Background and purpose:Researches had indicated that about over 85%of malignant tumors highly express telomerase activity. So telomerase has become one of the important methods in the research field of tumor diagnosis and treatment. Nowadays, several reports about malignant tumor which over expresses hTERT targeting imaging with radionuclide labeled hTERT ASON had been published. In these reports, high quality of pictures can hardly be acquired because of poor anatomical and spacial resolution in nuclear imaging itself. Accordingly, in this study, we developed a method of detecting human telomerase in vivo with magnetic resonance imaging (MRI) and evaluate its feasibility. Methods:Firstly, Uniformly phosphorothioate-modified human telomerase reverse transcriptase antisense oligonucleotide (hTERT ASON) was labeled with Gd3+ through the bifunctional chelator 1, 4, 7, 10-tetraazacyclododecane-N, N’, N’’, N’’’-tetraacetic acid (DOTA) and iv vitro experiments were performed to characterize the antisense probes (for biodistribution and cellular uptake, 99mTc-DOTA-ASON was used in stead of Gd-DOTA-ASON). Then Gd-DOTA-ASON was injected intraperitoneally in pulmonary adenocarcinoma A375 nude mice tumor-bearing BALB/c for in vivo imaging using 7.0 T Micro MRI periodically, tumors and their surrounding tissues were defined as region of interest (ROI) to calculate the signal to noise ratio (SNR) of tumor to muscle using Gd-DTPA as control. Finally, immunohistochemical analysis of telomerase activity of each xenograft was operated 2 days after imaging. Results:The binding efficiency of Gd-DOTA-ASON reached was as high as 65%(63.2±2.4, n=6). And it can maintain 61%in fresh human serum and normal saline at 37℃over 24 h;A375 cells showed an uptake of 8.5%when incubated with 99mTc-DOTA-ASON;In comparing with DOTA-ASON and Gd-DTPA, cells transected with Gd-DOTA-ASON had higher SI when performed MRI with T1WI. The hTERT-expressing xenografts were obviously enhanced by Gd-DOTA-ASON at 0.5-6 h after injection and the SNR can reach 2.37, whereas obvious enhancement only could be found within 2 h after injection of Gd-DTPA. Both labeled and non-labeled antisense probes can suppress the activity of telomerase of A375 cells either in vitro or in vitro. Conclusion:Our research offers proof that Gd-DOTA-ASON can be used as tumor specific targeting MR probe for diagnosing malignant tumors with high expression of telomerase.

Objective To investigate the phenotypes and the HIV-1-specific T cell responses of KIR3DL1 positive CD8 cells in patients with early HIV-1 infection. Methods Fifty-six HIV-1 antibody negative individuals and thirty-two patients with early HIV-1 infection were enrolled in the study. Fluores-cence-activated cell sorting (FACS) was performed to detect the phenotypes of KIR3DL1 receptor expressed on the surface of CD8 cells. The levels of IFN-γwere measured by intracellular cytokine staining assay after the PBMCs were stimulated with an HIV-1 Gag peptide pool. Results The percentages of KIR3DL1+CD8 T cells in HIV-1 negative individuals and patients with early HIV-1 infection were 1. 45% (0. 12%-8. 4%) and 0. 82% (0. 14%-6. 14%), respectively, and there was no significant difference between them. The percentages of KIR3DL1+CD8 Temra cells in HIV-1 negative individuals and patients with early HIV-1 infec-tion were (4. 55±3. 84)% and (6. 71±8. 50)%, respectively, which were significantly higher than the per-centages of KIR3DL1+CD8 Tem cells, which were (0. 50±0. 59)% and (1. 18±1. 39)%, respectively (all P<0. 01). Moreover, the percentages of KIR3DL1+CD8 Tem cells in patients with early HIV-1 infection were higher than those in HIV-1 negative individuals (P=0. 001 2). The percentage of KIR3DL1+CD8 Temra cells was positively correlated with the HIV-1 viral load in patients with early HIV-1 infection ( rs=0. 576,P=0. 000 9). The percentages of KIR3DL1+CD8 Temra cells in HIV-1 patients, whose viral loads were larger than 4. 0log, were much higher than those in HIV-1 patients with viral loads less than 4. 0 log (P=0. 002). Additionally, the levels of IFN-γsecreted by KIR3DL1 positive CD8 cells were much lesser than those secreted by KIR3DL1 negative CD8 cells (P<0. 000 1). Conclusion The receptor of KIR3DL1 was mainly expressed on CD8 Temra cells in both HIV-1 negative subjects and patients with early HIV-1 infec-tion. High HIV-1 viremia was associated with the high percentage of KIR3DL1+CD8 Temra cells. The KIR3DL1 positive CD8 cells induced lower HIV-1-specific T cell responses.

