Objective To investigate the expression of glucose-regulated protein 78(GRP78)and cysteine aspartic acid protease 12(Caspase-12) and evaluate the endoplasmic reticulum stress (ERS) in rats with contrast-induced nephropathy (CIN),and observe the protective effects of hydroxytyrosol on CIN rats.Methods Eighty-four Wistar rats,(220±20) g,were randomly divided into control group,CIN group,hydroxytyrosol treated group (group C+H).At 12th,24th,48th,72th day after the rats model were established,BUN and Scr were detected.ELISA were used to detect the expression of methane dicarboxylic aldehyde (MDA),superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px).HE staining were used to evaluate the pathological change of kidney.TUNEL were used to detect the apoptosis of tubular ceils.Real-time PCR were used to detect the expression of GRP78 mRNA and Caspase-12 mRNA in tubular cells.Immunohistochemistry and Western blotting were used to detect the expression of GRP78 and Caspase-12 protein in tubular cells.Results BUN,Scr,the mRNA and protein expression of GRP78,Caspase-12 in hydroxytyrosol treated group were higher than that in control group(P ＜ 0.05),but were significantly lower than that in CIN group (P ＜ 0.05).Pathological changes and the apoptosis of tubular cells in CIN group were more serious than that in hydroxytyrosol treated group (P ＜ 0.05).Conclusions Endoplasmic reticulum stress may be associated with contrast-induced nephropathy.Hydroxytyrosol can protect kidney from contrast medium via reducing the endoplasmic reticulum stress.

Objective To observe the efficacy and safety of recombinant tissue type plasminogen activa-tor (rt-PA)for the treatment of the patients with wake-up ischemic stroke (WUS)under the guidance of multimode CT. Methods Eighteen patients with WUS (a thrombolytic group)suitable for intravenous thrombolysis after multimode CT imaging screen at the Department of Neurology,Shiyan Hospital of Integrated Traditional and Western Medicine,Hubei Province from October 2012 to October 2014 were enrolled retrospectively. Twenty patients with WUS (a control group)who underwent multimode CT imaging screen were suitable for intravenous thrombolysis,but because of exceeding time window or rejecting thrombolysis and other reasons without having intravenous thrombolysis from February 2012 to February 2014 were enrolled retrospectively. The control group was treated with conventional therapy and the thrombolytic group was treated with rt-PA (0. 9 mg/kg)intravenous thrombolytic therapy. The indicators including fibrinogen (Fib),coagulation function (prothrombin time [PT ]),activated partial thromboplastin time (APTT ), platelet (PLT ),high-sensitivity C-reactive protein (hs-CRP ),National Institute of Health Stroke Scale (NIHSS )scores,and activities of daily living scores (Barthel index)at before treatment and 24 h,7 and 14 days after treatment were observed respectively. The adverse events and complications were documented and compared with the control group. Results There were no significant differences in Fib,PT,APTT, PLT,hs-CRP,NIHSS score and Barthel index before treatment between the thrombolytic group and the con-trol group (all P>0. 05);at day 7 and 14 after treatment in the thrombolytic group,compared with before treatment,Fib (14 d after treatment),PLT,and hs-CRP were decreased,PT and APTT were prolonged,the NIHSS scores were decreased,and Barthel indexes were increased. There were significant differences (all P<0. 05). At day 14 after treatment,there were significant differences in Fib,PT,APTT,hs-CRP,NIHSS scores,and Barthel indexes (Fib:3. 25 ± 0. 38 g/L vs. 3. 55 ± 0. 28 g/L;PT:15. 7 ± 3. 2 s vs. 12. 9 ± 2. 5 s;APTT:42. 7 ± 3. 5 s vs. 38. 7 ± 2. 6 s;PLT:[189 ± 26]× 109/L vs. [201 ± 23]× 109/L;hs-CRP:5. 7 ± 0. 6 mg/L vs. 11. 3 ± 2. 2 mg/L;NIHSS scores:5. 6 ± 2. 4 vs. 9. 2 ± 4. 5;and Barthel indexes:68 ± 15 vs. 47 ± 5)between the two groups (all P <0. 05). Except 1 patient occurred symptomatic intracerebral hemorrhage after thrombolysis,no other serious complications were observed in the thrombolytic group. One patient in the control group had stress gastric ulcer and bleeding,no symptomatic intracerebral hemorrhage occurred. Conclusion Multimode CT guidance can be used as a reliable imaging evidence for patients with WUS expanding intravenous thrombolytic time window. Under the multimode CT guidance, using rt-PA for intravenous thrombolytic therapy has a certain efficacy.

