Objective To establish the methods of isolating and culturing human ovarian endometriosis-derived microvascular endothelial cells (OEMEC).Methods The tissues of human endometriotic cyst of ovary were finely minced with scissors,then digested by collagenase Ⅰ,Ⅱ and trypsinethylene diamine tetraacetic acid (EDTA).The cells were purified by using centrifugation of 2000 r/min speed.OEMEC were identified by light microscope and transmission electron microscope observing CD34,FⅧ-Rag and Weibel-Palade in microvascular endothelial cells.Results The OEMEC grew as confluent monolayer like cobblestones under light microscope.CD34 and FⅧ-Rag were expressed strongly,and the percentages of CD34 and FⅧ-Rag positive cells were 91.4％ and 92.5％.Weibel-Palade bodies could be observed under transmission electron microscope.The time of cell doubling proliferation was 4.5 days.Conclusion The established system of isolating OEMEC would provide lab base for studying the mechanisms of angiogenesis in endometriosis lesions.

Objective To conduct an in vitro study of the growth of Pseudomonas aeruginosa,Staphylococcus aureus and Acin- etobacter spp.,and evaluate their response to various concentrations of tumor necrosis factor(TNF)-?,interleukin (IL)-1?, and IL-6.Methods To monitor the growth of bacteria incubated with the cytokines TNF?,IL-1?and IL-6 that were added to RPMI 1640 medium in various concentrations (10,50.100,500 pg,1 and 10 ng) at 2,4 to 6,8 and 16-18 h.The bacterial concentration was estimated when the mixtures of cytokines and specific neutralizing monoclonal antibodies (MoAbs) were in- cubated.Results We found that all three bacterial species showed concentration-dependent growth enhancement when incubated with one or more tested cytokines.Blockade by specific neutralizing cytokine significantly inhibited cytokine-induced growth. When compared with control,the 6 h growth response was maximal with IL-1?for Staphylococcus aureus and Acinetobacter spp.,and with IL-6 for Pseudomonas aeruginosa.Conclusions In this study we provide additional evidence for a newly de- scribed mechanism for bacterial proliferation in the presence of exaggerated and protracted inflammation.The effect that cyto kine-induced growth enhancement inhibited by specific neutralizing cytokine MoAbs may be useful for antimicrobial therapy.

Objective To prepare,purify and validate specific monoclonal antibodies(McAb) against fragment in C-terminal telopeptides of collagen type Ⅱ(EKGPDP)for clinical diagnostics and related research of osteoarthritis(OA).Methods 8 BALB/c mice were immunized with EKGPDP-KLH antigen complex.McAb against EKGPDP fragements were prepared by hybridoma technique.Immunoglobulin classes and subclasses were determined using an Immuno-Type~(TM)mouse McAb isotyping kit.Ascites were producted and McAb were purified by saturation ammonium sulfate(SAS)precipitation and protein G chromatography.Specificity and immunoactivity of antibodies were detected by indirect enzyme-linked imrnunosorbant assay(ELISA).Cartilage specimens were analyzed by immunohistochemical method.Results The hybridoma cells were obtained.10 IgG and 3 IgM single colonies were picked out by limiting dilution and ELISA kit.The titers of McAb in ascites were from 2.8?10~4 to 5.1?10~5.The McAb showed the characteristics of no cross reactions with KLH,BSA,cell culture supernatant,type Ⅰ,Ⅲ collagens and whole type Ⅱ collagen.The method of SAS could get better recovery,immunoactivity of the McAb than protein G chromatograply(t=25.26;P

OBJECTIVESTo investigate whether antisense human telomerase reverse transcriptase (hTERT) could inhibit the activity of telomerase and the proliferation of K562 cells.

METHODSThe antisense plasmid was constructed by reverse insertion of hTERT PCR product into plasmid pLNCX-neo. Then the constructed plasmid was introduced into K562 cells by liposomes-mediated DNA transfection. The inhibition effects of telomerase on the proliferation of K562 cells were analyzed by MTT and colony formation assay, the telomerase activity of K562 cells by TRAP-PCR ELISA methods.