Objective:To study the antimalarial mechanism of benflumetol (B). Methods: Flow cytometry (FCM) was used to analyze the effects of B and chloroquine (CQ) on DNA content of Plasmodium berghei and pH value of the lysosome of malarial parasites. Results: DNA content of the plasmodia not treated with any drugs was not changed in 24 hours,while benflumetol could decrease the DNA content: the DNA content began to decrease 2 h after the drug administration and reached the minimum by 16 h, but somewhat increased at 24 h after administration. The pH in the lysosome increased 1 h and restored premedication level 4 h after benflumetol administration. Chloroquine had the same effects on DNA and lysosome pH of malarial parasites.Conclusions: The antimalarial mechanism of benflumetol is directly related to its effect to inhibit the synthesis of DNA.

Objective To identify the disease-causing gene mutation and investigate the genotypephenotype correlation in a Chinese pedigree with familial hypertrophic cardiomyopathy.Methods In this study we collected a large multigenerational Chinese family with FHCM.Total genome DNA was extracted from 67 subjects' peripheral leucocytes.The exons and boundary introns of MYH7 gene was amplified by PCR and directly sequenced by ABI PRISM 3700 DNA sequencer.Then,the mutation was examined.Results Fourteen family members had hypertrophic cardiomyopathy,including 4 deceased 2 of whom died from sudden death at young age.Analysis by echocardiography showed all the 10 living affected individuals have a maximal leftventricular-wall thickness of at least 13 mm.Three single nucleotide polymorphisms (SNP) which had been reported in NCBI SNP database,were found mutated.No mutation co-seperated with the disease was identified.Conclusion FHCM of this family was not caused by MYH7 mutation.Other genes should be screened to further identify the disease-causing gene mutation in hypertrophic cardiomyopathy.

OBJECTIVE Respiratory depression hinders the use of anaesthetics and sedative hyp-notics.To explore the mechanism of LCX001 on protection against respiratory depression,a novel AMPA receptor modulator LCX001,synthesized by our Institute of Medicinal Chemistry,is expected to relieve suppressed respiration. METHODS LCX001 was tested to alleviate respiratory depression triggered by opioid(fentanyl and TH-030418),propofol and pentobarbital in the plethysmography recording.The acetic acid writhing and hot-plate tests were conducted to evaluate analgesic effect of LCX001.Binding assay and whole-cell recording were used to analyze the property of LCX001 on positive modulation. The function of AMPA receptors were determined by location of receptors in the membrane and state of channel opening, and both processes were impressed by AMPA receptor regulatory proteins. Ac-cording to the theory,the effect of LCX001 on the expression of stargazin was measured firstly by west-ern blotting. The variation of receptor surface location was observed by live cell imaging. The regula-tion on neuronal Ca2+and cell function was investigated intensively by Ca2+imaging to clarify mecha-nism of LCX001. RESULTS LCX001 effectively rescued and prevented opioid (fentanyl and TH-030418), propofol, and pentobarbital-induced respiratory depression by strengthening respiratory fre-quency and minute ventilation in rats. The acetic acid writhing test and hot-plate test revealed potent anti-nociceptive efficacy of LCX001,in contrast to some ampakines that did not affect analgesia. Fur-thermore,LCX001 potentiated[3H]AMPA and L-glutamate binding affinity to AMPA receptors,and facili-tated glutamate-evoked inward currents in HEK293 cells stably expressing GluA2(R).Importantly,appli-cation of LCX001 generated a significant increase in GluA2(R) surface expression in a mechanism of stargazin up-regulation,and restrained opioid-induced abnormal intracellular Ca2+load,which might par-ticipate in breathing modulation. CONCLUSION The novel pharmacological effect and potential new mechanism of LCX001 might promote ampakines to be a therapeutic option for protection against respi-ratory depression.

