Cytokine-Induced Cell Surface Expression of Adhesion Molecules in Vascular Endothelial Cells In vitro

Regulation of the adhesion molecules expression by cytokine in vascular endothelial cells was investigated. Human umbilical vein endothelial cells (HUVEC) were stimulated with cytokines, TNF α (1-250 U/ml) or IL 1β (0.1-50 U/ml) for 24 h. HUVEC were also cultured with cytokines, TNF α (100 U/ml) or IL 1β (10 U/ml), for 4-72 h, cell surface expression of adhesion molecules (ICAM 1 and VCAM 1) were detected and quantitated by immunocytochemical methods and computerized imaging analysis technique. Adhesion molecules expression were up regulated by TNF α, IL 1β in a concentration and time dependent manner. Some significant differences were observed between the effects of cytokines on the ICAM 1 and on VCAM 1 expression. Cytokines might directly induce the expression of ICAM 1 and VCAM 1 in vascular endothelial cells. Our observations indicate differential functions of the two adhesion molecules during the evolution of inflammatory responses in stroke.

Changes in serum cellular adhesion molecule and matrix metalloproteinase-9 levels in patients with cerebral infarction following hyperbaric oxygen therapy A case and intergroup control study

BACKGROUND： Animal studies have confirmed that hyperbaric oxygen （HBO） therapy can reduce matrix metalloproteinase activity and blood brain barrier permeability, thereby exhibiting neuroprotective effects. However, at present, consensus does not exist in terms of its clinical efficacy. OBJECTIVE： To validate the significance of changes in serum cellular adhesion molecule and MMP-9 levels in patients with cerebral infarction following HBO therapy. DESIGN, TIME AND SETTING： This randomized, controlled, neurobiochemical study was performed at the Department of Neurology, Affiliated Hospital of Qingdao University Medical College between December 2002 and March 2006. PARTICIPANTS： A total of 112 patients with acute cerebral infarction of internal carotid artery, comprising 64 males and 48 females, averaging （67 ±11） years, were recruited and randomized to a HBO group （n = 50） and a routine treatment group （n = 62）. An additional 30 gender- and age-matched normal subjects, consisting of 17 males and 13 females, averaging （63 ± 9） years, were enrolled as control subjects. METHODS： The routine treatment group received routine drug treatment and rehabilitation exercise. HBO treatment was additionally performed in the HBO group, once a day, for a total of 10 days. MAIN OUTCOME MEASURES： Serum levels of soluble intercellular adhesion molecule, soluble vascular cell adhesion molecule, soluble E-selectin, and matrix metalloproteinase-9 were detected by enzyme linked immunosorbent assay. RESULTS： Upon admission, serum levels of soluble intercellular adhesion molecule, soluble vascular cell adhesion molecule, soluble E-selectin, and matrix metalloproteinase-9 were significantly increased in patients with cerebral infarction, compared with control subjects （P 〈 0.01）. Following HBO and routine treatments, serum levels of the above-mentioned indices were significantly reduced in the HBO and routine treatment groups （P 〈 0.01）. Moreover, greater efficacy was observed in the HBO group, compared with the routine treatment group （P 〈 0.05 or P 〈 0.01）. CONCLUSION： Intergroup comparison and case-control results indicated that HBO noticeably reduced serum levels of soluble intercellular adhesion molecule, soluble vascular cell adhesion molecule, soluble E-selectin, and matrix metalloproteinase-9.

Higher CO_2-insufflation pressure inhibits the expression of adhesion molecules and the invasion potential of colon cancer cells

