Background Stromal-derived factor 1α /chemokine receptor 4(SDF-1α/CXCR4) axis is one of the important signals which mediates several different activities such as chemotaxis,adhesion,proliferation and survival resulting in recruitment to sites of immune and inflammatory reactions.Considerable evidence suggests that CXCR4/SDF-1α axis is involved in tumor angiogenesis and plays a key role in the development of ocular neovascularization.Objective The purpose of this study was to explore the effect of CXCR4 antagonist on the development of cxperimental corneal neovascularization(CNV).Methods CNV model was established in the left eye of 8-weekold clean BALB/c mouse by putting the filter with 1 mol/L NaOH at the central cornea for 40 seconds.The animals were randomizcd into hyaluronate group and CXCR4 antagonist group,and the edydrops was topically administered respectively on the day of modeling 4 times per day for 14 days.CNV was examined under the slit lamp at the fourteenth day,and then the corneal suspension and section were made.Expressions of CXCR4 mRNA and protein in corneas were detected using RT-PCR and Western blot.The CD31 level in cornea was assayed by flowcytometry and immunochemistry.The expression of VEGF in burned corneas and suspension from mouse peritoneal macrophages stimulated with CXCR4 antagonist in vitro was detected by ELISA.The use of the animal followed Ragulations for the Administration of Affairs Concerning Experimental Animals by State Science and Technology Commission.Results Two weeks after corneal alkali burn,the growth of CNV peaked under the slit lamp.Compared with hyaluronate group,CNV was obviously decreased in the CXCR4 antagonist group.Immunochemistry showed that intensity of positive staining for CD31 in cornea in the CXCR4 antagonist group was weaker than the hyaluronate group.Flowcytometry clarified that CD31 positive cells rate was 9.50％ ±2.34％ in the CXCR4 antagonist group and 17.50％ ±3.16％ in the hyaluronate group,showing a significant difference between them (t=-7.312,P＜0.05).In 2,4,7 days after cornea alkali burn,the expressions of CXCR4 mRNA and protein were significantly enhanced in burn corneas compared with normal corneas(P＜0.01 ;P＜0.05).ELISA showed that the VEGF expression level in corneal tissue and supernatant of mouse peritoneal macrophages in vitro were significantly lower in the CXCR4 antagonist group than that of hyaluronate group(t =10.927,5.151,P＜0.05).The expression level of VEGF in corneal suspension was lower in the GM-CSH+CXCR4 antogonist group than that in the GM-CSH group (P＜0.05).Conclusions CXCR4 antagonist can reduce experimental CNV by down-regulating VEGF expression in cornea.

Objective To study the molecular mechanism of interleukin 25 (IL-25)expression in the lung of asthmatic rats.Methods The expressions of IL-25 mRNA and protein in the lungs were detected by Real-time PCR and ELISA,respectively.The levels of IL-25 mRNA and protein were detected by ovalbumin (OVA)in human bronchial epithelial cells.And the transcription factors that regulate IL-2 5 expression were explored through site prediction.Results The expressions of IL-25 mRNA and protein in the lung of OVA-induced asthma rats were significantly increased during animal experiments.Cell experiments showed that OVA could increase the expression of IL-2 5 in human bronchial epithelial cells in a dose-dependent manner,and OVA could upregulate the expression of transcription factor AP1.AP1 was found in the promoter region of IL-25 by site prediction.The AP1 inhibitor (T5224)significantly reduced the expression of IL-25 in OVA-induced human bronchial epithelial cells. Conclusion The molecular mechanism of IL-25 expression induced by OVA in asthma is related to the increase of transcription factor AP1 .