Objective To investigate the expression and distribution of plasmacytoid dendritic cells(pDC) in peripheral blood and renal tissues in children with Henoch-SchSnlein purpura(HSP),and explore the role of pDCs in the pathogenesis of Henoch-Schtnlein purpura nephritis(HSPN).Methods Among the 40 children with HSP,28 cases were in the active phase(renal biopsy performed in 8 cases of them) and the other 12 in remission phase.Peripheral blood mononuclear cells were isolated,and the expression of pDC was detected by flow cytometry.The normal control group was established (n =15).Total RNA of peripheral blood was extracted and transcripted into cDNA.Sybr green dye based real-time quantitative PCR method was used to compare the expression(indicated as 2-△Ct value) of CXC motif chemokine 10 (CXCL10),CC chemokine ligand 5 (CCL5),chemokine CXC subfamily receptor 3 (CXCR3),CC chemokine receptor 5 (CCR5) in children with HSP and those in the controls.Immunohistochemistry labeling technique was used to detect the distribution of pDC in renal tissues from renal biopsy,and the normal controls were established (n =3).Results The expression percentage of pDC in peripheral blood in active phase was 0.051 ± 0.039,significantly lower than those in remission phase (0.181 ± 0.082) and the normal controls (0.166 ± 0.079) (P ＜ 0.000 1).Chemokines genes CXCL10 and CCL5 were overexpressed in peripheral blood ceils of acute phase HSP children,but chemokine receptors CXCR3,CCR5 were lowly expressed compared with normal controls.There was almost no expression of pDC in the normal control renal tissues,while pDC was infiltrated in glomeruli of HSPN children.Conclusions The number of pDC and chemokines' expression in peripheral blood is abnormal,and the pathogenesis of nephritis may be involved with the pDC in peripheral blood to migrate to the renal tissues.

Objective To investigate the expression of 4-hydroxynonenal (4-HNE) in the kidney of diabetic rats and the effect of probucol.Methods The rats were being intraperitoneal injected with STZ (60 mg/kg) to establish diabetic models.Then diabetic rats were randomly divided into diabetic group (group D,n =24),probucol treated group (group P,n =24).Normal rats were taken as control group (group C,n =24).Rats in group P were treated by probucol (110 mg·kg-1·d-1); rats in group D and group C were given equal volume water instead.Scr,BUN,triglyceride (TG),total cholesterol (TC) and 24-hour urinary proteinin were measured at the 4th,8th and 12th week.PAS staining and HE staining were used to evaluate the pathological changes of the kidney.The immunohistochemistry and Western blotting were used to detect the expression of 4-HNE in renal tissue.Results Levels of Scr,BUN,TG,TC and 24-hour urinary protein in group D were higher than those in group C at the 4th,8th and 12th week(all P ＜ 0.05); Levels of Scr,BUN,TG,TC and 24-huor urinary protein in group P were lower than those in group D at 4th,8th and 12th week (all P ＜ 0.05).The pathological changes of the kidney in group D were more serious than that in group P.The expression of 4-HNE in group D were higher than group C at the 4th,8th and 12th week (all P ＜ 0.05);The expression of 4-HNE in the kidneys of group P decreased significantly compared to that of group D at the same time (P ＜ 0.05).Conclusions As an indicator of lipid peroxidation,the expression of 4-HNE significantly increases in the kidney of diabetic rat.Probucol may protect the diabetic kidney through decreasing the expression of 4-HNE and the level of lipid peroxidation.

