AIM: To investigate whether small GTPase RhoA's downstream effector Rho kinase mediates burn serum-induced endothelial hyperpermeability. METHODS: Primary cultured rat dermal microvascular endothelial cells (DMECs) were exposed to serum isolated from burned or sham burn rats for 6 hours and 8 hours, respectively, and did or didn't pretreated or post-treated with Y-27632 (30 ?mol/L), a specific inhibitor of Rho kinase. ECs were then prepared for routine scanning electron microscopy observation, or stained with rhodamine-phalloidin for F-actin visualization. Permeability to FITC-albumin was evaluated using EC monolayers. RESULTS: Stimulation with 15% burn serum for 6 h changed the ultrastructure on cellular surface of DMECs with appearance of ripple marks instead of microvillus. The small protuberances at cellular lateral were shorten and the gaps were seen between adjacent cells. Post-treatment of Y-27632 reversed the changes of ultrastructure on the cellular surface. Burn serum induced a striking reorganization of actin cytoskeleton with a weakening of fluorescent intensity of the peripheral filament bands and formation of the long and thick stress fibers, lamellipodia and filopodia. The stress fibers were diminished by pretreatment or post-treatment of Y-27632. But lamellipodia and filopodia were not influenced by pretreatment or post-treatment of Y-27632. Pre-treatment of Y-27632 also attenuated significantly the increase in EC monolayer permeability stimulated by burn serum for 6 h. However, post-treatment of Y-27632 could not attenuated burn serum-induced endothelial hyperpermeability response although their Pa values were lower than simple burn serum group's. CONCLUSION: These findings indicate that Rho kinase is involved in the mediation of burn serum-induced endothelial actin cytoskeleton reorganization and early stage of barrier dysfunction. [

Objective To explore the effects of cotransplantation with osteoblasts on hematopoietic reconstitution in mice after bone marrow transplantation (BMT). Methods The typical model of syngeneic BMT was established. 18 Balb/c mice were used to prepare the bone marrow nuclear cells and osteoblasts for BMT. The 42 Balb/c mice were randomly divided into 3 group:normal group (6 mice, without any treatment), the single BMT group ( 18 mice, given 2 × 106 bone marrow nuclear cells/each mouse) and the cotransplantation group of HSC with osteoblaats (18 mice,given 2 × 106 bone marrow nuclear cells and osteoblasts/each mouse). The following factors were measured on day 7, 14, 21 after BMT: peripheral blood cells, bone marrow mononuclear cells (BMMNC), the percentage of CD34+ cells in BMMNC (assayed by flow cytometry), the hematopoietic tissue changes (detected by HPIAS-1000 image analysis system) and micro vascular density (MVD) of bone marrow tissue (with immunohistochemistry). Results The levels of periphral WBC, RBC, PLT, BMMNC in the contransplantation group were higher than those in the single BMT group (P＜0. 01 or P＜0. 05). In the contransplantation group, the percentage of CD34+ cells in BMMNC, the hematopoietic tissue area and the MVD of bone marrow were also higher than the single BMT group on the 7th, 14th, 21st day after BMT(P＜0.01 or P＜0.05). Conclusion Cotransplantation with osteoblasts could significantly promote hematopoietic reconstruction in mice after BMT. Cotransplantation may represent a promising means of achieving higher engraftment rate after BMT.

Objective To study the expression of NM-23 ,PCNA,p53 and S-100 in colorectal cancer tissues and their relationship.Methods Three hundred and seventy eases of elderly eolorectal cancer were labelled with NM-23,PCNA,p53 and S-100 and staged by pTNM classification.Results The positive rates of NM-23,PCNA,p53 and S-100 were 74.9% ,60.3% ,58.1% ,68.1% respectively.In elderly group,the expression of NM-23 had a neg-ative correlation with that of PCNA (r = -0.101 ,P = 0.026).However the expression of NM-23 had a positive corre-lation with that of p53 (r =0.111 ,P =0.016 ),the expression of S-100 had a positive correlation with that of p53 (r=0.112,P = 0.015 ),the expression of S-100 had a positive correlation with that of PCNA (r =-0.229,P =0.000).Conclusions In the elderly patients,NM-23,p53 and S-100 were significantly correlated with the inva-sion,metastasis and prognosis of eoloreetal cancer.PCNA may be a stimulative factor in the invasion and metastasis of colorectal cancer.NM-23 ,PCNA,p53 and S-100 can serve as effective markers in reflecting the invasion,metas-tasis and prognosis of colorectal carcinoma in the elderly.

