Effects of cartilage oligomeric matrix protein on BMP-2 induced cell differentiation of mesenchymal stem cells / 中华骨科杂志

Objective To investigate the effect of overexpression of cartilage oligomeric matrix protein (COMP) on BMP-2 induced cell osteogenic and chondrogenic differentiation of mesenchymal stem cells (MSCs).Methods MSCs,transfected with plasmid DNA encoding recombinant human COMP,were induced to differentiate into osteocytes and chondrocytes by BMP-2.Realtime PCR of osteogenic related markers (Col1a1,RUNX2,OPN,BGP) and chondrogenic related markers (Col2a1,SOX9,Aggrecan) were performed to evaluate the process of cell differentiation.ALP staining,Alizarin red S staining for osteogenic differentiation and alcian blue staining for chandrogenic differentiation were conducted to evaluate the tendency of cell differentiation.Results Real-time PCR assay presented the significantly higher (P＜0.05) COMP expression of MSCs when COMP gene was transfected into cells.The expression level of OPN was significantly (P＜0.05) down-regulated at all the time points in experimental group compared with that in control group.A final significant (P＜0.05) up-regulation of expression appeared in experimental group at the late stage of induction (day 7,14) compared with that in control group,even though a decrease (P＜0.05) expression of Col1a1,RUNX2 and BGP in experimental group occurred at the early stage of induction (day 3).The expression of Aggrecan and Col2a1 in experimental group was up-regulated (P＜0.05) at different time points compared with that in control group.And a significant higher (P＜0.05) expression of SOX9 in experimental group only appeared at day 7 compared with that in control group.ALP staining and Alizarin red S staining were weakened while alcian blue staining was enhanced.Conclusion COMP may inhibit BMP-2 induced osteogenic differentiation and promote BMP-2 induced chondrogenic differentiation,which may provide new insight for cartilage tissue engineering.

Enhanced integrin-mediated human osteoblastic adhesion to porous amorphous calcium phosphate/poly (L-lactic acid) composite / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>The initial osteoblastic adhesion to materials characterizes the first phase of cell-material interactions and influences all the events leading to the formation of new bone. In a previous work, we developed a novel amorphous calcium phosphate (ACP)/poly(L-lactic acid) (PLLA) material that demonstrated morphologic variations in its microstructure. The aim of this study was to investigate the initial interaction between this material and osteoblastic cells. Cellular attachment and the corresponding signal transduction pathways were investigated.</p><p><b>METHODS</b>A porous ACP/PLLA composite and PLLA scaffold (as a control) were incubated in fetal bovine serum (FBS) containing phosphate-buffered saline (PBS), and the protein adsorption was determined. Osteoblastic MG63 cells were seeded on the materials and cultured for 1, 4, 8, or 24 hours. Cell attachment was evaluated using the MTS method. Cell morphology was examined using scanning electron microscopy (SEM). The expression levels of the genes encoding integrin subunits α1, α5, αv, β1, focal adhesion kinase (FAK), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were determined using real-time reverse transcription polymerase chain reaction (RT-PCR).</p><p><b>RESULTS</b>The ACP/PLLA material significantly increased the protein adsorption by 6.4-fold at 1 hour and 2.4-fold at 24 hours, compared with the pure PLLA scaffold. The attachment of osteoblastic cells to the ACP/PLLA was significantly higher than that on the PLLA scaffold. The SEM observation revealed a polygonal spread shape of cells on the ACP/ PLLA, with the filopodia adhered to the scaffold surface. In contrast, the cells on the PLLA scaffold exhibited a spherical or polygonal morphology. Additionally, real-time RT-PCR showed that the genes encoding the integrin subunits α1, αv, β1, and FAK were expressed at higher levels on the ACP/PLLA composite.</p><p><b>CONCLUSIONS</b>The ACP/PLLA composite promoted protein adsorption and osteoblastic adhesion. The enhanced cell adhesion may be mediated by the binding of integrin subunits α1, αv, and β1, and subsequently may be regulated through the FAK signal transduction pathways.</p>

Controllable synthesis of different size hydroxyapatite nanoparticles and their biological effects on osteoblast-like cells / 中华骨科杂志

