Background Herpetic stromal keratitis (HSK) is an immunopathologic eye disorder mediated by CD4+ T lymphocytes.Interleukin-9 (IL-9) is a cytokine linked to the process of many immune inflammatory diseases,but whether IL-9 is involved in the immunopathology of HSK remains unclear.Objective This study was to investigate the expression of IL-9 in murine cornea during the development of HSK and its relationship with the degree of HSK.Methods One hundred and eighty clean BALB/c mice were divided into the normal control group (20 mice) and experimental group (160 mice).HSK models were established by scratching on the surface of the cornea followed by inoculation of 1 × 106 plague forming unit(PFU) herpes simplex virus type 1 (HSV-1) KOS strain.Change of ocular surface was examined under the slit lamp biomicroscopy before and I day,3,5,7,8,10,14,21 days after inoculation of HSV-1.The corneas of mice were collected on the time points mentioned above.The relative expression level of IL-9 mRNA in the cornea of mice was detected by real-time PCR.Immunocytochemical localization of IL-9 protein in murine cornea was viewed by a confocal microscope after preparation of the corneal cryostat sections.All experimental manipulations were undertaken in accordance with the institutional guidelines for the care and use of laboratory animals.Results Punctiform,dendriform or geographic defect in corneal epithelium were seen 3 days and healing 6 days after inoculation of HSV-1.However,edema and opacity of the corneal stroma appeared 7 days and peaked 14 days following the inoculation.Real-time PCR assay showed that very little of IL-9 mRNA (A260/A280) was expressed in the cornea in the mice of the control group.But in 1 day,3,5,7,8,10 and 14 days,the relative level of IL-9 mRNA was 6.37±0.45,5.66±0.53,3.93±0.35,3.62±0.34,3.23±0.18,2.57±0.14,2.19±0.20,with a significant difference among the various time points (F=92.764,P=0.000),and IL-9 mRNA in 1 day,3,5,7,8,10 and 14 days after inoculation was significantly higher than before inoculation (P＜0.05);while no markedly difference was found in IL-9 mRNA expression between the 21-day group after inoculation and the normal control group (P =0.340).IL-9 protein was expressed in the epithelial layer,stromal layer and endothelial cell layer of the corneas,with a stronger green fluorescence in the HSK mice,but no expression of IL-9 protein was seen in the control mice.Conclusions HSK model can be successfully created by corneal scratching and inoculation of HSV-1 KOS strain in BALB/c mouse.IL-9 is involved in the pathogenesis and development of HSK mediated by immunoresponse.

OBJECTIVETo observe the neurological protective effects of progesterone (PROG) on focal cerebral ischemia/reperfusion injury in rats and to explore its possible mechanism.

METHODSOne handred and twenty male SD rats were divided into three groups randomly: sham-operated group, middle cerebral artery occlusion ( MCAO ) group and PROG + MCAO group( n = 40). The right temporary MCAO model was established by the line-embolism method. The PROG + MCAO group rats were according to 8 mg/kg intraperitoneal injection PROG, after that 30 min, the rats were suffered ischemia/reperfusion. After rats were suffered ischemia for 2 h and reperfusion 0, 24, 48, 72 h stress, the nervous functional defect degree were evaluated by longe scoring, and the expression of two-pore domain K channel 3 (TASK3) mRNA in brain tissue were detected by the real-time PCR.

RESULTSPROG (8 mg/kg) could significantly reduced the nervous functional defect degree in rats after ischemia/reperfusion 24, 48, 72 h (P < 0.05). The results of real-time PCR showed that the TASK3 mRNA expression in the brain tissue at all time points significantly decreased in MCAO group compared with sham-operated group (P < 0.05). However, compared with MCAO group, the expression of TASK3 mRNA in brain tissue at all time points dramatically increased in PROG + MCAO group (P < 0.05).

CONCLUSIONPROG can improve the nervous functional defect degree after focal cerebral ischemia/reperfusion injury in rats, and the mechanism might be associated with up-regulating the expression of TASK3 mRNA in brain tissue.

