Objective To construct contains human interleukin-10 gene recombinant lentiviral-vector(LV-IL-10)and to form a basis to further explore the therapy of chronic pain.Methods hIL-10 gene fragment was isolated and amplified from pCYIL-10 plasmid by PCR,and was cloned into pWPXL-GFP.The inserted hIL-10 fragment was verified by Pme I digestion and DNA sequencing.The recombinant plasmid pWPXL-IL-10-GFP,envelope plasmid pMD2.G and packaging plasmid psPAX2 were cotransfected into 293T cells,to pack out lentivirus particle that has the ability of duplicated-deficiency,then virus titer determination was undertaken.Results The 530bp IL-10 gene fragment was amplified from pCYIL-10 plasmid by PCR,and was recombinated into pWPXL-GFP plasmid.DNA sequencing confirmed that the cloned gene segment was 100% homologous to the published hIL-10 sequence in genebank.High titer(2?1010)and highly purified lentiviral particles was obtained.Conclusions The lentivirus vector LV-hIL-10 was constructed successfully,which form a basis of research of chronic pain therapy.

Objective To determine the changes in hemodynamics and plasma PGI2, TXA2 levels caused by uterus traction during operation under continuous spinal anesthesia. Methods Thirty ASA Ⅰ -Ⅱpatients undergoing hysterectomy under continuous spinal anesthesia (CSA) were studied. Aged ranged from 28 to 65 years, height from 150-171 cm and body weight 40-72kg. The patients were premedicated with intramuscular phenobarbital sodium 0.1g and atropine 0.5mg 30 min before surgery. Continuous spinal anesthesia (CSA) was performed at L2-3 with 22G Spinocath ( Braun) . Hyperbaric bupivacaine 0.5% (0.75 % bupivacaine 4ml+ 10 % glucose 2ml) was injected and the height of block was maintained at T8 . Venous blood samples were taken before anesthesia (T0 ), before skin incision ( T1 ), before traction on uterus (T2 ), uterus was being pulled for 10min (T3 ) and 15 min after the end of surgery (T4 ) for determination of 6-keto-PGF1** and TXB2 . BP, HR, SpO2 and ECG were continuously monitored during surgery. Results Continuous spinal anesthesia provided perfect analgesia and satisfactory muscle relaxation. MAP and HR decreased significantly when uterus was pulled (P

It is necessary to use objective and accurate methods to assess the changes of the consciousness of patients emergencing from general anesthesia. In this way, adverse medications during the waking period can be avoided, and it can ensure the stable and safe recovery of consciousness of the patients, quickly remove the adverse factors affecting the patients, and strive to reduce the occurrence of complications during the waking period. This article briefly reviews the research progress of bispectral index and other common clinical anesthesia depth monitoring techniques used to assess the changes of consciousness of patients awakening from general anesthesia, and explores the regular pattern of recovery of consciousness in patients awakening from general anesthesia, in order to reduce complications in the recovery period .

OBJECTIVE: To compare the hemodynamics and post-anesthetic recovery of total intravenous anesthesia (TIVA) with remifentanil or fentanyl combined with propofol administered by target controlled infusion (TCI) in neurosurgery. METHODS: A total of 80 patients undergoing selective neurosurgery were randomly divided into a remifentanil group (Group R, n=40) and a fentanyl group (Group F, n=40). In Group R,remifentanil and propofol was administered by TCI and the blood concentration were 3 approximately 5 microg/L and 3 approximately 5 mg/L each. In Group F, fentanyl was continuously infused at 2 approximately 3 microg/(kg.h) and propofol was administered by TCI with the same blood concentration as that in Group R.Vecuronium was injected at intervals to maintain muscle relaxant.Mean arterial pressure and heart rate during the anesthesia and post-anesthetic recovery were recorded. RESULTS: Mean arterial pressure of all the patients was decreased significantly from induction of anesthesia to termination of operation compared with that before the induction( P<0.01). The heart rate of Group R was increased obviously from recovery of respiration to extubation and heart rate of Group F was decreased obviously from fixed headframe to termination of operation compared with that before the induction (P<0.01). But there was no significant difference between the 2 groups (P>0.05). The eyes opened and extubed time of Group R were decurtated obviously and the scores of pain were increased significantly (P<0.01). CONCLUSION TIVA with remifentanil or fentanyl combined with propofol administered by TCI in neurosurgical operation can provided steadible hemodynamics. Resuscitation of remifentanil with propofol administered by target controlled infusion were more quickly but the scores of pain were more higher than that of fentanyl.
Adolescent ; Adult ; Aged ; Anesthesia Recovery Period ; Anesthesia, Intravenous ; methods ; Anesthetics, Intravenous ; administration & dosage ; Craniotomy ; Female ; Fentanyl ; Humans ; Infusion Pumps ; Male ; Middle Aged ; Neurosurgical Procedures ; Piperidines ; Propofol ; Remifentanil ; Vecuronium Bromide ; administration & dosage ; Young Adult