OBJECTIVE The present study was aimed to investigate the role of Wnt/β-catenin sig-naling in spinal VGLUT2 regulation and neuropathic pain. METHODS To elucidate the association be-tween VGLUT2 and neuropathic pain,we determined the expression and distribution characteristics of VGLUT2 in mice subjected to spared nerve injury(SNI),and then observed the effects of two VGLUT2 targeting shRNAs on mechanical allodynia and glutamate release.The effects of Wnt/β-catenin signal-ing on VGLUT2 expression and pain behavior were investigated by using Wnt agonist,Wnt1,and Wnt/β-catenin pathway inhibitor XAV939 in SNI mice.RESULTS SNI surgery induced significant up-regula-tion of VGLUT2 on postoperative days 7,14,and 21.Double immunofluorescence labeling of VGLUT2 with NeuN,MAP2,Iba-1,or GFAP showed that VGLUT2 was mainly expressed in neurons in the dor-sal horn of the spinal cord after SNI(NeuN,MAP2).Intrathecal administration of VGLUT2 shRNAs be-fore or after SNI surgery significantly decreased mechanical allodynia and glutamate release. Mean-while,Wnt1/β-catenin signaling increased significantly after SNI surgery.Over-expression of β-catenin in PC12 cells increased VGLUT2 protein level,intrathecal administration of Wnt agonist or Wnt1 signifi-cantly increased VGLUT2 protein expression in spinal cord, while Wnt/β-catenin pathway inhibitor XAV939 decreased VGLUT2 expression in PC12 cells and spinal cord.Additionally,intrathecal admin-istration of XAV939 7 days after SNI significantly attenuated mechanical allodynia in mice, which was in accordance with down-regulation of VGLUT2 protein levels.VGLUT2 shRNAs significantly attenuat-ed Wnt agonist or Wnt1 induced mechanical allodynia. CONCLUSION Wnt1/β-catenin signaling path-way up-regu-lates the spinal VGLUT2 expression,and this regulation is involved in neuropathic pain behavior.

BACKGROUNDNeonatal respiratory distress syndrome (NRDS) is caused by a deficiency in pulmonary surfactant (PS) and is one of the main reasons of neonatal mortality. This study was conducted to evaluate the efficacy and safety of intra-amniotic administration of pulmonary surfactant for prophylaxis of NRDS.

METHODSForty-five pregnant women who were due for preterm delivery and whose fetuses' lungs proved immature were divided into two groups. Fifteen women (study group) were administered one dose of pulmonary surfactant injected into the amniotic cavity and delivered within several hours. Nothing was injected into the amniotic cavity of 30 women of the control group. The proportion of neonatal asphyxia, NRDS, mortality and the time in hospital were analyzed to determine if there was any difference between the two groups.

RESULTSThere was no significant difference between the two groups for neonatal asphyxia. Foam tests showed that higher proportion of neonates in the study group than in the control group (56.3% vs 13.3%, P < 0.05) had lung maturity. A greater number of control neonates (11/30, 32.3%) had NRDS, compared with the neonates given PS via the amniotic cavity before delivery (1/16, 6.3%, P < 0.05). The neonates in the study group spent nearly 10 days less in hospital than the control group [(32.4 +/- 7.6) days vs (42.0 +/- 15.7) days, P < 0.05], but the difference in mortality between the two groups was not statistically significant.

CONCLUSIONSIntra-amniotic administration of pulmonary surfactant can significantly reduce the proportion of NRDS and the time in hospital of preterm neonates. Whether this method can reduce the mortality of preterm neonates needs to be evaluated further. Intra-amniotic administration of pulmonary surfactant provides an additional effectual means for NRDS prophylaxis.

