Objective: To examine the pathologcial changes of glial cells after chronic optic nerve compression, so as to discuss the interaction between the glial cells and neurons. Methods: Thirty adult cats were randomly divided into 6 groups (n= 5), namely, control group, sham operation group, 1-week compression group, 2-week compression group, 4-week compression group, and 8-week compression group. The chronic optic nerve injury model was produced by an inflatable balloon implanted under the optic chiasm in the latter 4 groups. All the animals were sacrificed and perfused; the optic nerves were removed and the cellular responses of the nerves were observed by electronic microscopy; and the expression of glial fibrillary acidic protein (GFAP), myelin basic protein (MBP), carbonic anhydrase II (CA II) and ED-1 was exmined at various time points by immunohistochemical staining. Results: Under the electron microscopy, the normal optic nerves had integrated myelin structure, clearly and closely arranged neural plate. The optic nerve presented slight demyelination 2 weeks after compression; myelin laminalle dissociating and glial cell degenerating occurred 4 weeks after compression; and the demyelination became more obvious and the most myelin became thiner 8 weeks after compression. No obvious immunohistochemistral changes were found in the optic nerves during the first two weeks of compression. The MBP staining was disturbed and lost at 4 weeks after compression, which became more obvious 8 weeks after compression. The CA II staining in the compressed region was irregular and lost at 4 weeks, which was more obvious at 8 weeks; the staining in non-compressed region was normal. The intensity of GFAP staining was reduced in the compressed region and increased at proximal portion of the nerve at 4 weeks, which became more significant at 8 weeks. The ED-1 positive cells were found in the normal nerve with low density. The positive cells increased around the compressed region at 4 weeks and became more significant at 8 weeks. Conclusion: Glial cell degeneration and death occur in the compressive region afte chronic compression of optic nerve. The proximal portion of the compressed nerve has astrocyte proliferation and microglia activation, indicating that functional change of glial cells may contribute to chronic optic nerve injury.

Objective: To clone, identify and analyze subtractive cDNA libraries of differentially expressed genes associated with chronic optic nerve compression in cats. Methods: The constructed cDNA libraries were amplified by E. coli transformation with calcium chloride and were subjected to blue and white screening. Three hundred positive bacterial clones were randomly chosen from each library and were identified by colony PCR. The positive clones were sequenced to screen for the differentially expressed genes. The identified sequences were then analyzed for homology using Blast program against the DNA database bank of Japanese through internet. Real-time PCR was performed to verify the expression of the 4 differential clones. Results: One thousand positive clones were identified by PCR and 674 ESTs were obtained by seqeuncing, with length ranging from 200 to 800bp. Results from Blast analysis revealed 14, 20, 23 and 19 homolog genes in 4-w forward, 4-w reverse, 8-w forward and 8-w reverse subtractive libraries, respectively. These genes fell into several functional groups, such as energy and metabolism, ion transport, gene transcription, signal transduction, cellular injury and repair, and MHC molecules. Result of real-time PCR verified that the 4 clones were differentially expressed. Conclusion: The constructed subtractive cDNA libraries lay an experimental basis for further identification and study of the differentially expressed genes related to chronic optic nerve injury.

Objective: To construct subtractive cDNA libraries of differentially expressed genes associated with chronic optic nerve injury in cats. Methods: Fifteen adult cats were randomly divided into 3 groups (n=5): control group, 4-w compression group and 8-w compression group. The chronic optic nerve injury was produced by an inflatable balloon implanted under the optic chiasm. The total RNA was prepared from optic nerves of each group by TRIzol method. Double-stranded cDNA was produced by SMART PCR cDNA synthesis protocol. Suppression subtractive hybridization (SSH) was used to isolate the cDNA fragments of differentially expressed genes in the optic nerves after 4-w and 8-w compression. The cDNA fragments were directly inserted into T/A cloning vector to establish the subtractive library,followed by amplification of the libraries through E. coli transformation with calcium chloride and screening of blue and white clones. Three hundred positive bacterial clones were randomly picked in each library and identified by colony PCR. Results: Analysis of the white clones by PCR showed that 80% clones contained 200-800 bp inserts in each library. Conclusion: Four subtractive cDNA libraries of differentially expressed genes associated with chronic optic nerve injury have been successfully constructed by SSH and T/A cloning techniques,which lays a solid foundation for screening and cloning specific differentially expressed genes associated with chronic optic nerve injury.