ObjectiveTo investigate the size effect of hydroxyapatite nanoparticles on proliferation and apoptosis of osteoblast-like cells.MethodsCetyltrimethylammnonium bromide (CTAB) was used to regulate the size of nano hydroxyapatite (nHAP) particles.All obtained particles were characterized by transmission electron microscopy (TEM),X-ray diffraction,dynamic light scattering and chemical analysis.HAP films were obtained by slowly coating cover glasses with 1％ HAP particle suspension.MG-63 cells on three different films(20HAP,40HAP and 80HAP) were cocultured for up to 5 days.Cell proliferation assay was obtained by methyl thiazolyl tetrazolium (MTT).Cell apoptosis was detected by flow cytometric detection.Cell ultrastructure morphology was observed by TEM observation.ResultsnHAP with diameter of 20 nm,40 nm and 80 nm were synthesized and and analyzed.The MG-63 cells were cultured on three different fihns.The optical density value of cells on 20HAP was 1.22±0.13 after 5 days incubation,and there was no different compared to the control group(F=6.843,P=0.124).Cell number and viability were significantly higher on 20HAP compared to large nHAP after 5 days incubation.The percentage of apoptotic cells increased with increasiug nHAP particle size.TEM images showed 20HAP was found in cytoplasm and cell morphology had no changes.ConclusionBoth cell proliferation and cell apoptosis are related to the size of the nHAP particles.20HAP was the most effective on promoting cell growth and inhibiting cell apoptosis.

Mental Status of the Patients with Irritable Bowel Syndrome / 中国康复理论与实践

@#ObjectiveTo analyze the mental status of the patients with irritable bowel syndrome (IBS).MethodsSixty IBS patients were evaluated with Self Rating Anxiety Scale (SAS), Self Rating Depression Scale (SDS), Symptions Check List-90 (SCL-90) and self-edited scale.ResultsThe primary causes of mental disorder of IBS patients were the stress events and lacking knowledge for IBS; the average SAS scores of sixty IBS patients were 52.32±10.19, SDS scores were 54.90±11.70; the average scores of SAS and SDS in the groups of midlife and young people were significantly higher than that in the group of agedness ( P<0.05). The total scores of SAS and SDS were not significantly difference among the patients with different culture level and sex ( P>0.05); except the scores of crankiness and opposition, the average SCL-90 scores of the sixty IBS patients were significantly higher than the normal ( P<0.05).ConclusionIBS relates to mental status and stress, proper mental intervention can ease the symptom and improve the quality of the living.

生活事件在老年期抑郁症发病中的作用 / 中国康复理论与实践

@#目的探讨生活事件心理社会因素在老年期抑郁症发病中的作用。方法应用生活事件量表(LES)对100例老年期抑郁症患者及100例正常老人进行对照研究。结果老年期抑郁症生活事件尤其负性生活事件频度及严重度均高于正常对照组;老年期抑郁症患者的生活事件中主要是健康问题和家庭问题。结论心理社会因素是老年期抑郁症发病的重要高危因素

Expression of heme oxygenase in injured lung following ischemia-reperfusion of limbs in rats / 中国病理生理杂志

AIM:To observe the changes in heme oxygenase-1(HO-1) expression in the lung after ischmia-reperfusion of hind limbs in rats.METHODS:Hind limbs ischemia was made by clamping infrarenal aorta with a microvascular clip and lung injury was made by following reperfusion. Lung tissue was obtained from the animals subjected to sham operation, 4 h ischemia without reperfusion and 4 h, 8 h, 16 h, 24 h, 48 h reperfusion following 4 h ischemia. The levels of HO-1 mRNA and protein were measured at different times by Northern blot and Western blot. Immunohistochemistry technique was used to determine the cell types responsible for limb ischemic reperfusion induced HO-1 expression. RESULTS:After ischemia-reperfusion of limbs, HO-1 mRNA increased by 4 h, reached a peak at 16 h, and returned toward baseline at 24-48 h. This time course correlated with increased HO-1 protein. Immunohistochemical studies showed HO-1expressed in a variety of cell types, including the airway epithelium, alveolar macrophages and vascular smooth muscular cells. There were no positive signals in sham group and ischemia group both in mRNA levels and protein levels. CONCLUSION:The expression of HO-1 in the lung is not induced by limb ischemia or sham operation, but induced by limb reperfusion after ischemia in rats.

