Objective To analyze the sandtray topics of the children with behavior problems according to the grounded theory method.Methods Collected and described children' s sandtray topics of different types of behavior problems,age and gender.And analyzed them base on a 3 levels coding list compiling according to the ground theory method.Results The frequency of “negative theme” (3.33 ± 1.25 ) was higher than “positive theme” (0.81 ± 1.25 ) with significant difference (P＜0.01) in feature level 1.Age factor had significant effect upon the frequency of “positive theme” in feature level 1 (P ＜ 0.05 ).And also had impact on “harmony”,“ development” in features level 2 (F =3.039,P ＜ 0.05 ).Gender factor,interaction of gender and age had significant effects on frequency of “negative theme” in feature level 1 (F =5.395,P ＜ 0.05 ; F =4.222,P ＜ 0.05 ).Conclusion The characteristics of the children' s sandtay with behavioral problems mainly are “negative theme”.And it shows different features in children with different gender or age.

Objective To explore the synergetic effect of HBX protein and M2 macrophages in inflammatory microenvironment on invasion and metastasis of hepatocellular carcinoma cells.Methods Hep3B cells were infected with recombinant lentivirus carrying HBx gene,following co-culture with THP-1 original M2 macrophages.The cells were divided into six groups:two infected groups (Hep3B +and Hep3B + + M2),four non-infected groups (Hep3B-,Hep3B-+ LV5,Hep3B-+ M2,Hep3B-+LV5 + M2).Western blot (WB) was used to assess the expression changes of E-cadherin and N-cadherin,markers of epithelial-mesenchymal transition (EMT).The cellular location of EMT markers was observed by immunofluorescence confocal microscopy.Transwell assay was used to evaluate the invasion ability of Hep3B cells.Results HBX protein overexpressed in Hep3B cells by lentivirus infection.After 72 h co-culture with M2 macrophages,WB results showed that E-cadherin descreased significantly in Hep3B+ (0.42 ±0.11) when compared with Hep3B-(1.00 ±0.18) (t =4.762,P ＜0.05),while N-cadherin was significantly higher in Hep3B + (2.85 ± 0.44) than in Hep3B-(1.00 ± 0.17) (t =4.762,P ＜ 0.05).M2macrophages decreased E-cadherin expression in Hep3 B + + M2 (0.1 ± 0.13) compared with Hep3 B + (t =3.255,P ＜0.05),while N-cadherin expression increased in Hep3B+ + M2 (4.18 ± 0.52) (t=10.009,P ＜ 0.05).Non-Infected groups didn't change the markers of E-cadherin and N-cadherin.It was suggested that invasion ability of Hep3B increased by HBx overexpression.Conclusions HBX protein and M2 macrophages synergetically mediated the invasion and metastasis of hepatocellular carcinoma cells by EMT.

Objective To investigate the regulation of protein kinase C(PKC) to the expression of β-1,4-galactusyhransferase- Ⅰ ( β-1,4-GalT- Ⅰ ) and the influence on cytoskeleton and adherence ability of human umbilical vein endothelial cells(HUVECs) when stimulated by lipopolysaccharide (LPS). Methods Cultured HUVECs were pretreated by various PKC inhibitors or phorbol 12-myristate 13-acctate( PMA), an excitomotor of PKC respectively for 30 min, then stimulated by LPS for 4 h. β-1,4-GalT-Ⅰ expression were detected by RT-PCR and Western blot, expression of β-1,4-galactosylated carbohydrate chains and cytoskeleton were assayed by immumofluorescence, and adherence ability of HUVECs was observed by endothelialmonocyte cell adherence test. Results Up-regulated expression of β-1,4-GalT- Ⅰ and β-1,4-galactosylated carbohydrate chains in HUVECs stimulated by LPS were suppressed by PKC inhibitors and increased by PMA. F-actin and β-1,4-GalT- Ⅰ were partly co-localized in HUVECs. PKC inhibitor inhibited the effect of LPS on the distribution of F-actin and β-1,4-GalT- Ⅰ. Adherence ability of HUVECs enhanced by LPS was significantly suppressed by PKC inhibitor. Conclusion PKC signal transduction pathway may participate in regulating β-1,4-GalT-Ⅰ expression in endothelial cells stimulated by LPS. Furthermore, polytypes of PKC may participate in this regulating process; PKC might regulate cytoskeleton reorganization and adherence ability of EC through β-1,4-GalT-Ⅰ during inflammation.