RESULTSThe growth rate of antisense hTERT transfected K562 cells was significantly lower than those of the controls, and the colony formation capacity of the transfected cells decreased significantly (P < 0.01), the colony number is (100.33 +/- 7.57)/10(3) cells, (92.67 +/- 5.86)/10(3) cells and (50.33 +/- 6.11)/10(3) cells for control K562 cells, K562 neo cells and antisense hTERT transfected HL60 cells, respectively. The telomerase activity of antisense hTERT transfected K562 cells was significantly inhibited.

CONCLUSIONThe expression of an antisense sequence to the mRNA sequence of telomerase protein subunit can inhibit the activity of telomerase, slow the cell growth and inhibit the capacity of colony formation of K562 cells.

Cell Division ; drug effects ; Humans ; K562 Cells ; Plasmids ; genetics ; RNA, Antisense ; genetics ; pharmacology ; RNA, Messenger ; genetics ; Telomerase ; drug effects ; genetics ; metabolism ; Transfection

Viral hepatitis was the most common infectious disease in china. But the diagnosis and treatment were varied because the viral hepatitis patients were hospitalized in different kinds of hospital such as infectious disease hospital, general hospital and Chinese medical hospital. It was necessary to know clinical characters and information of viral hepatitis patients in different hospitals. The general information, subtype distribution, prognosis, complication, medication and relations of onset with solar term from 41 180 viral hepatitis patients based on HIS data were analyzed. It was found that the age of patients between 18 to 59 years old was most; most patients were males. The national basic medical insurance was the most type of payment. The outcome of viral hepatitis in the youth and female were better than that in the old and male. Acute hepatitis was easer to restore than chronic hepatitis. Liver cirrhosis and hepatocellular carcinoma were the two most complications. The peak of onset was during summer solstice, slight heat and great heat. The most common Chinese medicine was Diammonium glycyrrhizinate and the most common western medicine was reduced glutathione. The combination of D. glycyrrhizinate with reduced glutathione, polyene phosphatidylcholine and thymosin was the main pattern. But It was not knew if the combination of western and Chinese medicine was the most effective therapy to protect liver function. It was necessary to take deeply research of the relationship between the combination therapy and their effectiveness.
Adolescent ; Adult ; Aged ; Antiviral Agents ; therapeutic use ; Female ; Glutathione ; therapeutic use ; Glycyrrhizic Acid ; therapeutic use ; Hepatitis, Viral, Human ; drug therapy ; Hospitals ; Humans ; Male ; Middle Aged ; Young Adult

OBJECTIVE: To establish an inductively coupled plasma-mass spectrometry (ICP-MS) method for determination of 24 elements in human hair. METHODS: The samples were digested with microwave digestion instrument. ICP-MS was applied to determine 24 elements in human hair using indium (115In) as an internal standard. The established method was applied to determine element concentration in normal group (56 samples) and heroin abuse group (10 samples). RESULTS: The limits of detection ranged from 0.000 3 microg/g to 10.14 microg/g. Measured value of the standard materials were basically consistent with the standard value. The contents of magnesium, gallium and barium in hair of heroin addicts decreased after rehabilitation treatment. CONCLUSION The method is sensitive and accurate for determination of 24 elements in human hair.
Adolescent ; Adult ; Aged ; Barium/analysis* ; Child ; Child, Preschool ; Female ; Gallium/analysis* ; Hair/chemistry* ; Heroin Dependence/rehabilitation* ; Humans ; Magnesium/analysis* ; Male ; Mass Spectrometry/methods* ; Metals, Heavy/analysis* ; Microwaves ; Middle Aged ; Sensitivity and Specificity ; Spectrophotometry, Atomic ; Trace Elements/analysis* ; Young Adult