Brain injury induced by ischemia-reperfusion of rat hindlimbs and its mechanisms / 中国病理生理杂志

AIM:To investigate the pathologic changes in the brain and its underlying mechansims during ischemia-reperfusion of rat hindlimbs.METHODS:SD rats were divided into the normal(N), sham(S), 4 h ischemia without reperfusion(I), and 4 h ischemia-2, 6,12,18 or 24 h reperfusion (I-R) groups at random. Ischemia and ischemia-reperfusion were established with the occlusion or/and re-opening of the terminal of abdominal aorta, respectively. The pathologic changes in the brain tissue were morphologically observed. The expression of inducible nitric oxide synthase (iNOS) mRNA, and iNOS protein and the nitrotyrosine, a marker of peroxynitrite (ONOO-)，in the brain tissue were detected with RT-PCR and immunohistochemical technique, respectively. The brain superoxide dismutase (SOD) activity and malondialdehyde (MDA) contents were spectraphotometrically measured.RESULTS:Hydropic degeneration and severe injury to neurons were only showed in I-R group. Expressions of iNOS mRNA and protein were demonstrated in I-R, I and S groups, which were maximal in I-R 6 h group. iNOS positive neurons and microglias were more spread in I-R 6 h group than those in S and I groups. NT positive neurons were localized in the cerebral cortex and hippcampus of I-R 6 h group. The contents of MDA markedly increased, while the activity of SOD significantly decreased in I-R 6 h group compared to the N, S and I groups. There were no significant changes in MDA and SOD in N, S and I groups.CONCLUSION:Severe ischemia-reperfusion of rat hindlimbs could induce brain injury, and its mechanisms might be related to enhanced expression of iNOS-NO-ONOO- in the brain.

Expression of heme oxygenase in injured lung following ischemia-reperfusion of limbs in rats / 中国病理生理杂志

AIM: To observe the changes in heme oxygenase-1(HO-1) expression in the lung after ischmia-reperfusion of hind limbs in rats.METHODS: Hind limbs ischemia was made by clamping infrarenal aorta with a microvascular clip and lung injury was made by following reperfusion. Lung tissue was obtained from the animals subjected to sham operation, 4 h ischemia without reperfusion and 4 h, 8 h, 16 h, 24 h, 48 h reperfusion following 4 h ischemia. The levels of HO-1 mRNA and protein were measured at different times by Northern blot and Western blot. Immunohistochemistry technique was used to determine the cell types responsible for limb ischemic reperfusion induced HO-1 expression. RESULTS: After ischemia-reperfusion of limbs, HO-1 mRNA increased by 4 h, reached a peak at 16 h, and returned toward baseline at 24-48 h. This time course correlated with increased HO-1 protein. Immunohistochemical studies showed HO-1expressed in a variety of cell types, including the airway epithelium, alveolar macrophages and vascular smooth muscular cells. There were no positive signals in sham group and ischemia group both in mRNA levels and protein levels. CONCLUSION: The expression of HO-1 in the lung is not induced by limb ischemia or sham operation, but induced by limb reperfusion after ischemia in rats.

Brain injury induced by ischemia-reperfusion of rat hindlimbs and its mechanisms / 中国病理生理杂志

AIM: To investigate the pathologic changes in the brain and its underlying mechansims during ischemia-reperfusion of rat hindlimbs.METHODS: SD rats were divided into the normal(N), sham(S), 4 h ischemia without reperfusion(I), and 4 h ischemia-2, 6,12,18 or 24 h reperfusion (I-R) groups at random. Ischemia and ischemia-reperfusion were established with the occlusion or/and re-opening of the terminal of abdominal aorta, respectively. The pathologic changes in the brain tissue were morphologically observed. The expression of inducible nitric oxide synthase ( iNOS ) mRNA, and iNOS protein and the nitrotyrosine, a marker of peroxynitrite (ONOO -),in the brain tissue were detected with RT-PCR and immunohistochemical technique, respectively. The brain superoxide dismutase (SOD) activity and malondialdehyde (MDA) contents were spectraphotometrically measured.RESULTS: Hydropic degeneration and severe injury to neurons were only showed in I-R group. Expressions of iNOS mRNA and protein were demonstrated in I-R, I and S groups, which were maximal in I-R 6 h group. iNOS positive neurons and microglias were more spread in I-R 6 h group than those in S and I groups. NT positive neurons were localized in the cerebral cortex and hippcampus of I-R 6 h group. The contents of MDA markedly increased, while the activity of SOD significantly decreased in I-R 6 h group compared to the N, S and I groups. There were no significant changes in MDA and SOD in N, S and I groups.CONCLUSION: Severe ischemia-reperfusion of rat hindlimbs could induce brain injury, and its mechanisms might be related to enhanced expression of iNOS -NO-ONOO - in the brain.

