin (PGI2) of the venous plasma and leucine-enkephalins (L-ENK) in the arterial walls were measured using radioiizimunoassay in 31 rabbits. Mouse morrality was observed. Results: 24 h mouse accumulative mortality (97. 1% )was greater than that of control (35%). TXB2 content was 4 times as much as in control group and 6-keto-PGF1α level was elevated too. These changes of arachidonic metabolism could be blorked by indomethacin. The contents of LENK in arterial walls of puhiionary, mesenteric and renal arteries dropped significantly in comparison with control group (P<0. 01 ). The albumin content in the bronchoalveolar lavage fluid rase significantly in comparison with control group (P<0. 01 ). Conclusion : The results demonstrate that the metabolites of cyclcoxygenase are not the major mediator of puhiionary permeability in the experiment. The alteration of TXA2 and L-ENK contents may be one of the mechanisms, by which the puhiionary artery pressure and pulmonary vascular resistance are increased during sepsis.

Plasma leucine enkephalin content in patients with “Liver-blood deficiency” syndrome and clinical significance

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Effects of methionine enkephalin on immune enhancement by reducing myeloid derived suppressor cells and reprogramming liver metabolism in colon cancer mice

OBJECTIVE To investigate enhanced immune function of methionine encephalin(MENK)and its anti-tumor mechanism in CT26 colon cancer mouse model.METHODS 3×10~6CT26 cells were implanted subcutaneously in BALB/c mice.Four days after,MENK was peritoneally administrated at the concentration of 20 mg·kg^(-1) for 14 d.The percentage of MDSCs in bone marrow,spleen,blood,tumor and liver were detected by flow cytometry.Non-esterified fatty acid(NEFA),triglycerides(TG)and total cholesterol(T-CHO)in liver homogenate were tested by a NEFA test kit,a TG test kit and a T-CHO test kit respectively.qRT-PCR and Western blot were used to measure m RNA and protein levels of inflammation-,glycometabolsim-and lipometabolsim-associated indexes in liver.RESULTS MENK decreased percentages of MDSCs in bone marrow,spleen,blood and tumor in colon cancer mice.MENK-treated mice displayed elevated ratio of CD4^+T and CD8^+T cells in spleen as well as increased T and B lymphocytes proliferation.Meanwhile,MENK also ameliorated liver damage reflected by lower levels of GPT and GOT in serum and reduced risks of cancer-associated index including inflammation,high lipid and high glucose.Furthermore,MENK lowered down the levels of NEFA,TG and T-CHO in liver homogenate.MENK treatment decreased expression of p-STAT3,increased expression of p-AKT,IRS1 and Glut4 at protein level as well as reduced lipogenesis-associated genes and elevated glycolysis-associated genes in liver of tumor bearing mice.Also,abated expression of genes associated with MDSCs generation(M-CSF,GM-CSF,IL-6,IL^(-1)β)and migration(S100A9,KC)was observed within shrunken subcutaneous tumor by MENK intervention.CONCLUSION MENK has the ability to strength immune function against colon cancer by reducing MDSCs and improving liver metabolism.

Regulation of vascular function by opioid receptor in the arterial wall during hemorrhagic shock in rats

Objective: The aim of this study was to investigate the action of the arterial wall opioid receptor in vascular function regulation during shock in rats. Metbods : Experiments were performed on rat model of hemorrhagic shock in vivo and arterial strips in vitro. Results: The mean arterial pressure of the rat was reduced rapidly to 3. 99 kpa by bleeding and was managed to maintain at that level. The mean maximum bleeding volume of the naloxone group was 25. 90±4. 23 ml/kg. It was significantly more than that of the control group which was 20. 26 ± 4. 43 ml/kg (P<0. 05). The vasoconstrictive phase of shock was 50. 00± 11. 53 min in the naloxone group and 31. 68 ± 9. 98 min in the control group. The differences between the two groups were considered significant (P<0. 05). The contents of leucine enkephalin (L-ENK) in the rat's arteries were measured by radioimmunoassay , which increased greatly during hemorrhagic shock. We found that the contractions of the rat's thoracic aorta strip evoked by electric field stimulation decreased significantly in the hemorrhagic shock group as compared with the control's. After giving naloxone (1. 37 ×10-5mol/L) , the enhancement rates of contraction of the shocked rat's artery strips increased greatly as compared with the control strips (P<0.01). Conclusion : The results indicated that opioid receptors in the arterial wall played a role in the decrease of vasoconstriction during hemorrhagic shock. The increasing content of enkephalin in the artery wall during hemorrhagic shork could be one of the important causes why vasoconstriction declined in hemorrhagic shock.Naloxone could act directly on the opioid receptors to block the inhibitory effects of opioid peptide on vasoconstriction and so to increase blood pressure and defy shock.