Aim To establish a co-incubation system in cardiac fibroblasts of SD neonatal rats and spleen CD4+ CD25 + regulatory T lymphocytes (Tregs) of normal adult SD rats,and to investigate the effects of eplerenone(EPL) on the interaction of two cells and the relationship with the Kvl.3 channel on Tregs cell membrane.Methods The spleen Tregs of normal adult SD rats were sorted by immunomagnetic bead sorting,and the myocardial fibroblasts of SD rats were isolated by differential adherence method.The experiment was conducted in the following groups:CFs,CFs + Tregs,CFs + Tregs + EPL,Tregs.The proliferation of CFs was detected by CCK-8 method.The expression levels of type Ⅰ collagen,type m collagen and matrix metalloproteinase 2 (MMP-2) secreted by CFs were detected by ELISA.The mRNA expression levels of Kv1.3,KCa3.1 on Tregs cell membrane and intracellular CRAC channel were detected by RT-qPCR technique.Tregs cell membrane Kvl.3 channel protein expression levels were determined by In-Cell Western blot.Results After 48 h incubation of the co-culture system,the cell proliferation was stable.CFs proliferation was marked(P ＜0.01),which could be inhibited by EPL(P ＜0.01).The type Ⅰ,type Ⅲ collagen and MMP-2 secreted by CFs increased (P ＜ 0.01).The expression levels of Kv1.3,KCa3.1 and CRAC channel mRNA in Tregs increased by 6.95,1.99 and 1.53 fold (CFs + Tregs vs Tregs,P ＜0.01),EPL decreased the mRNA level of each channel (CFs +Tregs + EPLvs CFs + Tregs,P＜0.01),and the decrease of Kv1.3 channel was the most significant (P ＜ 0.01).The Kv1.3 channel protein of Tregs increased by 67.9％ (CFs + Tregs vs Tregs,P ＜0.01),which could be inhibited by EPL(P ＜ 0.01).Conclusions Tregs cultured with CFs after 48 h can significantly promote the proliferation of CFs,and EPL can down-regulate the Kv1.3 channel expression on the Tregs membrane and inhibit the activation/proliferation of Tregs,indirectly inhibiting myocardial fibrosis.

The study was aimed to establish a new method of preparation of human placenta factor (PF) and to determine its physic-chemical properties, as well as effects on lymphocytes in vitro. PF was prepared by ultrafiltration. The contents and molecular weight of all constitutions were determined by Bradford method and SDS-PAGE, respectively. Cyclosporin A (CsA) was served as positive control, normal saline (NS) was used as negative control. PHA-stimulated lymphocyte proliferation and mixed lymphocyte reaction (MLR) were detected with MTT assay. The expression of CD69 on T cells was analyzed by flow cytometry. Cytotoxicity of natural killer (NK) cells against K562 tumor cells was examined with LDH release assay. The results indicated that PF was determined to be a group of low molecular weight polypeptides, consisting of two major components whose molecular weight were 9.187 and 4.794 kD respectively. The contents of PF were 5.7 - 6.9 mg/g fresh placenta. PF had similar suppressive effects on PHA-stimulated lymphocyte proliferation and MLR in vitro as compared with CsA (P > 0.05). Both PF and CsA could downregulate the expression of CD69 on T cells which had been stimulated by PMA plus ionomycin (PF vs CsA, P > 0.05). The cytotoxicity of NK cells against K562 cells in PF group was slightly higher or equivalent as compared with that in NS group (P > 0.05), but the cytotoxicity in CsA group was much lower than that in NS group (P < 0.05). It is concluded that a new method of preparation of PF has been established. This study first demonstrates that PF has strong immunosuppressive effects on T cell in vitro, and suppresses T cell proliferation and activation induced by mitogen and alloantigen. This study indicats that PF has no any inhibitory effects, but even enhances the cytotoxicity of NK cells against K562 tumor cells. These results suggest that PF may have suppressive effects on graft-versus-host disease (GVHD) without diminishing graft-versus-tumor (GVT) effects. Therefore, PF may probably be an ideal and promising agent against GVHD.
Cyclosporine ; pharmacology ; Ferritins ; immunology ; isolation & purification ; Graft vs Host Disease ; metabolism ; prevention & control ; Humans ; Immunosuppressive Agents ; immunology ; K562 Cells ; Killer Cells, Natural ; immunology ; Lymphocytes ; immunology ; T-Lymphocytes ; immunology