Objective To discuss whether mild hyperuricemia can lead to kidney damage and the protection of decreased uric acid,through observing that hyperuricemia did damage to glomerulus endothelial function and cell proliferation of vascular smooth muscle in rats. Methods Fifty-four male SD rats were divided into four groups,the control group,model group (Oxonate),allopurinol group and Oxonate+allopurinol group.Rats were administered on a low sodium diet and their systolic blood pressure (SBP) were measured each 10 days.ELISA was used to detect rat plasma markers of endothelial function damage [nitric oxide (NO),type-1 plasminogen activator inhibitor (PAI-1),endothelin 1 (ET-1)] and cell proliferation of vascular smooth muscle[plateletderived growth factor (PDGF),cycloxygenase 2 (COX2),monocyte chemotactic protein-1 (MCP-1)],and the markers of inflammatory reaction[interleukin-18 (IL-18),tumor necrosis factor α(TNF-α)].PDGF and nitric oxide synthase (NOS) levels of rats were detected by immunohistochemical method.Renal tissue pathology of rats was observed. Results Compared to the control group,the plasmic concentration of COX2,ET-1,IL-18,PAI-1,PDGF,TNF-o,MCP-1 increased,and NO decreased significantly in rats of model group (all P＜0.05),expression of NOS significantly reduced and PDGF increased (all P＜0.05).Under light microscope,vascular wall thickening,intimal proliferation and lumen slight stricture without uric acid crystals in renal tissue were found in model group,which were obviously improved by using allopurinol. Conclusion Mild hyperuricemia can do damage to endothelial function of glomerulus and lead to vascular cell proliferation,which can be improved through decreasing uric acid.

Objective To discuss the effect of self- care education at the early time of postoperation and the use of nursing packages during the later period on non- throat patients. Methods 40 patients having un-dergone total laryngectomy were divided into the control group and the experimental group according to the surgery date with 20 cases in each group. The control group received conventional nursing, while the experi-mental group was given the system intervention by specialized nurses. The intervention content included self-care education at the early time of postoperation and the use of nursing packages during the later period. The questionnaire survey was carried out 15 days and 3 months after operation.The coping style,self-care ability and survival quality were compared between the two groups. Results The comparison of coping style showed re-markable difference in the two groups. The comparison of self- care ability and level of health knowledge showed significant difference 15 days after operation except two other items,but the four items demonstrated significant difference 3 months after operation. Difference also existed in the total scorn of survival quality. Conclusions Early system serf- care education after operation and the use of nursing package during the later period can impel the patients to cope positively, enhance self-care ability and survival quality.

To study the influence of recombinant endostatin on angiogenesis and tumor growth of mice H22 hepatoma, tumor models were constructed by injecting H22 hepatoma cells into the leg muscle of mice. Recombinant endostatin was produced by gene engineering in E. coli. The recombinant protein was injected subcutaneously to treat transplanted hepatoma faraway. The weight of tumors was measured, and the changes of necrosis of tumor cells and vessel density were observed by immunohistochemistry. The results suggested that the growth of hepatoma models transplanted in the muscle of legs was suppressed by recombinant endostatin. The density of vascularity was decreased, but the necrosis of tumor cells increased. The inhibitory effect of recombinant endostatin on angiogenesis and tumor growth of hepatoma was not affected after chemotherapy.
Angiogenesis Inhibitors ; pharmacology ; Animals ; Antineoplastic Agents ; pharmacology ; Endostatins ; biosynthesis ; genetics ; pharmacology ; Escherichia coli ; genetics ; Liver Neoplasms, Experimental ; blood supply ; pathology ; Male ; Mice ; Neoplasm Transplantation ; Recombinant Proteins ; biosynthesis ; genetics ; pharmacology

OBJECTIVETo investigate the feasibility of reduction in tumor antigen peptide dose by dendritic cell (DC)-presenting so as to elucidate the characteristics of modifying DC by heat shock protein (Hsp70) and antigen peptide.

METHODSAntigen peptide bound to Hsp70 was used to modify DC in vitro. The metabolism of the modified DC and the cytokine secreted thereby was determined. Then the activation of lymphocytes by the modified DC and Hsp70-H22 peptide was tested. The cytotoxicity of the activated lymphocytes to H22 tumor cells and the inhibition of tumor in mice by DC injection and Hsp70-H22 peptide was tested.

RESULTS0.15 micro g of H22 peptide bound to Hsp70 could mature 2 x 10(5) DC. 4 x 10(3) matured DC could activate 2 x 10(6) lymphocytes. The same amount of lymphocyte could be activated to produce similar cytotoxicity to tumor cells by either DC modified by 0.003 micro g of peptides bound with Hsp70 or by direct stimulation with 0.15 micro g of peptides bound to Hsp70. The dose of peptide could be reduced to 1/50 if the modified DC injection was used instead of direct Hsp70-peptide injection. Peptide from the normal hepatocytes, if bound to Hsp70, could not mature DC, nor could it activate lymphocytes through DC.

