OBJECTIVETo investigate the relationship between the expressions of KAI1, nm23, ETS-1, vascular endothelial growth factor (VEGF) and microvascular density (MVD) and lymph node metastasis and prognosis in nasopharyngeal carcinoma (NPC).

METHODSThe Envision immunohistochemical method was used to detect the expressions of KAI1, nm23, ETS-1 and VEGF in 50 cases of non-keratinizing carcinoma (NKC) with cervical lymph node metastasis, 30 cases of NKC without cervical lymph node metastasis at the primary diagnoses and 30 cases of non-tumor nasopharyngeal tissues (NP). The microvascular density was counted by immunostaining with CD34.

RESULTS(1) The expression rates of KAI1 and nm23 protein in NKC with cervical lymph node metastasis group and without cervical lymph node metastasis group and NP group increased successively , the difference being significant (P < 0.05); The expression rates of ETS-1 and VEGF protein in NKC with cervical lymph node metastasis group and without cervical lymph node metastasis group and NP group increased successively, the difference being significant (P < 0.05). (2) In 80 NKC cases, the MVD was respectively lower in KAI1 and nm23 protein positive groups than those in the negative groups (P < 0.05); the MVD was respectively higher in ETS-1 and VEGF protein positive groups than those in the negative groups (P < 0.05 ). (3) There was significant difference between the MVD, the number of NKC without cervical lymph node metastasis cases in the single expression of KAI1 or nm23 protein and in common expression of KAI1 and nm23 protein (P < 0.05), in the same as between the single expression of ETS-1 or VEGF protein and in common expressions of ETS-1 and VEGF protein (P < 0.05). (4) There was positive correlation between the expressions of KAI1 and nm23 protein (P < 0.01), as well as between the expressions of ETS-1 and VEGF protein (P < 0.01). (5) the 5-year survival rates of the patients correlated with cervical lymph node metastasis and the expressions of KAI1, nm23, ETS-1 and VEGF proteins in NKC (P < 0.05).

CONCLUSIONSThe expressions of KAI1, nm23, ETS-1 and VEGF proteins were highly related to MVD in NPC,cervical lymph node metastasis and prognosis. They might be considered to be reference indicator for evaluating the cervical lymph node metastasis and prognosis of NPC.

Adult ; Aged ; Female ; Humans ; Kangai-1 Protein ; metabolism ; Lymphatic Metastasis ; Male ; Middle Aged ; NM23 Nucleoside Diphosphate Kinases ; metabolism ; Nasopharyngeal Neoplasms ; blood supply ; metabolism ; pathology ; Prognosis ; Proto-Oncogene Protein c-ets-1 ; metabolism ; Vascular Endothelial Growth Factor A ; metabolism ; Young Adult

Aedes ; virology ; Animals ; Dengue Virus ; classification ; isolation & purification ; Genome, Viral ; Phylogeny ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo explore the expression of MMP20 in endometrial carcinoma and its clinical significance.

METHODSBioinformatics analysis was used to explore the correlation of MMP20 expression with the prognosis of the patients with endometrial carcinoma. We collected 21 pairs of fresh endometrial carcinoma and adjacent endometrial tissues to detect the expression of MMP20 mRNA using real-time PCR. We also examined MMP20 protein expressions in 134 paraffin-embedded endometrial carcinoma tissues and 34 paraffin-embedded endometrial tissues using immunohistochemistry. The correlation of MMP20 expression with the clinicopathological features and prognosis of the patients were analyzed.

RESULTSBased on TCGA database analysis, the patients with endometrial carcinoma showing an increased MMP20 mRNA expression had a significantly poorer prognosis than those with a low MMP20 mRNA expression (=0.00065). Real-time PCR analysis and immunohistochemistry confirmed the up-regulated expressions of MMP20 at both the mRNA (=0.0062) and protein (=0.005) levels in endometrial carcinoma tissues compared to the adjacent endometrial tissues. An elevated MMP20 protein expression was positively associated with FIGO stage of endometrial carcinoma (=0.014) and inversely correlated with the survival time of the patients (=0.019). The Cox regression model analysis showed that an increased MMP20 expression was an independent predictor of a poor prognosis of the patients with endometrial carcinoma (=0.067).

CONCLUSIONSHigh expression of MMP20 is a potential prognostic factor for poor outcomes of patients with endometrial carcinoma.