Objective To evaluate the role of Fas/FasL signaling pathway in ulinastatin postconditioning-induced attenuation of apoptosis in the myocardial cells of patients undergoing cardiac valve replacement with cardiopulmonary bypass (CPB).Methods Forty patients of both sexes,aged 21-59 yr,of ASA physical status Ⅱ or Ⅲ (NYHA class Ⅱ or Ⅲ),scheduled for elective cardiac valve replacement with CPB,were randomly divided into 2 groups (n =20 each):control group (group C),and ulinastatin postconditioning group (group U).In group U,ulinastatin 10 000 U/kg was perfused via the aortic root at 4 000-5 000 U·kg-1 ·min-1 starting from 5 min before aortic unclamping.In group C,the equal volume of normal saline was infused instead of ulinastatin.Myocardial specimens were taken from the right auricle at 45 min after aortic unclamping for determination of Fas,Fas ligand (FasL),caspase-8,Bcl-2 and Bax expression and cell apoptosis.The ratio of Bcl-2 expression to/Bax expression (Bcl-2/Bax) and apoptotic index were calculated.Results Fas,FasL,caspase-8 and Bax expression and apoptotic index were significantly lower,and Bcl-2 expression and Bcl-2/Bax were higher in group U than in group C.Conclusion Ulinastatin postconditioning attenuates apoptosis in the myocardial cells through inhibiting Fas/FasL signaling pathway in the patients undergoing cardiac valve replacement with CPB.

Objective To evaluate the effects of ulinastatin postconditioning and combination of ulinastatin preconditioning and postconditioning on myocardial apoptosis in patients undergoing cardiac valve replacement with cardiopulmonary bypass (CPB).Methods Eighty New York Heart Association (NYHA) class Ⅱ or Ⅲ patients of both sexes,aged 21-59 years,scheduled for cardiac valve replacement with CPB,were randomly divided into four groups (n =20 each):normal saline control group (group C),ulinastatin preconditioning group (group U1),ulinastatin postconditioning group (group U2) and ulinastatin preconditioning plus postconditioning group (group U3).In group U1,uinastatin 20000 U/kg was infused via the central vein at 500-1000 U·kg-1 · min-1 after endotracheal intubation until 10 minutes before blocking the ascending aorta.In group U2,ulinastatin 10000 U/kg was infused via the aortic root at 4000-5000 U· kg-1 · min-1 at 5-7 minutes before opening the aorta.In group U3,ulinastatin preconditioning and postconditioning were performed as described in groups U1 and U2.In group C,the same volume of normal saline was infused instead of ulinastatin.Blood samples were taken from the radial artery at 10 minutes before blocking the ascending aorta,40 minutes after blocking the ascending aorta,45 minutes after opening the aorta and at the end of operation for determination of plasma concentrations of tumor necrosis factor-alpha (TNF-α) and soluble tumor necrosis factor receptor 1 (sTNF-R1).Myocardial tissues were obtained from the right atrial appendage at 45 minutes after opening the aorta for determination of the expression of TNF-α,bcl-2,bax,caspase-3,and apoptosis.The bcl-2/bax ratio and apoptotic index were calculated.Results Plasma concentrations of TNF-α and sTNF-R1 and the expression of TNF-α,bax,caspase-3 and apoptotic index were lower and the expression of bcl-2 and bcl-2/bax ratio were higher in groups U1,U2 and U3 than in group C and they were lower in group U3 than in groups U1 and U2 (P ＜ 0.05).Conclusion Ulinastatin postconditioning can inhibit myocardial apoptosis in patients undergoing cardiac valve replacement with CPB,and the efficacy of combination of ulinastatin preconditioning and postconditioning is stronger than that of ulinastatin postconditioning.The mechanism is involved in balancing the expression of bax and bcl-2 and down-regulating the expression of TNF-α and its receptor.