Animals ; Brain ; drug effects ; pathology ; Brain Ischemia ; drug therapy ; Infarction, Middle Cerebral Artery ; Male ; Neuroprotective Agents ; pharmacology ; Potassium Channels, Tandem Pore Domain ; metabolism ; Progesterone ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Real-Time Polymerase Chain Reaction ; Reperfusion Injury ; drug therapy

OBJECTIVETo investigate effects of the variation of immune function in high humidity environment in different time, and lay a foundation for further study of the related mechanism.

METHODThirty SD rats were divided into 3 groups (n = 10): 20 day group, 40 day group in 90% relative humidity chamber and control group in normal relative humidity. Peripheral blood and spleens were collected to detect the levels of T lymphocyte subsets by Flow Cytometery.

RESULTSIn peripheral blood of the 20 day group rats, the CD3+ %, CD4+ %, CD8+ % and CD4+/CD8+ were 52.91 +/- 6.27, 37.80 +/- 4.11, 14.85 +/- 3.73 and 2.72 +/- 0.82 separately. Expect CD3+ %, they all had significant differences (P < 0.05). In addition, the data of the 40 day group rats showed no diversity in statistics. In spleen, CD8+ % of the 20 day group rats was 6.23 +/- 2.87 with significant differences (P < 0.05) and IgG, IgA and IgM did not change a lot in blood serum of the high humidity groups except C3 of the 20 days group (P < 0.05).

CONCLUSIONIn high humidity environment, the immune function of the rats increased in the initial stage. As time went on, the immune function gradually went to normal level through the self adjustment.

Acclimatization ; Animals ; Humidity ; Rats ; Rats, Sprague-Dawley ; Spleen ; immunology ; T-Lymphocyte Subsets ; immunology

Twelve compounds were isolated from the herb of Chenopodium ambrosioides, and their structures were identified by spectroscopic methods as kaempferol-7-O-alpha-L-rhamnopyranoside (1), kaempferol-3,7-di-O-alpha-L-rhamnopyranoside (2), patuletin (3), quercetin-7-O-alpha-L-rhamnopyranoside (4), grasshopper ketone (5), 4-hydroxy-4-methyl-2-cyclohexen-1-one (6), syringaresinol (7), benzyl beta-D-glucopyranoside (8), dendranthemoside B (9), N-trans-feruloyl tyramine (10), N-trans-feruloyl 4'-O-methyldopamine (11), and 4-hydroxy-N-[2-(4-hydroxyphenyl) ethyl] benzamide (12). Among them,compounds 3, 6-8,10, and 12 were isolated from the genus Chenopodium for the first time, and compounds 2-12 were isolated from this plant for the first time.
Chenopodium ambrosioides ; chemistry ; Drugs, Chinese Herbal ; chemistry ; isolation & purification

AIM:To study the effect of roscovitine on the inflammatory hyperplasia of carotid artery intima in rats and the related mechanisms .METHODS: SD rats ( n=60 ) were randomly divided into 3 groups including control group, model group and treatment group .The rat model was established by trypsin digestion injury .The rats in control group were given sham operation .The rats in treatment group were administered with 0.5 mL roscovitine (2 g/L) slow-re-leasing gelatin.The rats in each group were fed normally for 4 weeks, then killed to take out carotid arteries for further ob-servations .The effects of roscovitine on the inflammatory hyperplasia of carotid artery intima and the related mechanism via nuclear factor-κB ( NF-κB) in the rats were detected by Western blot .RESUITS:Roscovitine inhibited the activation of NF-κB and the expression of inflammatory factors cyclooxygenase-2 (COX-2), vascular cell adhesion molecule-1 (VCAM-1),TNF-αand IL-6 via blocking the phosphorylation activation of NF-κB and inhibiting the degradation of IκB-α.CON-CLUSION:Roscovitine inhibits inflammatory hyperplasia of carotid artery intima in the rats via suppressing NF-κB activa-tion.