The myelodysplastic syndromes (MDS) include a diverse groups of clonal and potentially malignant bone marrow disorders. Evidences exist that microenvironment cells from MDS marrow show functional abnormalities, which may be relevant to the incidence of such a disease. Mesenchymal stem cells (MSCs) are a very important component of hematopoietic microenvironment. This study was supposed to investigate the biological characteristics and functions of MSC derived from patients with MDS in low-risk. MSCs from bone marrow samples of 11 low-risk MDS patients were isolated, cultured and expanded. Morphology, immunophenotype and osteoblasts differentiation were analyzed. Their capacity of proliferation and hematopoietic supporting in vitro were measured. A real-time quantitative reverse transcriptase polymerase chain reaction method (RQ RT-PCR) was used for detecting the expression levels of relative cytokines and chemokines in MSC. MSCs from healthy donors were used as controls. The results showed that the culture-expanded cells from MDS patients displayed a typical fibroblast-like morphology. Cells were positive for SH2 (CD105), SH3 (CD73), Thy-1 (CD90), while negative for CD34 and CD45. After induction, these cells could differentiate into osteoblasts. The proliferative ability of MSCs in MDS patients were not different from those of MSC isolated from normal bone marrow (p > 0.05), however, their capacity of hematopoietic supporting in vitro were significantly weaker (p < 0.05). RQ RT-PCR detection indicated that the SDF-1 gene expression level in MSCs of low-risk MDS patients was significantly higher than that in MSC derived from healthy donors (p < 0.01). It is concluded that the abnormal function of MSC influences the regulation of hemotopoiesis in the bone marrow microenvironment of MDS patients. It is worthy to further investigate the new clue in etiological mechanism and therapeutic strategies for MDS.
Bone Marrow Cells ; pathology ; Chemokine CXCL12 ; genetics ; metabolism ; Hematopoiesis ; Humans ; Mesenchymal Stromal Cells ; pathology ; physiology ; Myelodysplastic Syndromes ; pathology ; physiopathology ; Risk Factors

OBJECTIVETo study magnetic resonance enhancement features of inflammatory lymph nodes using different doses of ultrasmall superparamagnetic iron oxide (USPIO) particles in order to establish a standardized protocol for USPIO enhanced magnetic resonance imaging of lymph nodes.

METHODSA total of 12 healthy New Zealand rabbits were injected complete Freund's adjuvant in foot pad to establish popliteal inflammatory lymph node model. Different doses (45, 90, 135 micromol Fe/kg) of USPIO were injected intravenously. Magnetic resonance scans were performed before and after USPIO injection to observe the enhancement features of different groups. T2 signal intensity, T1 signal intensity, T2 x value, and T2 value were measured and T2 enhancement ratio was calculated at different time points.

RESULTSTwenty-four hours after USPIO injection, there was no statistical difference in T2 signal intensity and T2 enhancement ratio between 90 and 135 micromol Fe/kg dose groups, but both were superior to 45 micromol Fe/kg group (P < 0.05). There were no statistical differences in T2 signal intensity, T1 signal intensity, T2 value, and T2 enhance ratio among different postcontrast time delays from 6 to 24 hours in 90 micromol Fe/kg group (P > 0.05), and signal reduction of lymph nodes peaked 18 hours after USPIO injection. Better images were acquired with a postcontrast delay of 18-24 hours.

CONCLUSIONSLymph nodes can be enhanced well with a dose of 90 micromol Fe/kg. Postcontrast delay of 18-24 hours is appropriate for acquiring satisfactory enhancement images.

Animals ; Contrast Media ; administration & dosage ; Dextrans ; administration & dosage ; Image Enhancement ; methods ; Lymphadenitis ; diagnosis ; pathology ; Magnetic Resonance Imaging ; methods ; Magnetite Nanoparticles ; administration & dosage ; Male ; Rabbits ; Random Allocation

OBJECTIVETo assess the characteristics of enhanced magnetic resonance image with ultrasmall superparamagnetic iron oxide (USPIO) in the inflammatory and tumor metastatic rabbit model, and explore its relevance with histologic ultrastructural findings.