EFFECT OF METHIONINE ENKEPHALIN ON MIGRATION OF MACROPHAGES FROM MICE WITH IMPAIRED LIVER

Objective To observe the effects of methionine enkephalin (M Enk) on migration of macrophages from mice with impaired liver and its immunomodulatory mechanisms. Methods Liver of mice was impaired by feeding CCl 4 and macrophage migration inhibitory factor (MMIF) was produced by Con A stimulated spleen lymphocytes. Inhibition of macrophage migration was measured in reaction system by adding M Enk. Results Migration of macrophages in both liver impaired and control group were suppressed by MMIF, but the suppression might be reversed by adding 1 μmol/L M Enk ( P <0.05).M Enk could significantly inhibit in vitro both of the combination of MMIF with macrophages and production of MMIF from lymphocytes ( P <0.01).Macrophages from liver imparied group showed a higher sensitivity compared to the control group ( P <0.05).Conclusion The study suggests that opioid peptieds play an important role in the modulation of the immune response under stress as liver impairment.

RELATIONSHIP BETWEEN ACUPUNCTURE ANALGESIA AND MET-ENKEPHALIN OR DYNORPHIN

subjective: The effect of 4～5 Hz electroacupuncture (EA) on alterations of both met enkephalin (MEK) and dynorphin (Dyn) in the patient plasma or mouse spinal cord and its relation with analgesic effect were studied.Methods: In acupuncture clinic 10 patients with acute pain were treated with 4 Hz EA at Zusanli(ST 36) and/or Hegu(LI 4) acupoints for 30 min. 20 BALB/C mice were randomly divided into 2 groups: a. EA group(n=10), treated with 4～5 Hz EA at bilateral “Zusanli"(ST 36) for 15 min; b. control group(n=10) treated with no EA, but also restrained for 15 min. Before and after EA or restraining acupoints, the pain threshold of the patients or mice was detected. 10 μl of the patient plasma before and after EA and each mouse spinal cord suspension, of the 2 groups were blotted onto nitrocellulose membrane (NCM) respectively. The protein dot blot signals were detected by immunoreactivity (IR) and using Shimadu TLC Scanner and analyzed statistically. Results: The results showed that an increase in patient plasma MEK IR or Dyn IR and a decrease in mouse spinal MEK IR or Dyn IR could be detected, and the alteration of plasma or spinal MEK IR was more significant than that of plasma or spinal Dyn IR. There was a positive correlation in alteration between plasma or spinal MEK IR and plasma or spinal Dyn IR with respective parallel levels in individuals. The increased plasma MEK IR or the decreased spinal MEK IR was positively or negatively correlated with the analgesic effect, while the correlation between plasma or spinal Dyn IR and analgesic effect was insignificant. Conclusion: The results suggest that under lower frequency EA the met enkephalin may play an important role in analgesia.

CLINICAL STUDY ON ACUPUNCTURE TREATMENT OF STOMACH CARCINOMA PAIN

Clinical observation on 48 cases of stomach carcinoma pain indicated that acupuncture including filiform needle group and point-injection group had better therapeutic effects in treatment of stomach carcinoma pain when patient's mind was concentrated at the site of disease. After treatment for 2 months, the long-term effective rates of analgesia in both the filiform needle group and the point-injection group were similar to that in the western medicine group, all being about 81%. While the long-term markedly effective rates in the two groups were superior to that in the western medicine group. Life quality of the patients in all the groups were improved. The toxic action and side effects caused by chemotherapy were prevented, the high viscous state showed by indexes of blood rheology was improved, and the lowered Cu-Zu-SOD activity in erythrocytes in patients of stomach carcinoma was increased in the filiform needle group and the point-injection group. Based on the results of clinical study, we consider that acupuncture analgesic effect on stomach carcinoma is related to the increase of PLEK, improvement of cellular immune function and the elevation of life quality after acupuncture.