OBJECTIVE: To construct the recombinant lentivirus vector containing short hairpin RNA (shRNA) inhibited DREAM expression and to investigate the gene therapy of neuropathic pain by inhibiting the expression of DREAM gene by RNA interference. METHODS: An effective short hairpin RNA targeting to rat DREAM was cloned into the plasmids on the base of Lentivirous vectors, pKCSHR-Puro/GFP, and both of the pKCSHR-Puro/GFP-DREAM and Lentivector package plasmids mix were transferred into the 293T cells. The culture supernatant was harvested, and the virus titer was detected 48 hours after transferring. Thirty-six sheer breed pathogen free adult Sprague Dawley rats were randomly divided into 6 groups (6 in each group): normal control group (N); sham-operated group (S); CCI group (C0 group):CCI model without any intervention; Saline control group (C1 group); empty vector control group (C2 group); and LV-shRNADREAM lentiviral vector treatment group (C3 group). The rats in the last 3 groups respectively accepted injection of normal saline, blank vector, LV-shRNADREAM lentiviral vector in the subarachnoid on the 7th day after CCI, and the pain behavior was observed after 3, 7, 10, 14, 21 d after CCI. Green fluorescent protein (GFP) expression was detected by fluorescence microscope and the contents of DREAM mRNA and DREAM protein were detected by Realtime PCR and Western blot respectively in the rat lumbar spinal cord. RESULTS: The short hairpin RNA sequences targeting at rat DREAM were cloned into the vectors, and an entry clone and an expression clone were constructed successfully confirmed by sequence analysis. Lentiviral packaging was successful in 293 T cell line and the transfection titer of the lentivirus was 1 x 10(8) IFU/mL. LV-shRNADREAM lentivirus vector was transfected successfully in the rat spine with Intrathecal injection of LV-shRNADREAM. Compared with the other 3 groups, heat pain threshold and mechanical pain threshold in Group C3 improved significantly (P<0.01), and the expression of DREAM mRNA and DREAM protein in the lumbar spinal cord in Group C3 were lowered significantly (P<0.01). CONCLUSION Lentivirus vectors containing rat DREAM gene are constructed successfully, and lentivirus mediated shRNA can inhibit the DREAM expression in the rat spine, which may prove to be an effective method for neuropathic pain.
Analgesia ; methods ; Animals ; Base Sequence ; Genetic Therapy ; methods ; Genetic Vectors ; genetics ; Kv Channel-Interacting Proteins ; biosynthesis ; genetics ; Lentivirus ; genetics ; metabolism ; Male ; Molecular Sequence Data ; Pain ; etiology ; Pain Management ; RNA Interference ; RNA, Small Interfering ; genetics ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Repressor Proteins ; biosynthesis ; genetics ; Sciatic Nerve ; injuries ; Transfection

Objective:To investigate the value of application of combined ultrasound and spinal CT/MRI-assisted lumbar puncture in spinal muscular atrophy (SMA) patients with spinal deformities.Methods:Twelve SMA patients with spinal deformities were evaluated for combined ultrasound and spinal CT/MRI imaging-assisted lumbar intrathecal administration of nusinersen between November 2021 and September 2022.The degree of difficulty of lumbar puncture in SMA patients was graded according to the spinal imaging, and then different puncture methods were designed according to different puncture difficulty grades.The success rate, preparation time, operation time, the number of attempts, and complications of lumbar puncture were counted.Results:In this study, 48 lumbar punctures were performed in 12 patients with SMA, with a success rate of 100%.The average preparation time for puncture was 13.60 min, the average puncture time was 4.96 min, and the average number of attempts was 1.33. No complications such as injury to the spinal cord, nerve and viscera were found in all patients.Conclusions:Combined ultrasound and spinal CT/MRI-assisted lumbar puncture has high value when applied for SMA patients with spinal deformities.