METHODSTotally 36 New Zealand white rabbits were randomly divided into lymphadenitis group and metastatic group. Complete Freund's adjuvant was injected into the bilateral dorsal footpads of 18 rabbits to set up ipsilateral lymphadenitis model. The other 18 rabbits received a subcutaneous implantation of VX2 tumor cell suspension (1.5 x 10(7) cells/ml) in both thighs to set up metastatic lymph node model. Magnetic resonance scan were performed 24 hours before and after USPIO (90 micromol Fe/kg) injection. T2 values of each lymph node were measured and lymph node T2 enhancement rate was calculated as well. HE staining, Prussian blue staining, and electronic microscopy were performed to observe the pathological microstructure changes and the distribution of the iron particle in lymph node. Relationship between lymph nodes USPIO enhancement and its microstructures were further analyzed. Results Thirty-six lymph nodes in lymphadenitis group showed different degrees of reactive hyperplasia. Twenty-six lymph nodes in metastatic group were invaded by tumor cell. Non-enhanced scan showed mild difference between T2 signal intensity of the two pathological lymph node types. After USPIO enhancement, inflammatory lymph nodes showed distinct T2 signal reduction at the center, and metastatic lymph nodes showed homogenous and faint T2 signal reduction. Enhancement rate of benign and malignant lymph nodes were 57.39% and 29.45% respectively (P < 0.01). HE staining and Prussian blue staining indicated USPIO particles located mainly in the macrophages at inflammatory lymphatic medulla, while paracortical area and cortical area contained relatively much less USPIO particles due to less macrophages distribution. MRI findings were correlated with the pathological results. Electronic microscopy also verified that the majority of USPIO particles were located in the numerous cytophagic bubbles of macrophages. Lymph nodes metastasis including 4 lymph nodes with completed structure destruction due to entire tumor infiltration, 19 lymph nodes with partially lymph node structure destruction but reduced USPIO-contained macrophage numbers or reduced USPIO particles in macrophages, and 3 lymph nodes with only localized foci tumor metastasis at subcapsular area. Conclusions USPIO enhancement pattern of different lymph nodes is closely related to distribution and functional status of the intra-node macrophages. It may affect the accuracy of the lymph node property diagnosis based on USPIO enhanced image.

Animals ; Dextrans ; metabolism ; Female ; Image Enhancement ; methods ; Lymph Nodes ; ultrastructure ; Lymphadenitis ; diagnosis ; pathology ; Lymphatic Metastasis ; diagnosis ; ultrastructure ; Magnetic Resonance Imaging ; methods ; Magnetics ; Magnetite Nanoparticles ; Male ; Nanoparticles ; Rabbits ; Random Allocation

AIMTo study the effects of TNF receptor blocking peptide on adjuvant arthritis in rats.

METHODSThe model of rat adjuvant arthritis was induced by injection of complete Freund's adjuvant. The TNF receptor blocking peptide was injected locally in the ankle. The ankle swelling, the pathologic changes in the ankle joint and the expression of IL-1 beta mRNA and TNF-alpha mRNA by peritoneal macrophages (RT-PCR) were observed.

RESULTSThe model of rat adjuvant arthritis induced by injection of complete Freund's adjuvant was similar to human rheumatoid arthritis. The treatment with TNF receptor blocking peptide for 10 days resulted in complete inhibition of joint swelling, a decrease in infiltration of inflammatory cell into joint tissue, an obvious alleviation of inflammatory pathological damages and an apparent decline of TNF-alpha mRNA and IL-1 beta mRNA of peritoneal macrophages of rats.

CONCLUSIONThe TNF receptor blocking peptide can protect the joint from inflammatory damage induced by adjuvant arthritis by suppression of TNF-alpha and IL-1 production, thereby alleviating the pathological injury of joint and controlling effectively the clinic course of arthritis.

Animals ; Ankle Joint ; pathology ; Arthritis, Experimental ; immunology ; pathology ; Interleukin-1 ; biosynthesis ; genetics ; Macrophages, Peritoneal ; metabolism ; Male ; Peptides ; pharmacology ; RNA, Messenger ; genetics ; Rats ; Rats, Wistar ; Receptors, Tumor Necrosis Factor ; antagonists & inhibitors ; Tumor Necrosis Factor-alpha ; biosynthesis ; genetics