AIM： To investigate the influence of CO2-insufflation pressure on adhesion, invasion and metastatic potential of colon cancer cells based on adhesion molecules expression. METHODS： With an/n vitro artificial pneumoperitoneum model, SW1116 human colon carcinoma cells were exposed to CO2-insufflation in 5 different pressure groups： 6 mmHg, 9 mmHg, 12 mmHg, 15 mmHg and control group, respectively for 1 h. Expression of E-cadherin, ICAM-I, CD44 and E-selectin was meas- ured at 0, 12, 24, 48 and 72 h after CO2-insufflation using flow cytometry. The adhesion and invasion capacity of SW1116 cells before and after exposure to CO2-insufflation was detected by cell adhesion/invasion assay in vitro. Each group of cells was injected intraperitoneally into 16 BALB/C mice. The number of visible abdominal cavity tumor nodules, visceral metas-tases and survival of the mice were recorded in each group. RESULTS： The expression of E-cadherin, ICAM-1, CD44 and E-selectin in SWl116 cells were changed significantly following exposure to CO2 insufflation at different pressures （P 〈 0.05）. The expression of E-cadherin, CD44 and ICAM-1 decreased with increasing CO2-insufflation pressure. The adhesive/ invasive cells also decreased gradually with increasing pressure as determined by the adhesion/invasion assay. In animal experiments, the number of abdominal cavity tumor nodules in the 15 mmHg group was also significantly lower than that in the 6 mmHg group （29.7± 9.91 vs 41.7±14.90, P = 0.046）. However, the survival in each group was not statistically different. CONCLUSION： CO2-insufflation induced a temporary change in the adhesion and invasion capacity of cancer cells in vitro. Higher CO2-insufflation pressure inhibited adhesion, invasion and metastatic potential in vitro and in vivo, which was associated with reduced expression of adhesion molecules.

Growth-associated protein 43 and neural cell adhesion molecule expression following bone marrow-derived mesenchymal stem cell transplantation in a rat model of ischemic brain injury

BACKGROUND： Transplantation of bone marrow-derived mesenchymal stem cells （BMSCs） improves motor functional recovery, but the mechanisms remain unclear. OBJECTIVE： To investigate expression of growth-associated protein 43 （GAP-43） and neural cell adhesion molecule following BMSC transplantation to the lateral ventricle in rats with acute focal cerebral ischemic brain damage. DESIGN, TIME AND SETTING： A randomized, controlled, animal experiment using immunohistochemistry was performed at the laboratories of Department of Neurology, Renmin Hospital of Wuhan University and Doctoral Scientific Research Work Station of C-BONS PHARMA, Hubei Province, China, from January 2007 to December 2008. MATERIALS： Monoclonal mouse anti-rat 5-bromo-2-deoxyuridine and neural cell adhesion molecule antibodies were purchased from Sigma, USA; monoclonal mouse anti-rat GAP-43 antibody was purchased from Wuhan Boster, China. METHODS： Rat models of right middle cerebral artery occlusion were established using the thread method. At 1 day after middle cerebral artery occlusion, 20μL culture solution, containing 5×10^5 BMSCs, was transplanted to the left lateral ventricle using micro-injection. MAIN OUTCOME MEASURES： Scores of neurological impairment were measured to assess neural function. Expression of GAP-43 and neural cell adhesion molecule at the lesion areas was examined by immunohistochemistry. RESULTS： GAP-43 and neural cell adhesion molecule expression was low in brain tissues of the sham-operated group, but expression increased at the ischemic boundary （P 〈 0.05）. Transplantation of BMSCs further enhanced expression of GAP-43 and neural cell adhesion molecule （P 〈 0.05） and remarkably improved neurological impairment of ischemic rats （P 〈 0.05）. CONCLUSION： BMSC transplantation promoted neurological recovery in rats by upregulating expression of GAP-43 and neural cell adhesion molecule.

Novel strategies for the treatment of inflammatory bowel disease: Selective inhibition of cytokines and adhesion molecules

The etiology of inflammatory bowel disease （IBD） has not yet been clarified and immunosuppressive agents which non-specifically reduce inflammation and immunity have been used in the conventional therapies for IBD. Evidence indicates that a dysregulation of mucosal immunity in the gut of IBD causes an overproduction of inflammatory cytokines and trafficking of effector leukocytes into the bowel, thus leading to an uncontrolled intestinal inflammation. Such recent advances in the understanding of the pathogenesis of IBD created a recent trend of novel biological therapies which specifically inhibit the molecules involved in the inflammatory cascade. Major targets for such treatment are inflammatory cytokines and their receptors, and adhesion molecules. A chimeric anti-TNF-α monoclonal antibody, infiiximab, has become a standard therapy for CD and it is also likely to be beneficial for UC. Several anti-TNF reagents have been developed but most of them seem to not be as efficacious as infliximab. A humanized anti-TNF monoclonal antibody, adalimumab may be useful for the treatment of patients who lost responsiveness or developed intolerance to infliximab. Antibodies against IL-12 p40 and IL-6 receptor could be alternative new anti-cytokine therapies for IBD. Antiinterferon-γ and anti-CD25 therapies were developed, but the benefit of these agents has not yet been established. The selective blocking of migration of leukocytes into intestine seems to be a nice approach. Antibodies against α4 integrin and α4β7 integrin showed benefit for IBD. Antisense oligonucleotide of intercellular adhesion molecule 1 （ICAM-1） may be efficacious for IBD. Clinical trials of such compounds have been either recently reported or are currently underway. In this article, we review the efficacy and safety of such novel biological therapies for IBD.