To investigate whether mammalian target of rapamycin (mTOR) signaling pathway is involved in peripheral nerve injury-induced hyperalgesia through activation of spinal dorsal astrocytes in rats.
 Methods: A total of 30 male Sprague-Dawley (SD) rats were randomly divided into 6 groups (n=5): the 1 day group (D1 group), the 4 days group (D4 group), the 7 days group (D7 group), the 14 days group (D14 group), the normal group and the sham group. The sciatic nerve chronic constriction injury (CCI) model was established in the D1, D4, D7 and D14 group. The normal group received no treatment while the sham group was only exposed the sciatic nerve. Paw withdrawal mechanical threshold (PWMT) and paw withdrawal thermal latency (PWTL) were measured at the 1st, 4th, 7th, and 14th day after CCI in the different groups. Lumbar spinal cord were harvested on the 1st, 4th, 7th and 14th day in the D1, D4, D7, D14 group correspondingly, which were harvested on the 14th day in the normal group and the sham group. Distribution of mTOR in rat spinal cord was assessed by immunohistochemistry. The expressions of mTOR mRNA and protein in the spinal cord in different groups were determined by real-time PCR and Western blotting, respectively. Another 30 male intrathecal catheterized SD rats were randomly divided into 6 groups (n=5): a blank group, a CCI group, a CCI+early rapamycin (RAPA) group, a CCI+early dimethylsulfoxide (DMSO) group, a CCI+ later RAPA group, and a CCI+later DMSO group. The blank group didn't received any treatment; The CCI group was carried out the treatment of CCI model in the left hind limbs. 10 μL of 1% RAPA was given to the CCI+early RAPA group intrathecally at 4 hours after CCI for 3 days; the CCI+later RAPA group were treated with the same dose of RAPA on the 7th days after CCI for 3 days; the CCI+early DMSO group and the CCI+later DMSO group were injected with the same volume of 4% DMSO at the corresponding time as controls. The PWTL and PWMT were measured before and after intrathecal catheterization, and every other day after CCI. The lumbar spinal cords were selected and the expression of glial fibrillary acidic protein (GFAP) in spinal dorsal horn were examined by immunohistochemistry in the 14th day after CCI.
 Results: The immunohistochemistry positive particles of mTOR were widely distributed in the cytoplasm of the normal spinal neurons. Compared with the base line, the PWMT in the D14 group on the 1st, 4th, 7th and 14th day after CCI were significantly lower, and the PWTL on the 4th, 7th and 14th day after CCI were also significantly lower (P<0.05 or P<0.01). The expressions of mTOR mRNA and protein in the CCI groups (D1, D4, D7 and D14 group) were significantly increased than those in the normal group (P<0.05 or P<0.01). Compared with the CCI+early DMSO group, the PWMT and PWTL in the CCI+early RAPA group were obviously increased on 4th, 6th, 8th, 10th, 12th or 14th day after CCI (P<0.05 or P<0.01); compared with the CCI+later DMSO group, the PWMT and PWTL in the CCI+later RAPA group were also significantly increased at the 8th, 10th or 14th day after CCI (P<0.01 or P<0.05). The GFAP immunohistochemistry positive area and absorbance value in the dorsal horn of the lumbar spinal cord in the CCI rats were decreased in the CCI+early RAPA group compared with the CCI+early DMSO group (P<0.05 or P<0.01), and which were also decreased in the CCI+later RAPA group compared with the CCI+later DMSO group (P<0.05 or P<0.01).
 Conclusion: mTOR signaling pathway may be involved in hyperalgesia induced by peripheral nerve injury via spinal astrocyte activation in the dorsal horn of the spinal cord.
Animals ; Hyperalgesia ; Male ; Neuralgia ; Peripheral Nerve Injuries ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Spinal Cord ; TOR Serine-Threonine Kinases