To understand the sero-epidemiological features of rickettesiosis of humans and domestic animals in Yunnan province, blood samples from 237 adults in different geographic area, including Xundian country,Yulong country and Simao country and 81 children aged from 4 to 6 years old were collected for serological testing. In addition, 90 blood samples from dogs, goats and ox in each investigated area were also collected. Antibodies against 8 rickettsiae, including R.typhi, R.heilongjiangii or R.sibirica, Orientia tsutsugamushi Karp or Kato, Ehrlichia chaffeensis, Anaplasma phagocytosis, Bartonella henselae, Coxiella burnetii and Hainan spotted fever group of rickettsia were examined by using immunofluorescence assays(IFA).It was found that the sero-epidemiologic rates of R.typhi, B.henselae and C.burnetii (16.46%、6.33% and 9.28%) of adults were higher than those of other rickerrsiae investigated. The positive rates of IgG antibody against R.typhi for children also shared the higher rate (12.35%). Similar sero-epidemiologic features were found for domestic animals. Among the 8 rickettsia tested in this study, the positive rate of IgG antibody against R.typhi appeared to be the highest(61.48%) without significant difference among these investigated sites. From this investigation, it is evident that the rickettsial infection of farm population and domestic animals are common in Yunnan province, and the active surveillance of rickettsiosis and differential diagnosis of unknown febrile patients in clinical practice should be enforced.

BACKGROUNDThe amiloride-sensitive epithelial sodium channel α-subunit (α-ENaC) is an important factor for alveolar fluid clearance during acute lung injury. The relationship between adenosine receptor A(2a) (A(2a)AR) expressed in alveolar epithelial cells and α-ENaC is poorly understood. We targeted the A(2a)AR in this study to investigate its role in the expression of α-ENaC and in acute lung injury.

METHODSA549 cells were incubated with different concentrations of A(2a)AR agonist CGS-21680 and with 100 µmol/L CGS-21680 for various times. Rats were treated with lipopolysaccharide (LPS) after CGS-21680 was injected. Animals were sacrificed and tissue was harvested for evaluation of lung injury by analysis of the lung wet-to-dry weight ratio, lung permeability and myeloperoxidase activity. RT-PCR and Western blotting were used to determine the mRNA and protein expression levels of α-ENaC in A549 cells and alveolar type II epithelial cells.

RESULTSBoth mRNA and protein levels of α-ENaC were markedly higher from 4 hours to 24 hours after exposure to 100 µmol/L CGS-21680. There were significant changes from 0.1 µmol/L to 100 µmol/L CGS-21680, with a positive correlation between increased concentrations of CGS-21680 and expression of α-ENaC. Treatment with CGS-21680 during LPS induced lung injury protected the lung and promoted α-ENaC expression in the alveolar epithelial cells.

CONCLUSIONActivation of A(2a)AR has a protective effect during the lung injury, which may be beneficial to the prognosis of acute lung injury.

Acute Lung Injury ; metabolism ; Adenosine ; analogs & derivatives ; pharmacology ; Animals ; Blotting, Western ; Cell Line ; Epithelial Sodium Channels ; genetics ; metabolism ; Humans ; Male ; Phenethylamines ; pharmacology ; Pulmonary Alveoli ; cytology ; metabolism ; Purinergic P1 Receptor Agonists ; pharmacology ; Rats ; Receptors, Purinergic P1 ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo construct the multi-probe ribonuclease protection assay (RPA) template set to be used for detecting expression patterns of MD-2, TLR4, CD14 mRNAs in human peripheral blood mononuclear cells.

METHODSThe designed cDNA fragments of the three genes were generated by polymerase chain reaction (PCR) using specific primers and directionally cloned into EcoR I and Hind III sites of expression plasmid pSP72 containing the T7 promoter, the linearized plasmids was used as template to synthesize anti-sense RNA probes. Then we extracted total RNA from peripheral blood mononuclear cells (PBMC) and detected the dynamic expression patterns of the three genes with RPA method.

RESULTSThe proper sequence and orientation of the template set were confirmed by sequencing and the template set was successfully used to assay TLR4, MD-2 and CD14 mRNAs in human PBMC. The results showed that the three detected genes decreased transiently 1-3 hours after 100 ng/ml LPS stimulation.

CONCLUSIONSThese new RPA multi-probe set provided valuable tool for the simultaneous quantitative determination of expression of TLR4, CD14 and MD-2 mRNAs in both constitutive and inducible types.