OBJECTIVETo study the susceptibility of Aedes albopictus to dengue virus infection.

METHODSAedes albopictus from 15 collections in Guizhou province were challenged orally with dengue virus 1-4 types, respectively. The total RNA from mosquitos were extracted. The viral NS1 gene fragment was amplified with reverse transcriptase polymerase chain reaction (RT-PCR). Dengue virus in mosquitoes was isolated with C6/36 cells. Then the viral antigen was detected by indirect immunofluorescence assay (IFA). Antigen and nucleic acid of dengue virus from 15 geographic strains of Aedes albopictus orally infected with dengue viruses (DEN-1, DEN-2, DEN-3 and DEN-4) were detected by IFA, RT-PCR and the virus was isolated with C6/36 cells, respectively.

RESULTSThe rates of Aedes albopictus orally infected with DEN-1, DEN-2, DEN-3 and DEN-4 were 12/15, 12/15, 8/15 and 13/15, respectively.

CONCLUSIONSDifferent geographic strains of Aedes albopitus in Guizhou were susceptible to dengue viruses.

Aedes ; cytology ; virology ; Animals ; Antigens, Viral ; analysis ; Cell Line ; China ; DNA, Viral ; analysis ; Dengue Virus ; genetics ; isolation & purification ; Disease Susceptibility ; Ecosystem ; Reverse Transcriptase Polymerase Chain Reaction ; Viral Nonstructural Proteins ; genetics

OBJECTIVETo assess the efficacy and safety of Tangzu Yuyang Ointment (TYO) for treatment of chronic diabetic foot ulcers.

METHODSFifty-seven patients with chronic diabetic foot ulcers of Wagner's ulcer grade 1 to 3 were randomly assigned to the control group (29 cases) and the treatment group (28 cases). Patients in the control group received the standard wound therapy (SWT), while those in the treatment group received SWT plus TYO. The ulcer healing rate, the ulcer improvement rate and the incidence of adverse events were compared between the two groups. Totally 48 patients finished the final follow-ups and entered the data analysis.

RESULTSThe ulcer improvement rate was 79.2% in the TYO group and 41.7% in the SWT group (P=0.017) at the 12th week, and 91.7% vs. 62.5% (P=0.036) at the 24th week. There was no statistical difference in the ulcer healing rate and the incidence of adverse events between the two groups at week 4, 12, and 24, respectively. The ulcer healing time was 96 +/- 56 days in the TYO group and 75 +/- 53 days in the SWT group, showing insignificant difference (P=0.271).

CONCLUSIONTYO plus SWT was more safe and effective than SWT alone in the treatment of chronic diabetic foot ulcers.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Diabetic Foot ; drug therapy ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Humans ; Male ; Middle Aged ; Ointments ; Phytotherapy ; Treatment Outcome ; Wound Healing ; Young Adult

ObjectiveTo explore the potential suitable distribution area and spatial production area of high-quality medicinal materials of Miao medicine Laportea bulbifera. MethodBased on 200 distribution spots of L. bulbifera and 120 ecological factors, the maximum entropy model（MaxEnt） and ArcGIS software were applied to predict the potential suitable areas. The content of rutin, kaempferol-3-O-rutinoside, and quercitrin from 33 batches of L. bulbifera in Guizhou province was determined by high-performance liquid chromatography (HPLC). Combined with the previous content data of total polysaccharides and total flavonoids, the correlation between the main pharmacodynamic components of L. bulbifera and ecological factors was analyzed. Eventually, the quality regionalization of L. bulbifera was exhibited by the collocated Cokriging in ArcGIS. ResultThe optimum distribution areas of L. bulbifera were Yunnan, Guizhou, Sichuan, Hunan, and Hubei provinces in China, and the high and medium suitable areas accounted for 59.48% of the total distribution areas. October precipitation, slope, min temperature of the coldest month, altitude, solar radiation intensity in June, and April precipitation were the main ecological factors affecting the growth and distribution of L. bulbifera. Temperature, radiation intensity, precipitation, and soil composition and properties had great effects on the accumulation of secondary metabolites in L. bulbifera. The results showed that the high-quality areas of L. bulbifera mainly included Leishan county, Kaili city, Qingzhen city, Pingba district, Huishui county, Longli county, Kaiyang county, and Jiangkou county in Guizhou province. ConclusionL. bulbifera is widely distributed in China， but the suitable distribution areas are mainly concentrated in southwest China and part of central China. The planting bases of L. bulbifera can be selected in parts of cities and counties in southeast and central Guizhou province. In this study， the potential suitable growing areas and the optimum planting areas of L. bulbifera in Guizhou were predicted， which could provide scientific references for the cultivation and the site selection of large-scale planting for L. bulbifera.