Acupuncture-related techniques for the treatment of opiate addiction:a case of translational medicine

Drug addiction is a chronic brain disorder characterized by withdrawal symptoms that occur during drug abstinence and a high tendency of relapse.Compared with the currently available pharmacological interventions,acupuncture therapy has the potential to help drug addicts stay away from drugs without major adverse side effects.It has taken decades of research to optimize the parameters of electrical acupoint stimulation for detoxification and for relapse prevention,as well as to establish a safe and easy procedure by which drug addicts can use it on themselves.The discovery that acupuncture can trigger the release of opioid substances from the brain in the 1970s provided the inspiration.Following this,basic research on animals made it possible to understand the mechanisms of action and establish the procedure for treating drug addictions.This article reviews the past,present,and foreseeable future regarding the use of acupuncture-related technique for the treatment of opiate addiction from the perspective of translational medicine.

Construction and identification of immortalized rat astrocyte cell line expressing enkephalin

Objective:To provide a sound cell source for further ex-vivo gene therapy for chronic pain,we attempt to develop an immortalized rat astrocyte cell line that expresses enkephalin regulated by doxycycline. Methods:Retrovirus infection method was employed to develop an immortalized rat astrocyte cell line that could express enkephalin regulated by doxycycline.The hPPE gene expression level of immoralized astroyte cells(IAC)/ hPPE was detected by RT-PCR,indirect immunofluorescence staining and radioimmunoassay. Results:IAC carrying Tet-on system transfected with preproenkephalin gene could secrete enkephalin that was regulated by doxycycline in a dose-dependent manner and hPPE gene activation could be repeated in on-off-on cycles through administration or removal of doxycycline. Conclusion:An immortalized rat astrocyte cell line that secrete enkephalin under the control of doxycycline is established successfully,which provides a research basis for transgenic cell transplantation for analgesia.

Involvement of δ-and μ-opioid receptors in the delayed cerebral ischemic tolerance induced by repeated electroacupuncture preconditioning in rats

Background Preconditioning with repeated electroacupuncture （EA） could mimic ischemic preconditioning to induce cerebral ischemic tolerance in rats. The present study was designed to investigate whether mu（μ）-, delta（δ）- or kappa（κ）-opioid receptors are involved in the neuroprotecUon induced by repeated EA preconditioning. Methods The rats were pretreated with naltrindole （NTI）, nor-binaltorphimine （nor-BNI） or D-Phe-Cys-Tyr-D- Trp-Om-Thr-Pen-Thr-NH2 （CTOP）, which is a highly selective δ-, κ- or μ-opioid receptor antagonist respectively, before each EA preconditioning （30 minutes per day, 5 days）. Twenty-four hours after the last EA treatment, the middle cerebral artery occlusion （MCAO） was induced for 120 minutes. The brain infarct volume was determined with 2,3,5-tdphenyltetrazolium chloride staining at 24 hours after MCAO and compared with that in rats which only received EA preconditioning. In another experiment, the met-enkephalin-like immunoreactivity in rat brain was investigated by immunohistochemistry in both EA preconditioning and control rats. Results The EA preconditioning reduced brain infarct volume compared with the control rats （P=-0.000）. Administration of both NTI and CTOP attenuated the brain infarct volume reduction induced by EA preconditioning, presenting with larger infarct volume than that in the EA preconditioning rats （P〈0.001）. But nor-BNI administration did not block the infarct volume reduction induced by EA preconditioning, presenting with smaller infarct volume than the control group rats （P=-0.000）. The number of met-enkephalin-like immunoreactivity positive neurons in the EA preconditioning rats was more than that of the control rats （P=-0.000）. Conclusion Repeated EA preconditioning stimulates the release of enkephalins, which may bind 5- and p-opioid receptors to induce the tolerance against focal cerebral ischemia.

Endogenous enkephalin does not contribute to the cerebral anti-hyperalgesic action of gabapentin

The aim of this study was to investigate the role of endogenous enkephalin in the cerebral antihyperalgesic action of gabapentin.Neuropathic pain models and antihyperalgesic effect of gabapentin were confirmed by the presentation and changes of mechanical allodynia and thermal hyperalgesia of operated mouse hind paws.The results suggested that endogenous enkephalin may not be involved in the antihyperalgesic effect of gabapentin.

DEPRESSION OF RETRIEVAL OF AN OPERANT CONDITIONING IN RATS BY ENKEPHALIN

It is established that opioid peptides are involved in the regulation of various physiological processes including the higher function of the brain. We have reported that peripheral administration of enkephalin impaired memory retention in mice. This effect was reversed by naloxone. We further found that bestatin, an aminopeptidase inhibitor, when injected peripherally or into amygdala introduced memory impairment which mimiced the effect of enkephalin. The present study examined the influence of hippocampal and peripheral enkephalin on operant defence conditioning in rats, with an attempt to detect the action of central enkephalin on memory.