Antigens, Surface ; analysis ; Base Sequence ; Biological Assay ; Cells, Cultured ; DNA ; analysis ; genetics ; Gene Expression Profiling ; methods ; Humans ; Lipopolysaccharide Receptors ; analysis ; Lymphocyte Antigen 96 ; Membrane Glycoproteins ; analysis ; Molecular Probe Techniques ; Molecular Sequence Data ; Monocytes ; metabolism ; RNA Probes ; analysis ; genetics ; Receptors, Cell Surface ; analysis ; Receptors, Immunologic ; analysis ; Ribonucleases ; Toll-Like Receptor 4 ; Toll-Like Receptors

OBJECTIVETo explore the association of the expression of hypoxia-inducible factor 1α (HIF-1α) with microlymphatic vessel density(MLVD) and lymph node micro-metastasis in rectal cancer.

METHODSThe experimental group consisted of 40 middle-low rectal cancer specimens pathologically confirmed at the First Affiliated Hospital of Anhui Medical University between 2000 and 2003. Forty samples of normal tissues taken from the corresponding area around the cancer were used as the control group. Immunohistochemistry was used to detect HIF-1α expression and MLVD in both the tumor tissues and the adjacent normal tissues. Lymph node micrometastasis was ascertained using immunohistochemical staining with CK20.

RESULTSIn rectal cancer tissues, the HIF-1α expression was 77 386±14 911 and MLVD was 7.3±0.7, significantly higher than those in normal adjacent tissues(33 092±5877 and 0.3±0.2, both P<0.01). The HIF-1α expression was positively correlated with MLVD in rectal cancer(r=0.781, P<0.01). Thirty-one patients had no lymph nodes metastasis and 10 had micrometastasis. The HIF-1α expression and MLVD in specimens with lymph node micrometastasis was significantly higher than that in those without lymph node micrometastasis(P<0.05).

CONCLUSIONHIF-1α and MLVD play important roles in the development of rectal cancer,which may promote lymphatic micrometastasis in rectal cancer.

Adult ; Aged ; Aged, 80 and over ; Female ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Lymph Nodes ; pathology ; Lymphatic Metastasis ; Lymphatic Vessels ; pathology ; Male ; Middle Aged ; Neoplasm Micrometastasis ; Rectal Neoplasms ; metabolism ; pathology

To evaluate the significance of FCM in minimal residual disease (MRD) detection, the immunophenotyping and leukemia-associated immunophenotypes (LAIP) of leukemia cells from 273 adult and 142 childhood patients with B lineage acute lymphoblastic leukemia (B-ALL) were detected by four to six antibody combinations of 4-color CD45/SSC gating multiparametric flow cytometry (FCM). The results showed that the B-ALL patients could be classified into 4 subtypes based on different expression CD34 and CD10: subtype I (CD34(+)/CD10(-)), subtype II (CD34(+)/CD10(+)), subtype III (CD34(-)/CD10(+)), subtype IV (CD34(-)/CD10(-)). The LAIP was observed in 100% and 92% patients of subtype I and subtype II, respectively, whereas only 79.2% in subtype III. The incidence of LAIP in total B-ALL cases was 90% by using the antibodies detected in this investigation. There was no significantce different for incidence of LAIP between adult and pediatric patients. LAIP was observed in 77.6% of patients by labeling only CD34/CD10/CD19/CD45 4-color antibody combination. It is concluded that in 90% of childhood and adult B-ALL patients LAIP can be found, which suits MRD detection by multiparameter flow cytometry.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Antigens, CD34 ; analysis ; B-Lymphocytes ; immunology ; Burkitt Lymphoma ; classification ; immunology ; pathology ; Cell Lineage ; Female ; Flow Cytometry ; methods ; Humans ; Immunophenotyping ; Male ; Middle Aged ; Neoplasm, Residual ; diagnosis ; Neprilysin ; analysis ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; classification ; immunology ; pathology

There are certain limitations in the application of liver transplantation， resection and radiofrequency ablation for liver cancer. Therefore，specific and selective new drugs are needed to provide better treatment. Curcumin is a hydrophobic polyphenol with a wide range of activities，such as anti-inflammatory，antibacterial，anti-oxidant and anti-tumor properties. Its nano-preparation has stronger growth inhibition and pro-apoptosis effects on tumor cells. Literature retrieval found that curcumin's anti-liver cancer molecular mechanisms include inhibiting cell proliferation by regulating the expressions of relevant miR，glyoxalase 1（GLO1），CD133 and vascular endothelial growth factor （VEGF），inhibiting signal transducer and activator of transcription （STAT3） and YAP expression to induce cell apoptosis，regulating the heat shock protein 70 （HSP70）-Toll-like receptor 4 （TLR4） signaling pathway，Wnt/β-catenin and transforming growth factor（TGF）/epithelial-mesenchymal transition（EMT） pathways，nuclear factor-κB （NF-κB） signaling pathway and nuclear factor E₂-related factor 2（Nrf2）/ Kelch-like ECH-related protein 1（Keap1） signaling pathway，inhibiting p38 mitogen-activated protein kinase （MAPK） phosphorylation，to reduce Lin28B expression，regulating phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt） signaling pathway and inhibiting G protein-coupled receptors （GPR81）/hydroxycarboxylic acid receptor-1 （HCAR-1） expression to reverse transformation therapy resistance. Curcumin nano-preparation mainly includes polymer micelles，liposomes，loaded microbubbles，nanocapsules and nanoparticles. Curcumin is mainly delivered to liver cancer cells to rapidly release the drug，enhance the anti-liver cancer effect and reduce toxic and side effects in normal liver cells. The mechanisms include activation of DR5/Caspase-mediated exogenous apoptosis pathway and VEGF/VEGF receptors （VEGFRs） signaling pathway，loss of mitochondrial membrane potential and increase of intracellular reactive oxygen species （ROS）. This paper summarizes the molecular mechanism of curcumin and its nano-preparation against liver cancer，in order to further define the molecular mechanism of liver cancer and provide a new reference for its clinical diagnosis and treatment.

