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OBJECTIVES@#Systemic lupus erythematosus (SLE) is a kind of autoimmune inflammatory connective tissue disease which seriously endangers human health. Genetic factors play a key role in the pathogenesis of SLE. This study aims to investigate a novel phospholipase D2 (PLD2) mutation associated with familial SLE, and further explore the underlying mechanism of the mutation in SLE.@*METHODS@#The blood samples from a SLE patient, the patient's parents, and 147 normal controls were collected and DNA was extracted. Whole genome high-throughput sequencing was performed in the patient and her parents and the results were further analyzed by various bioinformatics methods. The wild type (wt), mutant type (mu), and negative control PLD2 plasmids were further constructed and transfected into 293 cells. The expression level of HRAS protein in 293 cells was detected by Western blotting.@*RESULTS@#In this SLE family, the female SLE patient and her mother, 1 in generation II and 1 in generation III had typical clinical manifestations of SLE, and all of them had lupus nephritis at early stage. The genetic characteristics are consistent with autosomal dominant inheritance. A novel PLD2 heterozygous mutation (c.2722C>T) was found in the patient and her mother, but not in her father and other normal controls. Compared with wtPLD2 plasmid and negative control PLD2 plasmid, the expression of HRAS in 293 cells transfected with muPLD2 plasmid was significantly up-regulated (both @*CONCLUSIONS@#PLD2 c.2722C>T mutation may be one of the pathogeny of SLE in this family.
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Feminino , Humanos , Estudos de Casos e Controles , Sequenciamento de Nucleotídeos em Larga Escala , Lúpus Eritematoso Sistêmico/genética , Nefrite Lúpica , Mutação , Fosfolipase DRESUMO
Objective To study the effect and mechanism of dexmedetomidine combined with sufentanil on postoperative analgesia in elderly patients with abdominal surgery. Methods Ninety-six elderly patients having undergone abdominal surgery in the First Affiliated Hospital of Wenzhou Medical University from January 2016 to June 2017 were enrolled, and they were divided into a control group and an observation group by random number table method, 48 cases in each group. General anesthesia was performed in the operation, and after surgery venous analgesic pump was used for analgesia in both groups. Analgesic method: the control group was given sufentanil 2 μg/kg and tropisetron 5 mg; the observation group was given dexmedetomidine 2 μg/kg, sufentanil 2 μg/kg and tropisetron 5 mg. The changes of pain and sedation score, conventional indexes [systolic blood pressure (SBP), diastolic blood pressure (DBP), heart rate (HR), respiratory rate (RR), pulse blood oxygen saturation (SpO2)], oxidative damage indexes [lipid peroxide (LPO), glutathione peroxidase (GSH-Px), Cu-Zn superoxide dismutase (Cu-Zn SOD)], stress response indexes [cortisol (Cor), epinephrine, norepinephrine (NE)], platelet activation indexes granule membrane protein-140 (GMP-140), thromboxane A2(TXA2) and the incidence of adverse reactions were observed in both groups. Results ① After surgery the visual simulation score (VAS) and Ramsay score in both groups were higher than those before surgery, and showed a tendency firstly increased and then decreased, and reached to peak value 2 hours after operation [VAS score:the control group was 3.24±0.98 vs. 1.95±0.93, observation group was 3.19±1.03 vs. 1.98±0.95; Ramsay score:the control group was 3.26±0.51 vs. 1.90±0.45, observation group was 3.77±0.53 vs. 1.92±0.42], began to decline 6 hours after operation, reached to valley value 48 hours after operation, and there was no significant difference in VAS scores between the two groups (2.02±0.64 vs. 1.98±0.95), Ramsay score was significantly higher in observation group than that in control group (2.59±0.41 vs. 2.10±0.21). ② Since 2 hours after the operation, the SBP, DBP, HR and RR in the observation group began to be lower than those in control group, 12 hours after