Expression of cell adhesion molecule CD44 in gastric adenocarcinoma and its prognostic importance

AIM: To evaluate the relation of cluster of differentiation 44 (CD44) expression with clinicopathological features of gastric adenocarcinoma, and also its effect on prognosis with an emphasis on the differences between intestinal and diffuse types. METHODS: From 2000 to 2006, 100 patients with gastric adenocarcinoma, who had undergone total or subtotal gastrectomy without any prior treatment, were studied. Haematoxylin & eosin (HE) staining was used for histological evaluation, including the type (Lauren's classifi cation) and grading of the tumor. The expression of CD44 in the gastric adenocarcinoma mucosa and the adjacent mucosa were determined by immunohistochemistry. The survival analysis was obtained using the Kaplan-Meier test. RESULTS: Of 100 patients, 74 (74%) patients were male. The tumors were categorized as intestinal type (78%) or diffuse type (22%). Sixty-five percent of patients were CD44-positive. CD44 expression was not detected in normal gastric mucosa. Rather, CD44 was more commonly expressed in the intestinal subtype (P = 0.002). A signifi cant relation was seen between the grade of tumor and the expression of CD44 (P = 0.014). The survival analysis showed a poor prognosis of patients with CD44-positive tumors (P = 0.008); and this was more prominent in the intestinal (P = 0.001) rather than diffuse type. CONCLUSION: Cell adhesion molecule CD44 is highly expressed in gastric adenocarcinoma. CD44 expression is correlated with a poor prognosis in patients with the intestinal type of gastric adenocarcinoma. CD44 can, therefore, be utilized as a prognostic marker for this group of patients.

Role of adhesion molecules and dendritic cells in rat hepatic/renal ischemia-reperfusion injury and anti-adhesive intervention with anti-P-selectin lectin-EGF domain monoclonal antibody

AIM: To investigate the role of P-selectin, intercellular adhesion molecule-1 (ICAM-1) and dendritic cells (DCs)in liver/kidney of rats with hepatic/renal ischemiareperfusion injury and the preventive effect of anti-Pselectin lectin-EGF domain monoclonal antibody (anti-PsLEGFmAb) on the injury.METHODS: Rat models of hepatic and renal ischemiareperfusion were established. The rats were then divided into two groups, one group treated with anti-PsL-EGFmAb(n = 20) and control treated with saline (n = 20). Both groups were subdivided into four groups according to reperfusion time (1, 3, 6 and 24 h). The sham-operated group (n = 5) served as a control group. DCs were observed by the microscopic image method, while P-selectin and ICAM-1 were analyzed by immunohistochemistry.RESULTS: P-selectin increased significantly in hepatic sinusoidal endothelial cells and renal tubular epithelial cells 1 h after ischemia-reperfusion, and the expression of ICAM-1 was up-regulated in hepatic sinusoid and renal vessels after 6 h. CD1a+CD80+DCs gradually increased in hepatic sinusoidal endothelium and renal tubules and interstitium 1 h after ischemia-reperfusion, and there was the most number of DCs in 24-h group. The localization of DCs was associated with rat hepatic/renal function.These changes became less significant in rats treated with anti-PsL-EGFmAb.CONCLUSION: DCs play an important role in immune pathogenesis of hepatic/renal ischemia-reperfusion injury.Anti-PsL-EGFmAb may regulate and inhibit local DC immigration and accumulation in liver/kidney.