BACKGROUND:The limb ischemia model in rats has been widely applied in China to study the pathological process and treatment method of limb ischemia, but there are some controversies on the model construction and evaluation, so a reliable, convenient and economical disease model is needed for the related research. OBJECTIVE:To compare limb ischemia degree, duration and changing rule of acute hind limb ischemia models prepared by different ways in Sprague-Dawley rats, and find a method to prepare models that have moderate and stable limb ischemia and maintain a longer time. METHODS: 72 Sprague-Dawley rats were randomly divided into four groups, with 18 rats in each group. Group A: sham operation group, abdominal aorta below renal artery and iliolumbar artery, right superficial femoral artery, popliteal artery, saphenous artery were separated. Group B: the right superficial femoral artery, popliteal artery, saphenous artery were cut off, and the right femoral artery was resected to establish acute hind limb ischemic model. Group C: abdominal aorta and bilateral abdominal perineal artery were ligated to establish acute hind limb ischemic model. Group D: abdominal aorta, iliolumbar artery and lumbar artery were ligated to establish acute hind limb ischemic model. RESULTS AND CONCLUSION:Right hind limb muscle strengths of rats in groups B, C, D were weaker than that in group A at weeks 2, 4, 6 after operation; muscle strengths in group D was weaker than that in groups B, C at week 4 after operation; muscle strengths in groups B, D was stil weaker than that in group C at week 6 after operation. The partial venous oxygen pressure of right hind limbs in groups B, C, D was lower than that in group A at weeks 2, 4, 6 after operation; the partial venous oxygen pressure in group D was lower than that in groups B and C at weeks 2, 4 after operation; and group D was stil lower than group C at week 6 after operation. At weeks 2, 4 after operation, some muscle cels in the right hind limb muscle tissue were disrupted, fibrous connective tissue hyperplasia and capilary hyperplasia were detected, inflammatory cels infiltrated in the groups B, C, D. These pathological findings were more apparent in group D. Fiber connective tissue hyperplasia was attenuated, capilary hyperplasia, expansion and congestion were found in each group at week 6 postoperatively. The method of ligating abdominal aorta, iliolumbar artery and lumbar artery below renal artery for hind limb ischemia model can obtain moderate and stable ischemia conditions, maintain a long time, and is convenient to prepare.

Objective:Facial nerves can be dissected using anterograde and retrograde approaches. The optimal technique for the facial nerve dissection of a patient with benign parotid tumor has not yet been determined. This study focused on facial nerve dysfunc-tion and recovery rate after anterograde and retrograde facial-nerve dissections. Methods:The data of 110 patients with benign carotid adenoma from the Head and Neck Department of this hospital who were hospitalized between January 2011 and January 2013 were col-lected. These patients were divided into groups A (n=52) and B (n=58). Anterograde and retrograde dissections of the facial nerve were performed on group A and group B patients, respectively. Based on the preferential order of dissection, group B was divided into groups B1, B2, and B3 representing the zygomatic, buccal, and marginal mandibular branches, respectively. The patients were postoperatively observed to check for potential symptoms, such as facial paralysis along with its severity and recovery. The House-Brackmann grading system was used to assess all patients. Results:The operation could be successful, with better nerve exposure, using these approaches. Statistical differences were observed in the nerve injury and recovery rates between the groups, with group A better than group B, and group B2 better than the other two groups (P<0.05). Conclusion:Anterograde facial nerve dissection should be routinely used in be-nign parotid tumor, and the buccal branch of facial nerve dissection should be preferentially considered when no other option apart from retrograde dissection is available.

Objective To compare the effect of thin-sliced enhanced CT scanning and 3D-DSA data sources in the 3 D printing data reconstruction of intracranial arteriovenous malformation (AVM ). Methods Five patients with AVM were selected prospectively,3 were Spetzler-Martin grade II and 2 were grade III. Two of them used 256-slice spiral CT thin slice enhanced scanning. Three used the 3D-DSA rotating imaging,and the DICOM raw data of the examination results were extracted. Digital processing was performed by using the Mimics software,and the 3 D printing was performed according to the ratio of 1∶1 obtaining the solid model and the effects were compared. Results Using the data source 3 D printing of 256 slice spiral CT thin-slice enhanced scan could obtained skull and blood vessel image information and could reveal the smallest diameter of 0. 9 mm vessel,however,the fine branch structures of the vessel were difficult to distinguish. The 3D printing based on 3D-DSA data,although the digital subtraction did not have the skull data information,the vascular branches showed more abundant. It could reveal the smallest diameter of 0. 9 mm vessel. Conclusions Using the CT thin-slice enhanced scan or 3D-DSA data source can obtain reconstruction images of AVM nidus,and 3D-DSA shows that the better effect for spatial structure of AVM nidus. It is helpful to the design of preoperative treatment scheme and the development of corresponding auxiliary tools.