surgery their values reached their valleys [SBP (mmHg, 1 mm Hg = 0.133 kPa): 113.2±13.5 vs. 122.1±10.3, DBP (mmHg): 67.5±9.9 vs. 76.4±8.6, HR (bpm): 64.5±6.9 vs. 71.4±7.5, RR (bpm): 14.8±1.1 vs. 15.8±0.8, all P < 0.05]; while SpO2did not change a great deal. ③ LPO, Cor, epinephrine, NE, GMP-140, TXA2of two groups after operation were higher than those before operation, GSH-Px and Cu-Zn SOD were lower than those before operation. However, LPO, Cor, epinephrine, NE, GMP-140, TXA2in observation group were significantly lower than those in control group [LPO (μmol/L): 6.4±0.8 vs. 9.8±1.1, Cor (ng/L): 148.2±19.1 vs. 239.3±27.8, epinephrine (μg/L): 124.2±13.9 vs. 207.1±23.5, NE (μg/L): 109.2±14.8 vs. 183.3±21.6, GMP-140 (μg/L): 27.13±3.82 vs. 39.06±4.83, TXA2(ng/L): 422.30±53.74 vs. 610.43±73.21, all P < 0.05], GSH-Px and Cu-Zn SOD in observation group were significantly higher than those in the control group [GSH-Px (U/L): 426.7±58.7 vs. 307.9±51.2, Cu-ZnSOD (μg/L): 311.3±42.5 vs. 231.6±34.1, all P < 0.05].④ The incidence of adverse reaction nausea in the observation group was significantly lower than that in the control group [4.17% (2/48) vs. 37.5% (18/48)]. Conclusion Dexmedetomidine combined with sufentanil used in elderly patients after abdominal surgery has significant analgesic effect, can effectively inhibit platelet activation, and decrease the contents of GMP-140 and TXA2.
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Objective To analyze the effect of GC-rich DNA fragments on the level of transgenic expression in Chinese hamster ovary (CHO) celts and its position effect.Methods The synthetic DNA fragment with GC-rich was cloned into the 5'or 3'or both 5'and 3'ends of expression cassette of expression vector.Three new expression vectors (pIRES-G1,pIRES-G2 and pIRES-G3) which was inserted with the GC-rich DNA fragments in different position were transfected CHO ceils,respectively,and then was observed under fluorescence microscope;the control vector was pIRES-EGFP.Stable transfected cell lines were screened under G418,and enhanced green fluorescent protein(EGFP) expression was analyzed by flow cytometry and the transgenic copy number was detected by quantitative real-time quantitative polymerase chain reaction (qRT-PCR).Results Three expression vectors with a GC-rich DNA fragments in different position were constructed successfully.The insertion of GC-rich DNA fragments at 3'end and both 5',3'ends of the box of expression vector could obviously improve the expression level of vector in CHO cells;and the expression level of the stably transfected CHO cells increased 1.39 fold and 1.32 fold compared to the control vector,respectively;the transgene copy number increased 1.32 fold and 1.24 fold compared with the control vector.While the insertion of GC-rich DNA fragments at 5'end of expression cassette had no obvious effect on the level of gene expression.Conclusion The role of DNA fragment with GC-rich in improving the transgenic expression of CHO cells is related to its position in the vector.The insertion of GC-rich DNA fragments at 3'end and both 5',3'ends of the box of expression vector can improve transgenic expression.
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Objective:To explore the effects of the natural plant ingredients lemon essential oil(LEO),limonene(LIM)and tea poly-phenols(TP)on the cell surface hydrophobicity and adherence of Streptococcus mutans(S.mutans).Methods:S.mutans were treated by sub-minimal inhibitory concentration(MIC)levels of LEO,LIMand TP respectively.Adsorption to hexadecane was used to measure the hydrophobic interaction of S.mutans.A classical 96-cell microtitre plate production assay using crystal violet staining was employed to visualize the adherence of S.mutans to hard tissue surface.Results:LEO,LIMand TP at sub-MIC levels could inhibit the cell sur-face hydrophobicity and adherence of S.mutans in a dose-dependent manner(P <0.05).At 1 /2 MIC and 1 /20 MIC,the inhibitary effect of LEO was stronger than that of LIMand TP(P <0.05).Conclusion:LEO may possess anticariogenic potential.