Modern drying technology was used to explore suitable drying process to provide scientific basis for improving drying processing methods of Scrophulariae Radix. Controlled temperature and humidity drying, vacuum drying apparatus, microwave vacuum drying apparatus, short infrared drying device were used to gain samples for analyzing. The character appearance, concentration of main components and power consumption indicators were chosen for preliminary judging. Six major components, including iridoids and phenylpropanoids were analyzed by UPLC-MS/MS method. The contents of polysaccharides were determined by UV-visible spectrophotometry. The character appearance with controlled temperature and humidity drying and short infrared drying meet the pharmacopoeia standard (Ch. p, edition 2015), while samples with vacuum and microwave vacuum drying apparatus didn't. Compared to fresh sample, concentrations of harpagide, harpagoside, aucubin and catalpol were lower in the dried samples. Angoroside-C showed no significant change before and after drying. Concentration of acteoside increased after drying. Samples with controlled temperature (70 degrees C) and humidity (15% - 10%) drying had high content and short drying time. The better drying process of Scrophulariae Radix was controlled temperature and humidity drying. The method will provide the reference for the drying technology standard of roots medicine.
Desiccation ; methods ; Drugs, Chinese Herbal ; chemistry ; Plant Roots ; chemistry ; Quality Control ; Scrophularia ; chemistry

OBJECTIVETo explore the changes in the threshold of auditory brainstem response (ABR) and [Ca(2+)]I and calmodulin (CaM) in cochlear nucleus of newborn mice infected by murine cytomegalovirus (MCMV) in the brain.

METHODSSixty-nine newborn mice were randomized into model group and control group. The model group (54 mice) was established by intracranial injection with MCMV viral suspension 20 l and the same volume of 0.9% sodium chloride was injected in the control group (15 mice). After 1 month, the ABR was tested in a sound-electric screen environment and the threshold was recorded. Then intracellular free calcium [Ca(2+)]i and the mRNA level of CaM in the cochlear nucleus were assayed by flow cytometry and RT-PCR.

RESULTSCompare to the control group [(64.0 ± 1.3) dBSPL], the threshold of ABR in the model group [(84.5 ± 2.7) dBSPL] was increased (F = 2.789,P = 0.000). Moreover, in the model group the intracellular free calcium [Ca(2+)]i and the mRNA level of CaM in the cochlear nucleus were increased (F = 1.290, P = 0.000; F = 4.252, P = 0.023), and the differences were statistically significant.

CONCLUSIONSThe intracranial injection of MCMV can lead to abnormal changes in the threshold of ABR in mice, and the change of [Ca(2+) ]I/CaM in cochlear nucleus may be the important pathological basis of sensorineural hearing loss induced by MCMV infection.

3T3 Cells ; Animals ; Auditory Threshold ; Calcium ; metabolism ; Calmodulin ; metabolism ; Central Nervous System Viral Diseases ; metabolism ; virology ; Cochlear Nucleus ; metabolism ; Cytomegalovirus ; Cytomegalovirus Infections ; metabolism ; Evoked Potentials, Auditory, Brain Stem ; Female ; Male ; Mice ; Mice, Inbred BALB C

OBJECTIVETo study pharmacological effects of PEC (the oral liquid which consists of Panax quinquefolium, Epimedium brevicornum, Schisandra chinensis Bail and Cervus eplaphus) on mice.

METHODExperiments were carried out through swimming test, step-through, spontaneous activity and sleeping time.

RESULTWhen 5-10 mL x kg(-1) of PEC was given orally for 7 days, it could prolong swimming duration of mice in water tank, and increase the tolerant ability against oxygen-deficiency. PEC could also improve cognitive-deficiency induced by taking off sleep with force in mice after given orally for 7 days. The PEC could increase the spontaneous activity in mice, antagonize the inhabitation induced by Valium, and shorten the sleeping time caused by sodium pentobarbital.

CONCLUSIONPEC has strong potential neuro-pharmacological activities such as anti-fatigue, improving cognitive-deficiency in mice.

Animals ; Behavior, Animal ; drug effects ; Drug Combinations ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Epimedium ; chemistry ; Fatigue ; drug therapy ; Hypoxia ; drug therapy ; Male ; Memory Disorders ; drug therapy ; Mice ; Panax ; chemistry ; Pentobarbital ; pharmacology ; Phytotherapy ; Plants, Medicinal ; chemistry ; Schisandra ; chemistry ; Sleep ; drug effects ; Swimming

OBJECTIVETo screen out related microRNAs in keloid tissue, and identify their effect on the proliferation of keloid fibroblasts.