Amnion ; Female ; Humans ; Infant, Newborn ; Pulmonary Surfactants ; administration & dosage ; Respiratory Distress Syndrome, Newborn ; prevention & control ; Safety ; Treatment Outcome

Schizophrenia, described as the worst disease affecting mankind, is a severe and disabling mental disorder. Schizophrenia is characterized by complicated symptoms and still lacks a diagnostic neuropathology, so developing schizophrenia animal models which have quantifiable measures tested in a similar fashion in both humans and animals will play a key role in new therapeutic approaches. According to the symptoms of cognitive impairment and emotional disorder, the N-methyl-d-aspartate (NMDA)-receptor antagonist MK-801 was applied to induce schizophrenia-like behavior in mice. Locomotor activity and prepulse inhibition (PPI) were selected as indices and the effect of clozapine was also investigated in this model. The results showed that compared with the normal group, MK-801-treated mice exhibited significantly increased locomotor activity and impaired PPI, and pre-exposure to clozapine could ameliorate the abnormality and make it back to normal level. These findings suggest that the model we established could be a useful tool for antipsychotic drug screening.
Animals ; Antipsychotic Agents ; pharmacology ; Clozapine ; pharmacology ; Disease Models, Animal ; Dizocilpine Maleate ; Inhibition (Psychology) ; Male ; Mice ; Motor Activity ; drug effects ; Receptors, N-Methyl-D-Aspartate ; antagonists & inhibitors ; Schizophrenia ; chemically induced ; physiopathology

OBJECTIVE To explore the mechanism of Gαiand Gβγsubunits on dexmedetomidine (DMED)-induced sedation.METHODS Kunming mice were randomly placed into three groups(DMED group, DMED+dbcAMP/rolipram/gallein/M119 group, dbcAMP/rolipram/gallein/M119 group) to explore the regulation of dbcAMP/rolipram/gallein/M119 on DMED-induced sedation by establishing loss of righting reflex (LORR) model. DbcAMP/rolipram was intracerebroventricular injected and gallein/M119 was intraperitoneal injected 15 min before DMED intravenous injection. In CHO-α2A-AR cells, after administration of DMED/gallein/M119, the regulation on the cAMP accumulation stimulated by Forskolin (FSK) was detected, so was the intracellular calcium ion concentration ([Ca2+]i. The levels of pERK/pCREB were detected by Western Blot to explore the key signal molecules involved in DMED-induced sedation. RESULTS The ED50of DMED-induced LORR (200.0 nmol·kg-1) was increased to 375.0 or 433.3 nmol·kg-1by pre-treatment with cAMP analog dbcAMP(50 nmol/5μl per mouse)or phosphodies-terase 4 inhibitor rolipram(100 nmol/5μl per mouse).In addition,the ED50of DMED-induced LORR was decreased to 113.6 or 136.5 nmol·kg-1when pre-treated with Gβγsubunits inhibitor M119(100 mg·kg-1) or gallein(100 mg·kg-1)respectively.Administration of dbcAMP,rolipram,gallein or M119 alone had little effect on LORR of mice.Gallein(10 μmol·L-1)significantly inhibited forskolin-stimulated cAMP accumu-lation in CHO-α2A-AR cells.Compared with Gβγsubunits inhibitors or DMED alone,[Ca2+]iand pERK1/2 significantly increased after co-administration of Gβγsubunits inhibitors with DMED.DbcAMP(5 μmol·L-1) or rolipram (5 μmol·L- 1) alone had little effect on ERK1/2 phosphorylation, but decreased DMED-induced ERK1/2 phosphorylation after co-administration with DMED. Gβγsubunit inhibitors treatment increased DMED-induced phosphorylation of CREB, whereas dbcAMP or rolipram had little effect on pCREB induced by DMED.CONCLUSION Gβγsubunits might inhibit DMED-induced sedation through cAMP and pERK1/2 pathway,which was opposite to Gαisubuint.

Objective: To study the feasibility of evaluating the adequacy of hemodialysis using neural calculating method. Methods: The adequacy of hemodialysis patients were evaluated using Daugirdas, TACurea and neural calculating method respectively, the results of the 3 method; were compared with the clinical assessment of the patients. Results: The coincidence rate among the 3 methods was 84.6%, coincidence rate between neural calculating method and the clinical outcome of the patients was 92.3%, which was significantly higher than that of Daugirdas method (76.9%) and of TACurea (80.8%). Conclusion: Neural calculating method has higher accuracy in assessing the adequacy of hemodialysis patients and is clinically practical.