Relationship between cell adhesion molecules expression and the biological behavior of gastric carcinoma

AIM: To evaluate the relationship between the expression of cell adhesion molecules (CAMs) and the biological behavior of gastric carcinoma. METHODS: Expression of syndecan-1, E-cadherin and integrin β3 were evaluated by immunohistochemical study in a total of 118 gastric carcinomas and 20 non- tumor gastric mucosas. RESULTS: The expressions of syndecan-1 and E-cadherin were significantly lower in gastric carcinoma compared to non-tumor gastric mucosa, and the low expression rates were positively correlated to the tumor invasion depth, vessel invasion, lymph node metastasis and distant metastasis (P < 0.01 in all cases). However, the expression of integrin β3 was significantly higher in gastric carcinoma compared to non-tumor gastric mucosa, and the high expression rates were positively correlated to the tumor invasion depth, vessel invasion, lymph node metastasis and distant metastasis (P < 0.01 in all cases). In addition, the three protein expressions were correlated to the tumor growth pattern (P < 0.01, P < 0.01, and P < 0.05 respectively), but not correlated to tumor differentiation (P > 0.05, P > 0.05 and P > 0.05 respectively). Positive correlation was observed between the expressions of syndecan-1 and E-cadherin, but they which were negatively correlated to the expression of integrin β3 (P < 0.01 in all cases). Univariate analysis demonstrated that the mean survival time and 5-year survival rate were lower in the cases with low expressions of syndecan-1 and E-cadherin and high expression of integrin β3 (P < 0.01, in all cases). COX multivariate analysis showed that the expression level of syndecan-1 could be an independent prognostic index of gastric carcinoma (P < 0.01), whereas E-cadherin and integrin β3 could not be independent indexes (P > 0.05, P > 0.05 respectively). CONCLUSION: The low expression of syndecan-1 and E-cadherin and the high expression of integrin β3 are significantly correlated with the invasion and metastasis of gastric carcinoma, and they are highly correlated with each other. Therefore they may serve as important prognostic markers of gastric carcinoma.

Amelioration of experimental colitis by Astragalus membranaceus through anti-oxidation and inhibition of adhesion molecule synthesis

AIM: To investigate the protective effects of Astragalus membranaceus(Am) against hapten-induced colitis in male Sprague-Dawley rats as well as its underlying mechanism.METHODS: Experimental colitis was induced in rats by enema administration of 2,4-dinitrobenzene sulfonic acid (DNBS). Rats were either pretreated with Am extract (2 or 4 g/kg, p.o. once daily) starting from 10 d before DNBS enema, or received Am post-treatment (2 or 4 g/kg, p.o.twice daily) on the three consecutive days following DNBS administration. Colonic lesion area and histological damage were determined, while the activities of myeloperoxidase (MPO) and xanthine oxidase, as well as reduced glutathione (GSH) content were measured in the excised colonic tissues. Besides, protein expression of inducible nitrite oxide synthase (iNOS), intercellular adhesion molecule-1 (ICAM-1) and P-selectin was also detected by Western blot analysis.RESULTS: Our findings had shown that both macroscopic lesion area and histological colonic damage induced by DNBS were significantly reduced by both Am pre- and post-treatments. These were accompanied by attenuation of the elevated colonic MPO activity and downregulation of the iNOS, P-selectin, and ICAM-1 protein expression.Besides, deprivation of colonic GSH level under colitis condition was also preserved.CONCLUSION: These results demonstrate that Am possesses both preventive and therapeutic potential in experimental colitis. The anti-inflammatory actions involve anti-oxidation along with inhibition of adhesion molecule synthesis in the colonic tissues.

Expression pattern of epithelial cell adhesion molecule on normal and malignant colon tissues