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Objective:To explore the antioxidant activity of natural plant ingredients lemon essential oil(LEO),limonene(LIM) and tea polyphenol(TP).Methods:The UV-Vis spectrometry was used to determine the 3 agents on the scavenging activity of 1 ,1 diphenyl-2-trinitrobenzene hydrazine free radicals(DPPH·)and hydroxyl free radical (·OH).Results:LEO,LIM and TP showed scavenging effect on the 2 kinds of free radicals.The (DPPH·)scavenging effect:LEO(IC50 =0.01 4)>TP(IC50 =0.01 4)>LIM (IC50 =0.002);the (·OH)scavenging effect:TP(IC50 =0.079)>LEO(IC50 =0.01 3)>LIM(IC50 =0.004).In a certain con-centration range,scavenging effect was increased with the concentration increase.Conclusion:As a nontoxic natural extract,lemon essential oil has strong antioxidant activity.
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Objective To investigate the effect of Shenmai injection on purine content in rat cerebral cortex in order to provide a theoretical basis concerning its brain protective mechanism. Methods Sixteen Sprague-Dawley (SD) rats were randomly divided into two groups:normal saline control group and Shenmai injection group, with 8 rats in each group. Shenmai injection 15 mL/kg was injected intraperitoneally into the rats in Shenmai injection group, while in the normal saline group, an equal volume of normal saline was intraperitoneally injected. After the injection for 24 hours, the rats were sacrificed, and the cerebral cortex was removed on ice, homogenized and its supernatant was extracted;then high performance liquid chromatography (HPLC) was used to detect adenosine triphosphate (ATP), adenosine diphosphate (ADP), adenosine monophosphate (AMP), adenosine and inosine contents in the supernatant of cerebral cortex. Results Compared with normal saline control group, ATP, ADP, AMP, adenosine and creatinine content in the cerebral cortex of Shenmai injection group were significantly higher, the differences being statistically significant [ATP (ng/L): 31.62±5.12 vs. 20.25±4.53, ADP (ng/L): 37.04±6.72 vs. 25.12±7.35, AMP (ng/L): 87.82±20.37 vs. 33.23±10.34, adenosine (ng/L): 2.82±0.15 vs. 1.12±0.61, creatinine (ng/L): 11.72±1.05 vs. 6.05±2.55, P < 0.05 or P<0.01]. Conclusion Shenmai injection can elevate ATP, ADP, AMP, adenosine and creatinine contents in the cerebral cortex of rats, possibly that is the theoretical basis for brain protective mechanism of Shenmai injection.
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<p><b>OBJECTIVE</b>To investigate the effect of the serum of rats fed with Shenkang pill in regulating monocyte chemoattractant protein 1 (MCP-1) expression induced by advanced oxidation protein products (AOPP) in mouse podocyte clone 5 (MPC5) and explore the underlying mechanism.</p><p><b>METHODS</b>MPC5 cultured in vitro were incubated for different time lengths in the presence of different concentrations of serum of rats medicated with Shenkang pill, and the cell proliferation was assessed using MTT assay. In MPC5 treated with AOPP prior to exposure to the rat serum, the changes in the protein expressions of p38MAPK and IκBα were examined with Western blotting, NF-κB p65 nuclear translocation was analyzed with immunofluorescence assay, and MCP-1 expression in the supernatant was determined using ELISA kits.</p><p><b>RESULTS</b>The medicated rat serum time- and concentration-dependently promoted the proliferation of MPC5, with the optimal serum concentration of 5% and incubation time of 24 h. AOPP significantly increased MCP-1 expression in the cell supernatant in a time-and concentration-dependent manner; pretreatment with SB203580 (a p38 inhibitor) or parthenolide (a NF-κB inhibitor) significantly decreased MCP-1 expression, and treatment with the medicated serum significantly decreased AOPP-induced MCP-1 expression. AOPP concentration-dependently increased the protein expression of P-p38 but decreased that of IκBα. Both the medicated serum and SB203580 increased IκBα protein in AOPP-induced cells, but the effect was more obvious with the medicated serum. The medicated serum also decreased NF-κB p65 nuclear translocation in AOPP-induced MPC5.</p><p><b>CONCLUSION</b>Shenkang pill-medicated serum can decrease AOPP-induced expression of MPC-1 in MPC5 by regulating p38MAPK/NF-κB to mediate its anti-inflammatory effect. This finding provides a new theoretical basis for the application of Shenkang pill to treat diabetic nephropathy.</p>