OBJECTIVETo compare the leukemia-associated immunophenotypes (LAIP) in patients with B lineage acute lymphoblastic leukemia (B-ALL) at diagnosis and relapse, and investigate its implications for minimal residual disease (MRD) detection.

METHODSThe immunophenotype of leukemia cells from 410 newly diagnosed and 6 relapsed patients with B-ALL were detected by four to six antibody combination, mainly CD34/CD10/CD45/CD19 of 4-color CD45/SSC gating flow cytometry (FCM).

RESULTSThe proportion of CD45 under-expressed or negative in relapsed patients was much higher than that in newly diagnosed patients, being 69.2% and 37.8% respectively. Immunophenotypic changes occurred in 9 relapsed patients (including 8 hematological relapse and 1 central nerves system relapse) when analyzed by paired samples analysis at diagnosis vs at relapse: 4 cases showed CD45 down-modulation and 2 up-modulation; 4 CD34 down-modulation and 2 CD10 up-modulation, while the expression of CD19 remained no change. MRD was observed in all 7 cases of hematological relapse 2 - 4 months before relapse, and the immunophenotype of MRD cells was the same as that in relapse.

CONCLUSIONA high frequency of immunophenotypic changes occurred at relapse and even in MRD before relapse, however the accuracy of MRD monitoring seemed not affected by the FCM strategy used in this investigation.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Antigens, CD19 ; immunology ; Antigens, CD34 ; immunology ; Child ; Child, Preschool ; Female ; Flow Cytometry ; methods ; Humans ; Immunophenotyping ; methods ; Leukocyte Common Antigens ; immunology ; Male ; Middle Aged ; Neoplasm, Residual ; diagnosis ; immunology ; pathology ; Neprilysin ; immunology ; Precursor B-Cell Lymphoblastic Leukemia-Lymphoma ; diagnosis ; immunology ; pathology ; Recurrence ; Young Adult

OBJECTIVETo evaluate the clinical significance for minimal residual disease (MRD) detection by 4 color flow cytometry in B lineage acute lymphoblastic leukemia (B-ALL).

METHODSMRD was analyzed and followed up by using two panels of 4 color antibodies, mainly CD34/CD10/CD45/CD19, in 671 consecutive bone marrow specimens and 1 cerebrospinal fluid from 98 B-ALL patients. In 26 cases of them the immunophenotyping informations at diagnosis were not available.

RESULTSOf 671 bone marrow samples, 579 were MRD negative with leukemic cells below 0.0001 and 93 were MRD positive with leukemic cells over 0.0001. Of 93 MRD positive samples, leukemic cells below 0.05 were found in 64 bone marrow samples, meanwhile in the other 29 samples leukemic cells were over 0.05. Twenty patients relapsed, 19 were bone marrow relapse and one center nerves system. Fifteen of them were found MRD positive 7 - 17 weeks before relapse including 6 patients having no immunophenotyping data at diagnosis. The percentages of leukemia cells in these 15 patients were all over 0.0001. Two relapsed patients were MRD negative in 3 and 9 months before relapse, respectively. Two relapsed after MRD monitoring stopped. If MRD level was > 0.0001 at the end of induction chemotherapy and 12 weeks of treatment, the rate of relapse was 50% (6/12), while, it was 7.5% (3/40) in MRD negative patients (P = 0.000).

CONCLUSIONRelapses can be predicted by MRD monitoring, if MRD was positive in the early phase of treatment, the risk of relapse was higher. Based on the characteristics of B cells ontogeny, MRD detection can be done independently of immunophenotypic information at diagnosis.