Objective To establish an Apak gene stable and permanent knockout cell line using CRISPR/Cas9 system in human colon cancer cells ( HCT116 cells), and study the effect of Apak knock-out on p53 activity and apoptosis. Methods The lentiCRISPR v2-sgRNA Apak expression plasmid was co-transfected with lentivirus coated plasmids pSPAX2 and pMD2.G.The supernatant was collected, filtered, and used to infect HCT116 cells.The positive clones were screened out by puromycin culture and Western blot was used to detect Apak knockout cell lines.Luciferase reporter gene assay, flow cytometry analysis and colony formation assay were used to examine p53 activity and apoptosis of Apak knockout cells, respectively.Results Apak knockout HCT116 cell lines were generated in which p53 activity and apoptosis were increased,but the colony formation was decreased.Conclusion The Apak stable knockout cell lines of HCT116 are successfully generated by CRISPR/Cas9 system for further functional study.

Objective To approach Professor Zhu Guangli's regular pattern of differentiation of symptoms and signs in patients with hypertension accompanied by complications and in accord with the individual manifestations to add or subtract the ingredients in the prescription.Methods The patients with essential hypertension selected were those who the first time came to the Affiliated Guangxing Hospital of Zhejiang Chinese Medical University or Hangzhou Hospital of Traditional Chinese Medicine (TCM) of Zhejiang Province from September 2012 to January 2015, and were continuously followed up in the hospitals having their clinical data comprehensive. The complications, combination diseases and symptoms of hypertension were conducted as dependent variables, while the applied Chinese medicines were acted as independent variables. QUEST algorithms were employed to analyze the different dependent variables to evaluate the importance contribution rate (ICR) of every kind of Chinese medicine.Results There were 9 types of complications or combination diseases [including hyperlipemia, coronary heart disease, cardiac arrhythmia, hyperglycemia, trace albuminuria, lacunar cerebral infarction, elevation of creatinine in blood (azotemia), liver dysfunction and cardiac dysfunction] and 39 types of symptoms obtained (including dizziness, palpitation, chest oppression, insomnia, lack of strength, soreness of the waist and knees, dry mouth, loose stool, fatigue, apontaneous sweating, headache, shortness of breath, coprostasis, fullness in the stomach, blurred vision, limb numbness, low spirit, chest pain, poor appetite, abdominal distension, excessive dreaming, tinnitus, flush of the face, neck rigidity, hectic fever, bitterness in the mouth, cough, ructus, nocturia, fremitus, stenagma, pharyngalgia, heaviness in head, nocturnal sweating, frequent urination, coughing of sputum, chillness and cold limbs, aphthous stomatitis and sialosis) being the QUEST decision models. All kinds of the TCM employed in all the patients paying visits for 755 times were summed up, and the individual drug frequency of presentation in the prescription was recorded. In the study, 171 sorts of TCM and 10 620 of cumulative frequency (freq.) were involved in 755 prescriptions. The average sorts of TCM was 14.07±1.53 per prescription. The top ten kinds of TCM the most commonly used (freq.) were as follows: tuckahoe (588), Chinese yam (551), wolfberry fruit (496), white peony root (442), abalone shell (405), gastrodia tuber (395), lilyturf root (325), Huai wheat (293), curcuma root (281), rush pith (277), accounting for 38.16% in the total frequency. There were 33 kinds of TCM with frequency > 100, accounting for 74.65% in the total frequency. By the analyses of QUEST algorithms directing to the complications and combination diseases to add or subtract the items of TCM, it was discovered that the drugs with ICR > 5% were as follows: in lacunar cerebral infarction, the drug application frequencies or ICRs of batryticated silkworm and bitter cardamom were 32.69% and 50.74% respectively; in coronary heart disease, the ICRs of pseudostellaria root, astragalus root, white sandal wood and lepidium seed were 23.05%, 17.70%, 15.31% and 5.06% respectively; in cardiac arrhythmia, the ICRs of astragalus root, flavescent sophora root, Huai wheat and dragon tooth and dragon bone were 46.65%, 11.56%, 7.56%, 7.13% and 6.28% respectively; in cardiac dysfunction, the ICRs of lepidium seed and white hyaciath bean were 92.97% and 7.03%; in hyperlipemia, the ICRs of cassia seed, white sandal wood, dried rehmannia root and sargentgloryvine stem were 24.26%, 12.47%, 10.51% and 5.81% respectively; in hyperglycemia, the ICRs of trichosanthes root and dried hawthorn fruit were 51.02% and 30.18%; in trace albuminuria, the ICR of snow in June herb was 83.33%; in elevation of creatinine in blood, the ICRs of Chinese actinidia root, herb of snow in June, glossy privet fruit and centella were 81.37%, 6.21%, 6.21% and 6.21% respectively; in liver dysfunction, the ICRs of coral ardisia root, purse grass and herb of stringy stonecrop were 86.37%, 6.82% and 6.82% respectively. When the symptoms were used as the dependent variables, for the top 3 commonly seen symptoms, in the prescription, the items of TCM used with ICR > 5% were as follows: in cases with elevation of blood pressure, parasitic loranthus (ICR 89.59%) was used; with dizziness, gastrodia tuber (ICR 46.96%) and/or abalone shell (ICR 33.61%) were added; headache, rhizome of Chinese ligusticum and/or rhizome of Sichuan lovage and/or large gentian with ICR at 85.77%, 7.11% and 7.11% respectively. Conclusion Professor Zhu's essential thought in differentiation of disease and expectant treatment of hypertension in accord with the individual manifestations to add and subtract drugs in TCM is under the guidance of TCM principle of differentiation of symptoms and signs for treatment, and based on the classical theories in TCM, the reliable drugs whose actions are demonstrated clinically and experimentally are preferentially selected.