Carcinoma, Squamous Cell ; genetics ; Cell Division ; Chromosome Deletion ; Humans ; Laryngeal Neoplasms ; genetics ; Telomere ; genetics ; Tumor Cells, Cultured

Abstract?AlM: To observe the anterior chamber angle shape of patients with diabetic retinopathy ( DR) at different stages using OCT, and to explore a more sensitive and accurate glaucoma preventing measure.?METHODS: A total of 613 cases of DR patients were divided into proliferation phase group ( 353 cases ) and background phase group ( 260 cases ) , another 100 cases of patients with only diabetes were selected as control group. OCT was used to detect the anterior chamber angle related parameters, including angle opening distance ( AOD) , trabecular-iris space area ( TlSA) , angle recess area ( ARA ) , anterior chamber depth ( ACD ) and anterior chamber angle ( ACA) .?RESULTS: There were significant difference among AOD500, AOD750, TlSA500, TlSA750, ARA500 and ARA750 of control group, proliferation phase group and background phase group ( P< 0. 01 ). The proliferation phase group was the lowest, AOD500, AOD750, TlSA500, TlSA750, ARA500 and ARA750 were 0. 60 ± 0. 22mm, 0. 78 ± 0. 39mm, 0. 22 ± 0. 12mm2 , 0. 45 ± 0. 19mm2 , 0. 29 ± 0. 18mm and 0. 46 ± 0. 15mm, respectively. There were significant difference among ACA500, ACA750, ACD500 and ACD750 of control group, proliferation phase group and background phase group ( P< 0. 01 ). The proliferation phase group was the lowest, ACA500, ACA750, ACD500 and ACD750 were 30. 29o ± 8. 19o, 21. 20o ± 7. 40o, 2. 32 ± 0. 23μm and 2. 52±0. 16μm, respectively. ln addition, ROC results indicated that all these indices exert significant difference on evaluating the stage of DR. ?CONCLUSlON: Anterior chamber angle shape detected using OCT can reflect the change of retinopathy. OCT is an accurate and simple technique for detecting DR staging.

Humans ; Monitoring, Intraoperative ; methods ; Perioperative Care ; Sleep Apnea, Obstructive ; nursing ; therapy

Objective To establish an early diagnostic test for tuberculous meningitis (TBM) with good sensitivity and specificity. Methods Twenty-five patients with a clinical diagnosis of TBM and 49 controls, including 27 patients with other infectious diseases of central nervous system and 22 patients with noninfectious neurological diseases, were enrolled in our research. We simultaneously detected antimycobacterium bovis BCG IgG secreting cells in both cerebral spinal fluid (CSF) and peripheral blood (PBL)by enzyme linked immunospot assay(ELISPOT),repeated insertion sequence IS6110 specific for mycobacterium tuberculosis in CSF by PCR and anti-BCG IgG titre in both CSF and PBL by enzyme linked immunosorbent assay(ELISA).Results The sensitivity of ELISPOT was 84.0%,much higher than that of PCR(75.0%)and ELISA(52.3%).The specificities of the three tests were 91.8%,93.7%and 91.8%respectively.The numbers of CSF cells secreting anti-BCG IgG tested by ELISPOT were even higher in the early phase of TBM, but declined along with the disease progressing(t=-3.183,P=0.008),which allowed an early diagnosis to be made. Conclusion ELISPOT technique is proved to be the most valuable test for the early diagnosis of TBM.

Animals ; Calcitriol ; pharmacology ; Diabetes Mellitus, Experimental ; complications ; Diabetes Mellitus, Type 2 ; complications ; Liver ; drug effects ; Liver Diseases ; complications ; drug therapy ; Male ; Rats ; Rats, Sprague-Dawley

This study is to investigate the effect of ibudilast on apoptosis of airway eosinophil in asthmatic guinea pigs and its mechanism. Experimental asthma model of guinea pigs was induced with ovalbumin (OVA). Differential count in BALF was examined. The apoptosis of eosinophils (EOS) was labeled with TdT-mediated dUTP nick end labeling (TUNEL) technique. Fas mRNA expression of EOS was detected by reverse transcription-polymerase chain reaction (RT-PCR). The quantification of GM-CSF and IL-5 in BALF was conducted with ELISA. After treatment of ibudilast, the number of EOS and the quantification of GM-CSF and IL-5 decreased significantly. The number of apoptotic cells as well as Fas mRNA expression of EOS obviously increased. The results indicated that anti-asthma mechanisms of ibudilast can antagonize asthma through decreasing the number of EOS, inducing apoptosis of EOS, enhancing Fas mRNA expression of EOS and reducing the content of GM-CSF and IL-5.
Animals ; Anti-Asthmatic Agents ; pharmacology ; Apoptosis ; drug effects ; Asthma ; metabolism ; pathology ; Bronchoalveolar Lavage Fluid ; Eosinophils ; cytology ; drug effects ; metabolism ; Female ; Granulocyte-Macrophage Colony-Stimulating Factor ; metabolism ; Guinea Pigs ; Interleukin-5 ; metabolism ; Male ; Pyridines ; pharmacology ; fas Receptor ; metabolism

Stable isotope labeling by amino acids in cell culture (SILAC) is an in vivo metabolic labeling technique based on the abundance ratio of mass spectrometry. Combined with different methods of proteins affinity enrichment, SILAC can obtain sensitive, accurate and quantitative information of proteins. This article reviewed the history, rationale, characteristics and application of SILAC, with special focus on its role in studying protein interactions with small molecules, quantitative proteomics and post-translational modification of proteins.