Acute Disease ; Adolescent ; Adult ; Aged ; Antigens, CD19 ; immunology ; Antigens, CD34 ; immunology ; Child ; Child, Preschool ; Female ; Flow Cytometry ; methods ; Follow-Up Studies ; Humans ; Immunophenotyping ; Leukocyte Common Antigens ; immunology ; Male ; Middle Aged ; Neoplasm, Residual ; diagnosis ; immunology ; Neprilysin ; immunology ; Precursor B-Cell Lymphoblastic Leukemia-Lymphoma ; diagnosis ; immunology ; Young Adult

OBJECTIVETo investigate the correlation of vascular endothelial growth factor D (VEGF-D) expression, microlymphatic density (MLD) and microvessel density (MVD) levels with the development and metastasis of rectal cancer.

METHODSEighty specimens from resected middle-lower rectal cancer diagnosed by pathology were examined by immunohistochemistry for VEGF-D,MLD and MVD. Simultaneously, 40 biopsy specimens from rectal polyps and 80 specimens from normal rectal tissue were examined as controls. Correlation between the expression of above three factors and the tumor size, gross morphology, histological type, metastasis, differentiation grade, infiltration depth, Dukes stage, lymph node metastasis and long-distance metastasis before operation were investigated with Spearman method.

RESULTS(1) Positive expression rate of VEGF-D was 55 % (44/80) in rectal cancer, and zero in rectal polyps and normal rectal tissues. The expression of VEGF-D in rectal cancer was significantly higher than that in rectal polyps and normal rectal tissues(P<0.05). MLD was significantly higher in rectal cancer (2.80+/-1.31) than that in rectal polyps (0.50+/-0.72) and normal rectal tissues(0.25+/-0.44)(P<0.05).Meanwhile MVD was significantly higher in rectal cancer (80.10+/-23.18) than that in rectal polyps (27.00+/-11.01) and normal rectal tissues (10.45+/-5.34) (P<0.05). (2) VEGF-D, MLD and MVD were positively correlated with lymph node metastasis and long-distance metastasis before operation (P<0.05). (3) VEGF-D was positively correlated with MLD (P<0.05) and MLD was positively correlated with MVD as well(P<0.05).

CONCLUSIONSLymphangiogenesis exists in rectal cancer tissues. VEGF-D and MLD can be used as good predictors of lymphangiogenesis and they are the important factors affecting biological behavior of rectal cancer. Lymphangiogenesis and angiogenesis may have a cooperative function in the development of rectal cancer.

Female ; Humans ; Lymphangiogenesis ; Lymphatic Metastasis ; Male ; Microcirculation ; Microvessels ; pathology ; Middle Aged ; Neoplasm Staging ; Neovascularization, Pathologic ; pathology ; Rectal Neoplasms ; blood supply ; pathology ; Vascular Endothelial Growth Factor D ; genetics ; metabolism

BACKGROUNDDamaged articular cartilage has very limited capacity for spontaneous healing. Tissue engineering provides a new hope for functional cartilage repair. Creation of an appropriate cell carrier is one of the critical steps for successful tissue engineering. With the supposition that a biomimetic construct might promise to generate better effects, we developed a novel composite scaffold and investigated its potential for cartilage tissue engineering.

METHODSChitosan of 88% deacetylation was prepared via a modified base reaction procedure. A freeze-drying process was employed to fabricate a three-dimensional composite scaffold consisting of chitosan and type II collagen. The scaffold was treated with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide. Ultrastructure and tensile strength of the matrix were carried out to assess its physico-chemical properties. After subcutaneous implantation in rabbits, its in vivo biocompatibility and degradability of the scaffold were determined. Its capacity to sustain chondrocyte growth and biosynthesis was evaluated through cell-scaffold co-culture in vitro.

RESULTSThe fabricated composite matrix was porous and sponge-like with interconnected pores measuring from 100-250 microm in diameter. After cross-linking, the scaffold displayed enhanced tensile strength. Subcutaneous implantation results indicated the composite matrix was biocompatible and biodegradable. In intro cell-scaffold culture showed the scaffold sustained chondrocyte proliferation and differentiation, and maintained the spheric chondrocytic phenotype. As indicated by immunohistochemical staining, the chondrocytes synthesized type II collagen.

CONCLUSIONSChitosan and type II collagen can be well blended and developed into a porous 3-D biomimetic matrix. Results of physico-chemical and biological tests suggest the composite matrix satisfies the constraints specified for a tissue-engineered construct and may be used as a chondrocyte carrier for cartilage tissue engineering.

Animals ; Biodegradation, Environmental ; Cartilage ; cytology ; Chitosan ; chemistry ; Coculture Techniques ; Collagen Type II ; chemistry ; Immunohistochemistry ; Rabbits ; Tensile Strength ; Tissue Engineering ; methods