Sodium succinate (0.8 g?kg~ -1,ig) could significantly shorten afterdischarge duration (ADD), lower Racine’s score on amygdala electrical kindling in rat.Additional sodium succinate capsule (0.25g,po) was effective on patients with epilepsy administrated with common antiepileptic drugs which showed little effectiveness on these patients. The results supposed sodium succinate would be effective in epilepsy therapy.

AIM: To investigate the changes of the renal L-arginine /nitric oxide pathway and the relationship of L-arginine transport between kidney and erythrocytes in spontaneously hypertensive rat (SHR). METHODS: Sixteen week old SHR, 16 week old SHR with captopril (CAP) treated for four weeks and 16 week old WKY rats were used in the experiment. L-arginine transport, NO synthase(NOS) activity, nitrite and cyclic GMP (cGMP) content were measured in renal tissue or erythrocytes. RESULTS: In the renal tissue, compared with that of WKY group, the Vmax of high-or low-affinity L-arginine transporter, NOS activity, NO - 2 and cGMP content of SHR group were significantly decreased ( P

Objective To describe the status of fatigue and its influencing factors among hypertension patients. Methods A total of 1253 hypertension patients were investigated with global fatigue severity subscale,family APGAR index and self-designed questionnaire. Results 36.7% of hypertension patients were fatigue.Univariate analysis showed that the determinants of fatigue among hypertension patients included older, normal marriage, rural, low educational, mental, family dysfunction (P ＜ 0.05 ). Multivariate analysis revealed that age ( OR =2. 612,95% CI = 2. 047 ～ 3. 333 ), normal marriage ( OR = 1. 754,95% CI = 1. 259 ～ 2. 445 ), manual ( OR =0.810,95 % CI = 0.698 ～ 0.939) and family dysfunction ( OR = 1.717,95 % CI = 1. 396 ～ 2.111 ) were associated with fatigue. Conclusion The rate of status is high in the hypertension patients. The behavior is related to age,marital status, labor type and family function.

Objective: To observe the effect of pyridoxamine and telmisartan on the indexes of myocardial remodeling in spontaneous hypertensive rats (SHR). Methods: A total of 48 male SHR at 22 weeks of age were randomly divided into 4 groups:①Hypertension (H) control group, the rats received distilled water 2 ml/d,②telmisartan (T) group, the rats received telmisartan 6 mg/(kg.d),③pyridoxamine (P) group, the rats received pyridoxamine 200 mg/(kg.d),④Combination (TP) group, the rats received both telmisartan and pyridoxamine. All animals were treated for 16 weeks and the systolic blood pressure (SBP) was measured before and after treatment. After treatment, the serum levels of nitric oxide (NO) and super oxide dismutase (SOD) were examined by chemiluminescent method, advanced glycation end-products (AGEs) was detected by ELISA, left ventricular weight index and collagen volume fraction (CVF) in myocardial tissue were calculated, the mRNA expression of myocardial receptor of advanced glycation end products (RAGE) was evaluated by real time RT-PCR. Results: Compared with H group, SBP levels were decreased in T and TP groups,P<0.01, while SBP in P group stayed similar,P>0.05. Compared with H group, the serum levels of NO and SOD were increased in T, P and TP groups,P<0.01; while compared with T group and P group, the levels of NO and SOD were further increased in TP group,P<0.05. Compared with H group, left ventricular weight index and CVF were decreased in T, P and TP groups,P<0.01; while compared with T group and P group, left ventricular weight index and CVF were further decreased in TP group,P<0.05. Pyridoxamine and telmisartan had the coordinative effect on left ventricular weight index and CVF, P<0.05. The serum levels of AGEs were lower in T group (5.99 ± 0.51) mg/L, P group (5.57 ± 0.91) mg/L and TP group (5.24 ± 0.63) mg/L than that in H group, (6.71 ± 0.50) mg/L,P<0.01; while compared with T group and P group, AGEs was further decreased in TP group,P<0.05. The mRNA expressions of RAGE were reduced in T group (0.035 ± 0.010), P group (0.036 ± 0.005) and TP group (0.024 ± 0.007) than that in H group (0.053 ± 0.010),P <0.01; while compared with T group and P group, the mRNA of RAGE was further reduced in TP group, P<0.05. Conclusion: Pyridoxamine could improve the oxidative stress and the indexes of myocardial remodeling independently from decreasing blood pressure in SHR. Combined intervention of P and T may break the coordination systems between AGEs-RAGE and renin-angiotensin, therefore further improve the indexes of myocardial remodeling in SHR.