AEM: To investigate the expression pattern of epithelial cell adhesion molecule (Ep-CAM) on normal and malignant colon tissues to evaluate its diagnostic and therapeutic significance. METHODS: cDNA encoding Ep-CAM extracellular domain was doned by reverse transcription-polymerase chain reaction (RT-PCR) from excised malignant colon tissues and inserted into a glutathione S-transferase (GST)-tagged vector. Ep-CAM-GST fusion protein was induced by isopropyl-p-D-thiogalactopyranoside (IPTG) and purified with glutathione-sepharose. The Ep-CAM-GST fusion protein was mixed with Freund's adjuvant and Balb/c mice were immunized with it. Sp2/0 myeloma cells were fused with the spleen cells of the immunized mice. After having selected by indirect ELISA, the anti-Ep-CAM monoclonal antibodies (MAbs) were generaled and the corresponding ascites were obtained. Finally, the human colon carcinoma tissue array prepared from seventy individual patients was stained with the anti-Ep-CAM MAbs. RESULTS: The isdated Ep-CAM cDNA sequence was identical to the data in GenBank. The expressed fusion protein was almost soluble and had a molecular weight (MW) of 53 ku. Four MAbs against Ep-CAM were obtained and designated as FMU-Epl, FMU-Ep2, FMU-Ep3 and FMU-Ep4 respectively. Among them, FMU-Ep4 could recognize the natural Ep-CAM on Colo205 and SW480 cells, and all of them could be used for immunohistochemical staining of tissue sections. It was fbund that Ep-CAM was distributed differently in normal and various malignant colon tissues, induding squamous cell carcinoma, signet-ring cell carcinoma and adenocarcinoma. In normal colon gland epithelia, Ep-CAM antigen was mainly distributed on the basolateral membrane and in the region between the basolateral membrane and the cytoplastic part near the nuclei, whereas the expression pattern of colon malignancies was mainly on the whole surface of epithelia and the expression was much higher than the normal colon tissues. The staining pattern of tissue array showed in adenocarcinoma and papillary adenocarcinoma, and the expression of Ep-CAM was increased from grade I to grade Ⅲ. CONCLUSION: MAbs against Ep-CAM might be useful for research on the structure and function of Ep-CAM and may have diagnostic and therapeutic value to various colon carcinomas.

Regulation of LOX-1 on adhesion molecules and neutrophil infiltration in mouse Aspergillus fumigatus keratitis

AIM:To determine whether lectin-like ox-LDL receptor(LOX-1)regulates adhesion molecules expression and neutrophil infiltration in Aspergillus fumigatus(A.fumigatus)keratitis of C57 BL/6 mice.METHODS:C57 BL/6 mice were pretreated with a neutralizing antibody to LOX-1(5μg/5μL)or control nonspecific IgG(5μg/5μL),LOX-1 inhibitor Poly-I(2μg/5μL)or PBS by subconjunctival injection.Fungal keratitis(FK)mouse models of C57 BL/6 mice were established by scraping corneal central epithelium,smearing A.fumigatus on the corneal surface and covering the eye with contact lenses.The corneal response to infection was assessed via clinical score.The mRNA levels of the adhesion molecules intercellular cell adhesion molecule-1(ICAM-1),vascular cell adhesion molecule-1(VCAM-1),P-selectin and E-selectin were tested in control and infected corneas by reverse transcriptionpolymerase chain reaction(RT-PCR).The protein levels of ICAM-1 were evaluated by immunofluorescence(IF)and Western blot.Neutrophils were extracted from the abdominal cavity of C57 BL/6 mice followed by pretreatment using antibody to LOX-1(10μg/mL)or control nonspecific IgG(10μg/mL),the Poly-I(4μg/mL)or PBS.The cells were then stimulated with A.fumigatus and tested mRNA and protein levels of lymphocyte function-associated antigen-1(LFA-1)using RT-PCR and Western blot.IF and myeloperoxidase(MPO)assays were used to assess neutrophil infiltration in mice corneas.RESULTS:Pretreatment of LOX-1 antibody or the Poly-I reduced the degree of inflammation of cornea and decreased the clinical FK score compared with pretreatment of IgG or PBS(both P<0.01).And these pretreatment also displayed an obvious decline in the mRNA levels of ICAM-1,VCAM-1,P-selectin,E-selectin and LFA-1 expression compared with control groups(all P<0.01).Furthermore,pretreated with LOX-1 antibody or Poly-I,the protein levels of ICAM-1 and LFA-1 also decreased compared with control groups(all P<0.05).Neutrophil infiltration in the cornea was significantly reduced after pretreatment of LOX-1 antibody or Poly-I compared with control groups by IF and MPO assays(both P<0.01).CONCLUSION:Inhibition of LOX-1 can decrease the expression of adhesion molecules and reduce neutrophil infiltration in A.fumigatus infected corneas of C57 BL/6 mice.