Objective To detect the expressions of osteopontin (OPN) and inerleukin-18 in patients with bullous pemphigoid (BP),and to analyze their relationship with clinical and laboratory indices.Methods Enzyme linked immunosorbent assay (ELISA) was performed to quantify the serum levels of osteopontin (OPN) and inerleukin (IL)-18 in 30 patients with BP and 30 health controls.Results The serum level of OPN was statistically higher in the patients than in the healthy controls ((8.29 ± 2.76) vs.(3.88 ± 1.41 ) ng/ml,P ＜ 0.01 ),and was positively correlated with the severity of BP (r =0.658,P ＜ 0.01 ) and with some laboratory indices in the patients.Increased serum IL-18 level was observed in patients complicated by cardiovascular diseases compared with those without cardiovascular diseases ((37.49 ± 6.43) vs.(31.10 ± 5.40) pg/ml,P ＜ 0.01).Moreover,the BP patients with diabetes,tumor,hepatic and renal impairment displayed an enhanced level of serum OPN and IL-18 than those without (all P ＜ 0.05).Conclusions OPN may be positively correlated with the severity of BP,while IL-18 may be involved in the development of complications of BP.

Objective To evaluate the effects of ulinastatin on renal ischemia-reperfusion injury in patients undergoing operation on aorta with deep hypothermic circulatory arrest (DHCA ) .Methods Thirty patients ,aged 30-50 yr ,of ASA physical status Ⅲ or Ⅳ (NYHA Ⅱ or Ⅲ) ,scheduled for elective operation on aorta with DHCA ,were randomly divided into 2 groups ( n=15 each) using a random number table :control group (group C ) and ulinastatin group (group U ) .In group U ,ulinastatin 20 000 U/kg was infused via the central vein at 500-1 000 U·kg-1 ·min-1 from the time immediately after tracheal intubation until 10 min before ascending aortic cross-clamping .In group C ,the equal volume of normal saline was infused instead of ulinastatin .At 5 min before the beginning of DHCA (T1 ) and 15 min after the end of DHCA (T2 ) ,blood samples were taken from the extracorporeal circulation for determination of polymorphonuclear leukocyte counts , and plasma levels of intercellular adhesion molecule-1 , tumor necrosis factor-α, iterleukin-6 (IL-6 ) IL-8 , IL-10 , malondialdehyde , myeloperoxidase ,atrial natriuretic peptide ,cystatin C ,and creatinine .Results The polymorphonuclear leukocyte counts and plasma levels of intercellular adhesion molecule-1 , tumor necrosis factor-α, IL-6 , IL-8 , malondialdehyde , myeloperoxidase , cystatin C , and creatinine were significantly lower , and the plasma concentrations of IL-10 and atrial natriuretic peptide were higher in group U than in group C ( P< 0.05 ) . Conclusion Ulinastatin can attenuate renal ischemia-reperfusion injury in patients undergoing operation on aorta with DHCA and inhibition of inflammatory responses is involved in the mechanism .

BACKGROUND:Adipose-derived mesenchymal stem cells are gaining widespread interest in the Achil es tendon tissue engineering and regeneration, and an enabling environment (oxygen concentration) for cellinduction and differentiation is particularly necessary.
OBJECTIVE:To co-culture adipose-derived mesenchymal stem cells with primary tenocytes in standard culture condition (20%O 2 tension) and in an atmosphere of reduced oxygen (2%O 2 tension) in order to determine whether the two conditions differ in their effect on tenogenic differentiation.
METHODS:Tenocytes were isolated via serial expansion in culture from several Sprague-Dawley rats’ Achil es tendons. Adipose-derived mesenchymal stem cells were purchased. After one passage, adipose-derived mesenchymal stem cells were indirectly co-cultured with tenocytes in standard culture condition (20%O 2 tension) and in an atmosphere of reduced oxygen (2%O 2 tension). Col agen 1, col agen 3, Tenomodulin, Thrombospondin-4, Scleraxis levels were compared for each culture condition at 7, 14 and 21 days fol owing co-culturing. Immunofluorescence staining was performed to evaluate production of col agen 1 and Thrombospondin-4.
RESULTS AND CONCLUSION:Fol owing indirect co-culturing, hypoxic differentiated adipose-derived mesenchymal stem cells expressed higher levels of tendon-related genes and proteins than normoxic controls, which suggest that oxygen levels can significantly affect tenogenic differentiation, and hypoxia is advantageous for efficient differentiation of adipose-derived mesenchymal stem cells in vitro for tendon tissue engineering.