Caspase-8 dependent osteosarcoma cell apoptosis induced by proteasome inhibitor MG132 / 中国病理生理杂志

AIM: To investigate the effects of proteasome inhibitor Z-LLL-CHO(MG132) on human osteosarcoma cell line MG-63 and its possibly mechanism.METHODS: After treated with different concentration of MG132,the morphological change,ultrastructral morphology,cell viability,cell apoptosis,gene transcription and protein expression in MG-63 cells were accessed by fluorescence microscope,electron microscope,MTT assay,agrose gel electrophoresis,FCM,RT-PCR and Western blotting.RESULTS: Proteasome inhibitor MG132 was an effective inducer of apoptosis in human osteosarcoma MG-63 cells.Not only apoptotic changes,but also cell arrest at G2-M-phase,the accumulation of p27kip1,the accumulation of activated caspase-8 and increased ratio of Bax∶Bcl-2 were observed.However,to normal human diploid fibroblast cells,MG132 did not show apoptosis-inducing effect.CONCLUSION: Apoptosis induced by MG132 may be caspase-8,p27kip1and bcl-2-related.

Changes of HO - 1 expression in kidney following ischemia and reperfusion of limbs and their significance in rats / 中国病理生理杂志

AIM: To detect the changes of heme oxygenase-1 (HO-1) expression in kidney following ischemia-reperfusion of hindlimbs and to elucidate their significance. METHODS: Health SD rats were randomly divided into normal (N), sham (S), 4h ischemia without reperfusion (I), 4 h ischemia-2, 6, 12, 18 or 24 h reperfusion (I-R) groups and I-R18h+zinc protoporphyrin (ZnPP) group. I-R was established using the occlusion for 4 h and re-opening for 2-24 h of the femoral arteries. In I-R 18 h+ZnPP group, ZnPP (5 ?mol?kg~(-1) body weight) was intravenously injected 6 h and 12 h after reperfusion, respectively. The expression of HO-1 mRNA in kidney was detected with reverse transcription-polymerase chain reaction (RT-PCR). The expression and location of HO-1 protein were detected with immunohistochemical technique. The observation of pathologic changes of kidney was made following the inhibition of HO-1 by ZnPP. RESULTS: The relative expression level of HO-1 mRNA significantly increased in I-R group, compared to those in the control groups, It was maximal in I-R 18 h group, and thereafter expression level of HO-1 mRNA decreased, however significant expression was still detected in I-R 24 h group (P

Changes of iNOS gene expression in kidney following ischemia and reperfusion of limbs and their significance in rats / 中国病理生理杂志

AIM: To detect the changes of inducible nitric oxide synthase (iNOS) expression in kidney following ischemia-reperfusion(I-R)of hindlimbs and to elucidate their significance. METHODS: I-R was established using the occlusion of the femoral arteries for 4 h and re-opening for 2-24 h in rats. The expression of iNOS mRNA,and iNOS protein and the nitrotyrosine (NT),a marker of peroxynitrite (ONOO -),in renal tissue were detected with reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemical technique,respectively. The superoxide dismutase (SOD) activity and malondialdehyde (MDA) contents in renal tissue were spectraphotometrically measured. The observation of pathologic changes of renal tissue was made following the inhibition of iNOS by aminoguanidine (AG). RESULTS: Compared with control group,the relative expression level of iNOS mRNA significantly increased in I-R group. There were more iNOS and more NT positive product in the epithelial cells of renal proximal convoluted tubules and thick segments of Henle′ loops in I-R group than control group. The contents of MDA markedly increased,while the activity of SOD significantly decreased in I-R group,compared to those in the control groups. The pathologic changes of kidney became milder in I-R group following the inhibition of iNOS by AG. CONCLUSION: The expression of iNOS mRNA and protein in renal tissue were significantly upregulated,excess induction of NO contributed to the kidney injury during the I-R of hindlimbs.

Changes of HO-1 gene expression in brain following ischemia and reperfusion of limbs / 中国病理生理杂志

AIM: To detect the changes of heme oxygenase-1(HO-1) expression in brain following ischemia-reperfusion of hindlimbs in rats and to elucidate their significance. METHODS: Ischemia-reperfusion was established using the occlusion for 4 h and re-opening for 2-24 h of the femoral arteries. The expression of HO-1 mRNA in brain was detected with reverse transcription-polymerase chain reaction (RT-PCR). The expression and location of HO-1 protein were detected with immunohistochemical technique. The observation of pathologic changes of brain was made following the inhibition of HO-1 by zinc protoporphyrin (ZnPP). RESULTS: The relative expression level of HO-1 mRNA significantly increased in I-R group, compared to those in control group. It was maximal in I-R 12 h group, and thereafter expression level of HO-1 mRNA decreased, however significant expression was still detected in I-R 24 h group (P