METHODS8 cases of keloid tissue and 8 cases of normal skin tissue were collected as specimens. The differently expressed miRNA in keloid tissue from normal skin tissue were screened out with gene chip( Exiqon company), which was validated with quantitative real-time PCR. Then miRNA mimics was transfected into keloid fibroblasts line to stimulate high expression of mature miRNA in cells. The effect on the proliferation of fibroblasts in keloid was tested by Edu.

RESULTS(1) A total of 17 differently expressed microRNAs were found, including miR-199a-5p. (2) The expression of miR-199a-5p had been verified by qRT-PCR to be down-regulated in keloid, which was consistent with the result of array. (3) The positive rate of EdU in miR-199a-5p mimics transfected group and negative control group was (20.72 +/- 2.50)% and (27.68 +/- 4.92)%, respectively. The proliferative rate of keloid fibroblasts turned down in miR-199a-5p-transfected group (t = 2.183, P = 0.047). Besides that, the cell cycle changed after transfection. The percentage of S and G2/M phase in miR-199a-5p mimics transfected group was 33.93 +/- 1.30 and 10.87 +/- 0.80, respectively, while it was 31.39 +/- 0.79 and 9.27 +/- 0.46 in negative control group, and the difference was statistically significant.

CONCLUSIONS(1) The miRNA expression profile is different between keloid and normal skin; (2) The expression of miR-199a-5p is down-regulated in keloid and miR-199a-5p can affect the cell cycle and suppress proliferation of keloid fibroblasts. It indicateds that miR-199a-5p may be involved in regulating fibroblastic proliferation.

Cell Proliferation ; Cells, Cultured ; Down-Regulation ; Female ; Fibroblasts ; metabolism ; Gene Expression Profiling ; Humans ; Keloid ; genetics ; metabolism ; pathology ; Male ; MicroRNAs ; genetics ; metabolism

AIMTo detect the deletion distribution of dystrophin gene and dystrophin changes in muscle cells of the patients with Duchenne/Becker muscular dystrophy (DMD/BMD), furthermore to investigate the relationship between them and clinical symptoms.

METHODS42 patients with DMD/BMD were screened by 9 primers multiplex PCR. The patients from 5 DMD and 2 BMD were detected by immunofluorescence technique for analyzing dystrophin located in muscle cell membrane, compared with 2 normal males.

RESULTSThe deletion of one or more exons was found in 21 patients. 16 cases (76.2%) were detected in the central region and 5 patients (23.8%) in the 5' extreme region, especially in exon 48 (6 patients). Negative result of staining was seen in 5 DMD patients. Of these, one case of DMD had no detectable levels of dystrophin, but no deletion of DMD gene. Dystrophin immunostaining from two BMD patients consisted of a discontinuous staining pattern around most fibers.

CONCLUSIONIt might be possible that some correlation existed between the type of gene deletion and the degree of severity of the disease. The amount and size of exon deletion may not affect the symptoms. DMD/BMD are highly heterogeneous in clinical manifestation and in inheritance pattern. The pathologic foundation of DMD and BMD is the absence or abnormal expression of dystrophin. The consequence of that depends not only on the degree, but also on the function.

Adolescent ; Adult ; Child ; Child, Preschool ; Dystrophin ; analysis ; Exons ; Gene Deletion ; Humans ; Male ; Multiplex Polymerase Chain Reaction ; Muscular Dystrophy, Duchenne ; genetics ; Sequence Deletion ; Young Adult

OBJECTIVETo report a case of pure erythroid leukemia.

METHODSThe clinical features, treatment and prognosis of a rare case of pure erythroid leukemia were reported, and the related literature was reviewed.

RESULTSThe pure erythroid leukemia patient was diagnosed by 90.4% pronormoblasts in bone marrow, 99.5% for erythroid antigen CD71, 67.4% for glycophorin A were detected, while no differentiation antigen of myeloid, lymphoid and megakaryocyte lineages were observed. HAG (homoharringtonine + Cytarabine and G-CSF) regimen were administered with no effect. The patient developed multiple organ failure and died soon.

CONCLUSIONPure erythroid leukemia has a fulminant clinical course with poor response to chemotherapy and worse prognosis.

Humans ; Leukemia, Erythroblastic, Acute ; therapy ; Male ; Middle Aged ; Prognosis