Effect of Batroxobin on Expression of Neural Cell Adhesion Molecule in Temporal Infarction Rats and Spatial Learning and Memory Disorder

The effect of Batroxobin expression of neural cell adhesion molecule (NCAM) in left temporal ischemic rats with spatial memory disorder was investigated by means of Morri's water maze and immunohistochemical methods. The results showed that the mean reaction time and distance of temporal ischemic rats for searching a goal were significantly longer than those of sham-operated rats and at the same time NCAM expression of left temporal ischemic region was significantly increased. However, the mean reaction time and distance of Batroxobin-treated rats were shorter and they used normal strategies more often and earlier than those of ischemic rats. The number of NCAM immune reactive cells of Batroxobin-treated rats was more than that of ischemic group. In conclusion, Batroxobin can improve spatial memory disorder of temporal ischemic rats and the regulation of the expression of NCAM is probably related to the neuroprotective mechanism.

Antrodia Cinnamomea ameliorates neointimal formation by inhibiting infl ammatory cell infi ltration through downregulation of adhesion molecule expression in vitro and in vivo

The increased vascular infl ammation is a key event in the development of atherosclerotic lesions.Antrodia cinnamomea has been shown to promote anticancerogenic activity through decreasing infl ammation.However,the potential role of A.cinnamomea in cardiovascular diseases remains unexplored.Herein,using carotid arterial ligation models,we found that ethanol extract from A.cinnamomea(EEAC)signifi cantly inhibited neointimal hyperplasia in a dose-dependent manner,accompanied with the reduced expression of activated p65 and infl ammatory cytokines.We also show that EEAC ameliorated TNF-α-induced phosphorylation of p65 and pro-infl ammatory cytokine expression in both vascular smooth muscle cells(VSMCs)and macrophages in vitro.Mechanistically,EEAC suppressed expression levels of intercellular adhesion molecule-1(ICAM-1)and vascular cell adhesion molecule(VCAM-1)in VSMCs,which attenuates the ability of monocytes/macrophages adhesion to VSMCs.Furthermore,the expression level of these adhesion molecules and infi ltration of monocytes/macrophages were also decreased in neointimal VSMCs of arteries pretreated with EEAC.Altogether,our results reveal a novel function of A.cinnamomea in suppressing vascular infl ammation upon ligation injury during neointimal formation,likely through inhibition of infl ammatory cell infi ltration via downregulating the adhesion molecules in VSMCs.Thus,A.cinnamomea may offer a pharmacological therapy to slow down disease progression in patients with vascular injury.

Nerve bundle formation during the promotion of peripheral nerve regeneration:collagenⅥ-neural cell adhesion molecule 1 interaction

The formation of nerve bundles,which is partially regulated by neural cell adhesion molecule 1(NCAM1),is important for neural network organization during peripheral nerve regeneration.However,little is known about how the extracellular matrix(ECM)microenvironment affects this process.Here,we seeded dorsal root ganglion tissue blocks on different ECM substrates of peripheral nerve ECM-derived matrixgel,Matrigel,laminin 521,collagen I,and collagen IV,and observed well-aligned axon bundles growing in the peripheral nerve ECM-derived environment.We confirmed that NCAM1 is necessary but not sufficient to trigger this phenomenon.A protein interaction assay identified collagen VI as an extracellular partner of NCAM1 in the regulation of axonal fasciculation.Collagen VI interacted with NCAM1 by directly binding to the FNIII domain,thereby increasing the stability of NCAM1 at the axolemma.Our in vivo experiments on a rat sciatic nerve defect model also demonstrated orderly nerve bundle regeneration with improved projection accuracy and functional recovery after treatment with 10 mg/m L Matrigel and 20μg/m L collagen VI.These findings suggest that the collagen VI-NCAM1 pathway plays a regulatory role in nerve bundle formation.This study was approved by the Animal Ethics Committee of Guangzhou Medical University(approval No.GY2019048)on April 30,2019.

Perinodular ductular reaction/epithelial cell adhesion molecule loss in small hepatic nodules

AIM: To investigate if loss of epithelial cell adhesion molecule (EpCAM) is associated with microinvasion in hepatocellular carcinomas (HCCs) in the presence of chronic hepatitis B.