CONCLUSIONThe dose of Hsp70-H22 peptides can be reduced significantly by DC-presenting to activate lymphocytes. Peptides from normal cells, being unable to activate the lymphocytes by either Hsp70-presenting or DC-presenting, have little to offer in the induction of autoimmunity.

Animals ; Antigens, Neoplasm ; immunology ; Dendritic Cells ; immunology ; Disease Models, Animal ; HSP70 Heat-Shock Proteins ; chemistry ; immunology ; therapeutic use ; Immunity ; Lymphocyte Activation ; Mice ; Mice, Inbred BALB C ; Neoplasm Transplantation ; Neoplasms, Experimental ; prevention & control ; Peptides ; chemistry ; immunology ; therapeutic use ; Tumor Cells, Cultured

To investigate the inducement of cytotoxic T lymphocytes (CTLs) by antigen peptides mixture from different leukemia cells and the cross-reaction of the mixtures from different cell lines, antigen peptides mixtures were prepared from different leukemia cell lines respectively and then bound with Hsp70 in vitro. Activation and proliferation of PBMC were observed after stimulation with different Hsp70-peptide complexes. The ratio of CD8+ in proliferative cells was analyzed by flow cytometry. The cytotoxicity of the activated PBMC to different target cells was assayed. The results showed that the antigen peptides from different leukemia cell lines, bound with Hsp70, could activate PBMC effectively, and stimulate the activated PBMC to proliferate. The proliferative PBMC had specific cytotoxicity to corresponding leukemia cells. CD8+ cells, accounting for a high proportion in proliferative cells, had a specific cytotoxicity to leukemia cells from which antigen peptides were prepared, suggesting that these CD8+ cells were CTLs specific to leukemia cells. CTLs activated by Hut78-peptides or Molt4-peptides had a significantly stronger cytotoxicity to Hut78 cells, Molt-1 cells and Jurkat cells than that of CTLs activated by HL-60-peptides (P < 0.05). And the cytotoxicity of CTLs activated by Hut78/Molt4-peptides to Jurkat cells was significantly stronger than that of CTLs activated by either Hut78-peptides or Molt4-peptides alone (P < 0.05). It is concluded that antigen peptides mixtures from leukemia cells can induce specific antitumor CTLs. There exists cross-reactivity among antigen peptides mixtures from different cell lines of the same type leukemia and more cross-reactive antigen peptides could be obtained from more cell lines, suggesting that antigen peptides mixture with broad antigenic spectrum could be prepared by using multiple leukemia cell lines.
Antigens, Neoplasm ; metabolism ; pharmacology ; Cell Division ; Cells, Cultured ; Cross Reactions ; Cytotoxicity, Immunologic ; immunology ; HL-60 Cells ; HSP70 Heat-Shock Proteins ; metabolism ; Humans ; K562 Cells ; Leukocytes, Mononuclear ; cytology ; immunology ; Neoplasm Proteins ; immunology ; Peptides ; pharmacology ; T-Lymphocytes, Cytotoxic ; immunology

OBJECTIVETo prepare tumor antigen peptides from HL-60 cells and to induce specific immune response.

METHODSHL-60 antigen peptides were obtained using techniques including freezing and thawing, heat precipitation and acid precipitation. The stimulating effect of the in vitro Hsp70 binding HL-60 peptides on PBMC and the proliferation of stimulated PBMC were observed by T cell activation test. The cytotoxicity of proliferated PBMC is detected by incubating HL-60 cells or K562 cells with PBMC respectively.

RESULTSThe obtained tumor antigen peptides were a peptides mixture. The mixed peptides could activate PBMC and cause PBMC proliferation in vitro after presented by Hsp70. The proliferated PBMC showed specific cytotoxicity to HL-60 cells but not to K562 cells.

CONCLUSIONThe method for preparing of human leukemia tumor antigen peptides used in this paper is simple and easy; the obtained antigen peptides can induce specific immune response in vitro.

Cell Division ; HL-60 Cells ; HSP70 Heat-Shock Proteins ; immunology ; Humans ; K562 Cells ; Killer Cells, Natural ; immunology ; Leukocytes, Mononuclear ; cytology ; immunology ; Neoplasm Proteins ; immunology ; Peptides ; immunology