Effect of nitric oxide and peroxynitrite on lung injury induced by ischemia-reperfusion of hind limbs in rats / 中国病理生理杂志

AIM: To observe the changes in nitric oxide(NO) and peroxynitrite anion (ONOO - ) in the injuried lung following the ischemia-reperfusion of hind limbs and evaluate the contribution of NO and ONOO - to tissue injury. METHODS: A model of hind limbs ischemia was made by clamping infrarenal aorta with a microvascular clip and lung injury occurring after reperfusion. Lung tissue was obtained from the animals received sham operation(group 1),4 hours ischemia without reperfusion(group2), 1 hour reperfusion following 4 hours ischemia (group3) and 4 hours reperfusion following 4 hours ischemia (group4) . The contents of MDA, NO - 2/NO - 3 and the activities of SOD in the lung were examined. Immunohistochemical technique was used to determine the immunoreactivity to iNOS and nitrotyrosine(NT)-a specific "footprint" of peroxynitrite. RESULTS: Compared with group1 and group2,the contents of MDA and NO - 2/NO - 3 increased significantly (P

OBSERVATION ON ANTERIOR MEDULLARY VELUM OF THE RABBIT WITH GROSS DISSECTION AND SCANNING ELECTRON MICROSCOPY / 解剖学报

The present study intended to provide some informations about the relationship between the supraependymal structures (SES) and the periventrieular neural tissue (PVNT), and with gross dissection and scanning electron microscopy the anterior medullary velum (AMV) was observed on 20 adult rabbits. The AMV may be divided into three portions: 1. the posterior membraneous wall of the recess of the inferior colliculus; 2. the anterior roof of the fourth ventricle; and 3. the transiional portion between the two portions mentioned above. On the ependymal surface of the entire AMV. There were numerous microvilli and cilia, except for the second portion of AMV, there were also some spherical-like structures, which were 2-6?m in diameter and had lace-like processes on its surface, and supraependymal cells(SEC) which were stellate, triangular, and spindle in shape. On the cellular surface with very few SEC, the secretory granules may be seen, which were 0.1-0.3?m in diameter. The SEC often extended out 2-5 processes, and the distal parts of which expanded into a flattened shovel-like structure, which lay on or inserted into the ependymal surface. The SEC here are similar to type Ⅱ SEC seen in the third and fourth ventricles, but they may differ significantly in structures on cellular surface, e.g. the secretory granules, and in shapes of their processes. Thus, it may suggest that the SEC here may play an intermediary role between PVNT and CSF, and be another route of neurohumoral modulation.

THE CENTRAL CONNECTIONS AND FUNCTIONAL CONSIDERATIONS OF THE DORSAL COMMISSURAL NUCLEUS OF THE RAT——A HRP METHOD STUDY / 解剖学报

HRP solution was injected into the dorsal commissural nucleus (DCN) of segment L_6 or S_1 of the spinal cord and laterodorsal tegmental area(TLD)——take the Barrington's nucleus as its center and lateral parabrachial nucleus (PBL) of the rostral pons in different individuals of the rats. After HRP was injected into the DCN, labelled neurons and dense terminals were found in Barrington's nucleus, and labelled terminals appeared in the PBL. When the unilateral TLD was injected the labelled cells and terminals were found in the DCN and bilateral intermediate zone (IM), and formed a band of labelled neurons and terminals. When the PBL was injected the labelled neurons were observed in the DCN and bilateral IM. A few labelled neurons were found in lamina Ⅰ in the latter two experimental groups.Based on the present and previous studies, the authors got the following understandings:1. Morphylogically, the present study for the first time demonstrated that, the micturition reflex arch through the pontine consists of following parts: the primary afferent neurons of the bladder→secondary relay neurons of DCN→Barrington's nucleus→IM (mainly IML) parasympathetic preganglionic neurons→parasympathetic postganglionic neurons.2. According to the present and previous researches, the authors conjecture that, the secondary fibers of visceral sensation of the pelvic organs originate from the neurons of DCN, IM and lamina I and project into the PBL.3. According to the facts mentioned above, we presume that the DCN and bilateral IM constitute a complex and named it the 'visceral field', and which is closely associated with the pelvic organs. This field has widespread connections with the peripheral efferent and afferent nerves. On the other hand, i t contains a lot of relay neurons projecting into Barrington's nucleus and PBL and receives the terminals of descending fibers of the neurons of the Barrington's nucleus. The descending fibers also project into the Onuf's nucleus.In addition, the present study disscussed the complicated functions of the DCN systematically.