Objective To evaluate the effects of ulinastatin preconditioning on protamine-induced pulmonary injury in patients undergoing cardiac valve replacement under cardiopulmonary bypass (CPB).Methods Sixty NYHA class Ⅱ or Ⅲ and ASA physical status Ⅱ or Ⅲ patients of sexes,aged 21-59 yr,scheduled for elective cardiac valve replacement under CPB,were randomly divided into 3 groups (n =20 each):group control one protamine given via central vein (group C1) ; group control two protamine given via ascending aorta (group C2) ;group ulinastatin preconditioning (group U).Heparin was neutralized with protamine after termination of CPB.Ulinastatin 20 000 U/kg was infused via the central vein at a rate of 500-1000 U· kg-1 · min-1 starting from the time point after tracheal intubation until 10 min before cross-clamping of superior vena cava and inferior vena cava in group U.At 10 min after termination of CPB,protamine 4 mg/kg was infused over 8 min via the right internal jugular vein in groups C1 and U,or via the aortic root in group C2.Blood samples were obtained from the left atrium and right atrium at 5 min before neutralization of heparin with protamine (T1) and 15 min after neutralization of heparin with protamine (T2) for determination of polymorphonuclear leukocyte (PMN) and platelet (Plt) counts,and plasma concentrations of thromboxane B2 (TXB2) and 6-keto-prostaglandin F1α (6-keto-PGF1α).Blood samples were obtained from the left atrium at T1 and T2 for determination of the levels of TNF-α,IL-1,IL-8,CD11b/CD18,C3a,C5a,and malondialdehyde (MDA) and superoxide dismutase (SOD) activity and for blood gas analysis.Alveolar-arterial oxygen gradiant (A-aDO2),respiratory index (RI) and oxygenation index (OI) were calculated.Pulmonary arterial pressure (PAP) was recorded.Results Plt and PMN counts in the blood obtained from the left atrium were significantly lower,and plasma TXB2 concentrations in the blood obtained from the left atrium were higher at T2 in group C1,and the plasma 6-keto-PGF1α concentrations and SOD activity in the blood obtained from the left atrium were higher at T2 in groups C2 and U than those in the blood obtained from the right atrium (P ＜0.05).Compared with group C1,Plt and PMN counts and plasma 6-keto-PGF1α concentrations were significantly increased,the levels of plasma TXB2,TXB2/6-keto-PGF1α,TNF-α,IL-1,IL-8,C3a,C5a and MDA were decreased,CD11b/CD18 expression was down-regulated,PAP,A-aDO2 and RI were decreased,and OI was increased at T2 in C2 and U groups (P ＜ 0.05).There were no significant differences in the parameters mentioned above between groups C2 and U (P ＞ 0.05).Conclusion Ulinastatin preconditioning can inhibit protamine-induced pulmonary injury in patients undergoing cardiac valve replacement under CPB,and the effect is similar to that of protamine administered via the aorta.

Objective To investigate the role of phosphatidylinositol 3-kinase (PI3K)/protein-serine-threonine kinases (Akt) signal pathway in ulinastatin postconditioning-induced attenuation of apoptosis in myocardial cells in patients undergoing cardiac valve replacement with cardiopulmonary bypass (CPB).Methods Forty NYHA class and ASA physical status Ⅱ or Ⅲ patients of both sexes,aged 21-59 yr,scheduled for cardiac valve replacement with CPB,were randomly divided into 2 groups (n =20 each):normal saline control group (group C) and ulinastatin postconditioning group (group U).In group U,ulinastatin 10 000 U/kg was perfused via the aortic root at 4000-5000 U· kg-1 · min-1 starting from 5 min before aortic unclamping.In group C,the equal volume of normal saline was given instead of ulinastatin.Myocardial specimens were taken from the right auricle at 45 min after aortic unclamping for determination of the expression of Akt,phosphorylated Akt (p-Akt),cytochrome c,caspase-9,Bcl-2 and Bax,and cell apoptosis.Bcl-2/Bax ratio and apoptotic index were calculated.Results The expression of p-Akt and Bcl-2 and Bcl-2/Bax ratio were significantly higher,and the expression of cytochrome c,caspase-9 and Bax and apoptotic index were lower in group U than in group C (P ＜ 0.05).Conclusion Ulinastatin postconditioning attenuates apoptosis in myocardial cells in patients undergoing cardiac valve replacement with CPB through activating PI3K/Akt signal pathway.