Objective To investigate the clinical significance of serum interleukin-6(IL-6),C-reactive protein(CRP) and procalcitonin(PCT) in early diagnosis of postoperative infection after artificial femoral head replacement in rabbits.Methods Selected 64 healthy New Zealand white rabbits,and randomly divided them into the experimental group and the control group,with 32 cases in each group.All of the 64 rabbits underwent left femoral hip replacement.Rabbits in the experimental group were injected with staphylococcus aureus into the left hip joint immediately after artificial femoral head replacement,while rabbits in control group were injected with the same amount of saline.The changes of IL-6,CRP and PCT in synovial fluid of all rabbits were measured.Took synovial fluid and 3 pieces of synovial membrane to make bacterial culture 9 days after surgery.Results The bacterial culture of the experimental group was positive for staphylococcus aureus while it was negative in the control group.One day after surgery,the levels of IL-6,CRP and PCT in synovial fluid of two groups were significantly higher than those before operation(P<0.05),but the difference between the two groups was not statistically significant(P>0.05).The levels of IL-6,CRP and PCT in the synovial fluid of the experimental group were significantly higher than those of the control group at the 3rd,6th and 9th day after operation(P<0.05).There was a significant increase in IL-6,CRP and PCT levels in the two groups at the first and 3rd day after operation(P<0.05).At 6th days after operation,the levels of IL-6,CRP and PCT in synovial fluid of experimental group continued to increase(P<0.05),and they began to decrease in the control group (P<0.05).PCT gradually restored preoperative level at 6th days after operation(P>0.05),and IL-6 and CRP gradually restored preoperative level at 9th days after operation(P>0.05).The sensitivity and specificity of IL-6,CRP and PCT combined test were 96.9% and 90.6% respectively,which were significantly higher than those of individual indexes.Conclusion Combined detection of serum IL-6,CRP and PCT levels in the infection model of artificial femoral head replacement can provide some help for the early diagnosis of infection after clinical hip arthroplasty.

OBJECTIVETo construct the acid-sensitive potassium hannel-3(TASK3) eukaryotic expression plasmid and to establish a stable SH-SY5Y cell line expressing enhanced green fluorescent protein (EGFP)-tagged TASK3.

METHODSTASK3 coding region was subcloned into pEGFP-N1 plasmid to construct a recombinant vector alled pEGFP-TASK3. The correct recombinant expressing plasmid was transfected with X-feet transfection reagent to SH-SY5Y cells. The cell line stably expressiing EGFP tagged-TASK3 gene was established by screening with antibiotic G418 and fluorescence microscope. The expression and localization of the EGFP tagged-TASK3 fusion protein was detected by Western blot and confocal microscope. Exposure of the SH-SY5Y cell line expressing stably TASK3-eGFP fusion proteins was exposed to different pH media (7.0, 6.7, 6.4, 6.1) for 24 h, the cell viability was assessed with cell counting Kit-8 (CCK-8).

RESULTSAll the results of identification by PCR, digestion with restriction endonuclease and sequencing indicated that the recombinant eukaryotic expression plasmid pEGFP-TASK3 was constructed correctly. The stable SH-SY5Y cell line expressing EGFP tagged-TASK3 fusion protein was successfully established. Exposure of the wild type SH-SY5Y cells and the stable SH-SY5Y-GFP tag-TASK3 cell line to different pH media (7.0, 6.7, 6.4, 6.1) for 24 h, the cell viability of two group cells significantly reduced with pH declining, and the difference was statistically significant (P < 0.05). Compared with wild type SH-SY5Y cells, the cell viability of stable SH-SYSY-GFP tag-TASK3 cell line increased significantly with the same pH media, and the difference was statistically significant (P < 0.05).

CONCLUSIONThe eukaryotic expression vector pEGFP-TASK3 is successfully constructed and the cell line stably expressing TASK3-eGFP fusion is established which is important for their fundamental research and potential applications.

Blotting, Western ; Cell Line ; Gene Expression ; Genetic Vectors ; Green Fluorescent Proteins ; genetics ; Humans ; Plasmids ; Polymerase Chain Reaction ; Potassium Channels, Tandem Pore Domain ; genetics ; Transfection