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Animais , Camundongos , Ratos , Produtos da Oxidação Avançada de Proteínas , Farmacologia , Linhagem Celular , Proliferação de Células , Quimiocina CCL2 , Metabolismo , Regulação para Baixo , Medicamentos de Ervas Chinesas , Farmacologia , Proteínas I-kappa B , Metabolismo , Imidazóis , Inibidor de NF-kappaB alfa , Piridinas , Soro , Química , Sesquiterpenos , Fator de Transcrição RelA , Metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno , MetabolismoRESUMO
<p><b>OBJECTIVE</b>To explore the effects of parthenolide (PTL) on high glucose-stimulated cell proliferation, NF-κB activation and monocyte chemoattractant protein-1 (MCP-1) expression in rat mesangial cells (MCs).</p><p><b>METHODS</b>MCs were cultured in normal glucose (5.6 mmol/L), high glucose (30 mmol/L), or high glucose with PTL. MTT assay was used to detect the cell proliferation. MCP-1 content in the supernatant was measured by ELISA, and the level of IκBα was detected by Western blotting to reflect NF-κB activity. EMSA method was used to measure the activation of NF-κB.</p><p><b>RESULTS</b>MC proliferation, MCP-1 expression and NF-κB activation were significantly enhanced and the expression of NF-κB-binding protein IκBα was obviously reduced in cells cultured in high glucose. Application of PTL obviously abolished the effects of high glucose.</p><p><b>CONCLUSION</b>PTL can suppress high glucose-stimulated NF-κB activation and MCP-1 expression in rat MC. These data provide a theoretical basis for the clinical application of PTL in prevention and control of diabetic nephropathy.</p>
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Animais , Ratos , Anti-Inflamatórios não Esteroides , Farmacologia , Asteraceae , Química , Proliferação de Células , Células Cultivadas , Quimiocina CCL2 , Metabolismo , Relação Dose-Resposta a Droga , Glucose , Farmacologia , Proteínas I-kappa B , Metabolismo , Células Mesangiais , Biologia Celular , Metabolismo , Inibidor de NF-kappaB alfa , NF-kappa B , Metabolismo , Plantas Medicinais , Química , Sesquiterpenos , FarmacologiaRESUMO
In this research project, rats were made into animal models of acute focal cerebral ischemia and reperfusion (IR) by occlusion of their middle cerebral artery (MCAO). We observed the effect of endogenous endothelial progenitor cells (EPCs) and serum cytokines on cerebral ischemia rats treated by electro-acupuncture(EA). The results showed: MCAO model had high stability after EA treatment which was delivered via the acupuncture needles inserted into "quchi" and "zusanli" points, the nervous functions of cerebral IR rats recovered faster than those of rats not treated; EPCs in rats' blood increased after acute focal cerebral ischemia and reperfusion; and the growth rate was obvious in IR group. This phenomenon might be related to the inflammation elicited by injury of ischemia and self-repair. Besides, EA treatment could decrease induced nitric oxide synthase (iNOS) activity, alleviate injury after cerebral ischemia, and regulate the quantity of EPCs in blood. The quantity of EPCs in blood increased in IR-24hr. In IR-48 hr, the rise of EPCs quantity was significant (P < 0.01). The level of vascular endothelium growth factor (VEGF) in serum of rats after cerebral ischemia was escalated, which indicated to a certain extent that cerebral ischemia could stimulate stress reaction. EA treatment could raise VEGF level, which suggested that high expression of VEGF could accelerate mobilization, chemotaxis and homing of EPCs. At the same time, the levels of matrix metalloproteinase-9 (MMP-9) and basic fibroblast growth factor (bFGF) also changed. In conclusion, EA treatment could promote neovascularization after cerebral ischemia by mobilizing EPCs, decreasing iNOS activity and increasing VEGF level. This may be one of the ways by which EA could treat cerebral ischemia.