Alterations in the polysialylated neural cell adhesion molecule and retinal ganglion cell density in mice with diabetic retinopathy

AIM：To investigate the impact of polysialylated neural cell adhesion molecule（PSA-NCAM）on the survival of retinal ganglion cells（RGCs）in the experimentally induced diabetes in mice.METHODS：Diabetes was induced in 2.5 months old Swiss Webster mice by intraperitoneal injection of streptozotocin（STZ,90 mg/kg）once daily for two consecutive days.Examination of the proteins of interest in the retinas from diabetic mice at 2mo after diabetes induction was performed using immunohistochemistry and Western blot analysis.RGCs were counted in the wholemounted retinas,and Brn3a marker was used.RESULTS：Examination of retinas from diabetic mice at 2mo after diabetes induction revealed a considerable reduction in RGC density.Our experiments also demonstrated a redistribution of PSA-NCAM in the retina of diabetic animals.PSA-NCAM immunoreactivity was diminished in the inner part of the retina where RGCs were located.In contrast,an enhanced PSA-NCAM immunoreactivity was detected in the outer layers of the retina.PSA-NCAM signal was co-localized with glial fibrillary acidic protein immunoreactivity in the Müller cell branches.Previous studies have shown that matrix metalloproteinase-9（MMP-9）is responsible for the reduction in PSA-NCAM levels in neuronal cells.The reduced levels of PSA-NCAM in inner layers（nerve fiber layer,ganglion cell layer）were accompanied by the increased expression of MMP-9.In contrast,in the outer retinal layers,the expression of MMP-9 was much less pronounced.CONCLUSION：MMP-9 induces PSA-NCAM shedding in the inner part of the retina and the decreased level of PSA-NCAM in the inner part of the retina might be,at least in part,responsible for the loss of RGCs in diabetic mice.

Expression changes of nerve cell adhesion molecules L1 and semaphorin 3A after peripheral nerve injury

The expression of nerve cell adhesion molecule L1 in the neuronal growth cone of the central nervous system is strongly associated with the direction of growth of the axon, but its role in the regeneration of the peripheral nerve is still unknown. This study explored the problem in a femoral nerve section model in rats. L1 and semaphorin 3A m RNA and protein expressions were measured over the 4-week recovery period. Quantitative polymerase chain reaction showed that nerve cell adhesion molecule L1 expression was higher in the sensory nerves than in motor nerves at 2 weeks after injury, but vice versa for the expression of semaphorin 3A. Western blot assay results demonstrated that nerve cell adhesion molecule L1 expression was higher in motor nerves than in the sensory nerves at the proximal end after injury, but its expression was greater in the sensory nerves at 2 weeks. Semaphorin 3A expression was higher in the motor nerves than in the sensory nerves at 3 days and 1 week after injury. Nerve cell adhesion molecule L1 and semaphorin 3A expressions at the distal end were higher in the motor nerves than in the sensory nerves at 3 days, 1 and 2 weeks. Immunohistochemical staining results showed that nerve cell adhesion molecule L1 expression at the proximal end was greater in the sensory nerves than in the motor nerves; semaphorin 3A expression was higher in the motor nerves than in the sensory nerves at 2 weeks after injury. Taken together, these results indicated that nerve cell adhesion molecules L1 and semaphorin 3A exhibited different expression patterns at the proximal and distal ends of sensory and motor nerves, and play a coordinating role in neural chemotaxis regeneration.

Cognitive disorder and changes in cholinergic receptors, N-methyl-D aspartate receptors, neural cell adhesion molecule, and brain-derived neurotrophic factor following brain injury

BACKGROUND： Learning and memory damage is one of the most permanent and the severest symptoms of traumatic brain injury; it can seriously influence the normal life and work of patients. Some research has demonstrated that cognitive disorder is closely related to nicotine cholinergic receptors, N-methyl-D aspartate receptors, neural cell adhesion molecule, and brain-derived neurotrophic factor. OBJECTIVE： To summarize the cognitive disorder and changes in nicotine cholinergic receptors, N-methyl-D aspartate receptors, neural cell adhesion molecule, and brain-derived neurotrophic factor following brain injury. RETRIEVAL STRATEGY： A computer-based online search was conducted in PUBMED for English language publications containing the key words ＂brain injured, cognitive handicap, acetylcholine, N-methyl-D aspartate receptors, neural cell adhesion molecule, brain-derived neurotrophic factor＂ from January 2000 to December 2007. There were 44 papers in total. Inclusion criteria： ① articles about changes in nicotine cholinergic receptors, N-methyl-D aspartate receptors, neural cell adhesion molecule, and brain-derived neurotrophic factor following brain injury; ② articles in the same researching circle published in authoritative journals or recently published. Exclusion criteria： duplicated articles. LITERATURE EVALUATION： References were mainly derived from research on changes in these four factors following brain injury. The 20 included papers were clinical or basic experimental studies. DATA SYNTHESIS： After craniocerebral injury, changes in these four factors in brain were similar to those during recovery from cognitive disorder, to a certain degree. Some data have indicated that activation of nicotine cholinergic receptors, N-methyl-D aspartate receptors, neural cell adhesion molecule, and brain-derived neurotrophic factor could greatly improve cognitive disorder following brain injury. However, there are still a lot of questions remaining; for example, how do these factors change at different time points after brain injury, and what is the relationship between associated factors and cognitive disorder. CONCLUSION： It is necessary to comprehensively study some associated factors, to analyze their changes and their relationship with cognitive disorder following brain injury, and to investigate their effects at different time points after brain injury.

DIFFERENTIAL EXPRESSION OF ADHESION MOLECULES (CD44,CD29,ICAM-1 AND E-CADHERIN) IN OVARIAN CANCER SK-OV-3ip1 CELLS GROWN AS MONOLAYER AND MULTICELLULAR AGGREGATES

Objective: To detect mRNA levels and expression ofCD44, CD54, CD29 and E-cadherin (E-cad) and to discuss their relationship with formation and drug resistance ofovarian cancer SKOV3ip1 multicellular aggregates.Methods: Liquid overlay system was employed to obtainmulticellular aggregates. mRNA levels and expression ofCD44, CD54, CD29 and E-cad were investigated with RTPCR and flow cytometry (FCM) respectively. Results:Compared with monolayer cells, RT-PCR results showed a decrease in CD44 mRNA level by 0.626-fold and a decrease in CD29 mRNA level by 0.792-fold in multicellularaggregates. However, an increase in CD54 mRNA level by 1.815-fold and an increase in E-cadherin mRNA level by1.344-fold were found in multicellular aggregates. Theresults revealed the downregulation of CD44 and CD29 and the upregulation of CD54 and E-cad genes activity. CD44 expression in monolayer cells and multicellular aggregates were 75.995?.046 and 50.700?.351 (%) respectively andthere was a significant decrease in multicellular aggregates (P=0.001). Compared with control cells, no expression of CD54 was detected in monolayer cells (P=0.563) but markedly elevated CD54 expression was detected in multicellular aggregates (15.780?.217) (%) (P<0.01). High expression of CD29 was seen in monolayer cells and also in multicellular aggregates with positive rates of 96.290+1.201 (%) and 92.494?.055 (%). However, the expression of CD29 in multicellular aggregates was significantly reduced (P=0.014). Also no expression of E-cadherin was found in monolayer cells compared with control cells (4.490?.283) (%) (P=0.65) while significantly increased expression in aggregates cells (17.258?5.572) (%) (P=0.003) was observed. Conclusion: Significant differences in mRNA levels and expression of CD44, CD54, CD29 and E-cadherin clearly exist between monolayer cells and multicellular aggregates, which may be associated with the formation of multicellular aggregates and its drug resistance.

Effect of Troglitazone on Expression of Adhesion Molecules and eNOS in Human Saphenous Vein Gaft

To investigate whether peroxisome proliferators-activated receptor-γ （PPARγ） ligand Troglitazone can reduce endothelial injury and activation during storage of harvested saphenous vein grafts. Segments of human saphenous vein graft were collected from 9 patients undergoing coronary bypass surgery and then divided into two equal parts of control and test specimens, were stored in ei- ther heparinized blood （control group） or heparinized blood containing 20 μmol/L troglitazone （test group） for 1 h at room temperature. Tissue distribution and protein expression of VCAM-I, ICAM-I, and endothelial nitric oxide synthase （eNOS） were compared using immunohistochemistry and Western blot analysis. Myeloperoxidase （MPO） activity, a marker of neutrophil sequestration in human saphenous vein grafts, was also measured in each group. The expression of ICAM-1 （753±132 versus 7201±934; P〈0.01） , VCAM-1 （3731±294 versus 8292±793; P〈0.01）, and MPO activity （1.52±0.42 U/g, 5.04±1.26 U/g P〈0.01） were significantly lower in test group. In contract, eNOS expression （7983±834 versus 3989±1008; P〈0.01） was significantly higher in test group. PPARγ ligand troglitazone might reduce endothelial injury during the storage period of human saphenous vein grafts.