ObjectiveTo investigate the intervention effect of total glucosides of paeony （TGP） on the renal injury of MRL/lpr mice based on the Toll-like receptor 9 （TLR9）/myeloid differentiation factor 88 （MyD88）/nuclear transcription factor-κB （NF-κB） signaling pathway and explore the immunological mechanism of TGP in preventing and treating systemic lupus erythematosus （SLE）. MethodMRL/lpr female mice of SPF grade were randomly divided into a model group， a dexamethasone group （0.15 g·kg^-1）， and high- （0.078 g·kg^-1） and low-dose （0.039 g·kg^-1） TGP groups， and female C57BL/6J mice were assigned to a blank group， with 7 mice in each group. Mice in each group were treated with corresponding drugs or normal saline by gavage at the same time every day. After 4 weeks， samples were collected. The kidney and spleen were weighed， and the organ index was calculated. Serum creatinine （SCr） and blood urea nitrogen （BUN） levels in each group were detected by biochemical assay. Hematoxylin-eosin （HE） staining was used to observe the histopathological changes in the kidney. The degree of renal fibrosis was evaluated by Masson staining. The serum levels of interleukin （IL）-2， interferon （IFN）-α， IL-4， and anti-nuclear antibody （ANA） were detected by enzyme-linked immunosorbent assay （ELISA）. The mRNA expression of TLR9， MyD88， and NF-κB p65 in renal tissues was detected by real-time quantitative polymerase chain reaction （Real-time PCR）. The protein expression of TLR9 and NF-κB p65 in renal tissues was detected by immunofluorescence. The protein expression of TLR9， MyD88， and NF-κB p65 in renal and spleen tissues was tested by Western blot. ResultCompared with the blank group， the model group showed increased SCr， BUN， spleen index， and kidney index （P<0.05）， deteriorated pathological injury and fibrosis in renal tissues， elevated serum levels of IFN-α， IL-4， and ANA， decreased level of IL-2 （P<0.05）， and up-regulated TLR9， MyD88， and NF-κB p65 mRNA and protein levels in the kidney and spleen （P<0.05）. Compared with the model group， the TGP groups displayed reduced SCr， BUN， spleen index， and kidney index （P<0.05）， relieved pathological damage and fibrosis in renal tissues， decreased serum levels of IFN-α， IL-4， and ANA （P<0.05）， increased level of IL-2， and declining mRNA and protein expression levels of TLR9， MyD88， and NF-κB p65 in the kidney and spleen （P<0.05）. ConclusionTGP may inhibit the expression of downstream inflammatory factors to regulate immunity and resist SLE-induced renal injury by regulating the TLR9/MyD88/NF-κB signaling pathway.

OBJECTIVETo investigate the possible role and significance of soluble programmed death-1 (sPD-1)/soluble programmed death ligand 1 (sPD-L1) in the pathogenesis of oral lichen planus (OLP).

METHODSThirty-six patients with OLP (20 cases of reticular OLP and 16 cases of erosive OLP) were enrolled in this study, and 18 healthy people served as controls. Lymphocyte subsets (CD3⁺, CD4⁺, CD8⁺, CD19⁺, CD16⁺ + 56⁺) were examined by flow cytometric analysis and humoral immunity indexes (IgG, IgA, IgM, C3, C4) tested by nephelometry immunoassay. The levels of sPD-1 and sPD-L1 proteins in serum of patients with OLP were determined by enzyme-linked immunosorbent assay. The correlations between the level of sPD-1, sPD-L1 proteins and the immune status and clinical characteristics of patients with OLP were analyzed by SPSS 19.0.

RESULTSCD3⁺, CD4⁺, CD8⁺, CD16⁺ + 56⁺ in patients with OLP were decreased compared with the normal value, while CD19⁺ in patients with OLP was increased compared with the normal value (P < 0.05). C3 and C4 in patients with OLP were decreased compared with the normal value, but IgM in patients with OLP was increased (P < 0.05). The levels of sPD-1 and sPD-L1 proteins in patients with OLP were significantly higher than that in control group [26.10(8.81, 40.00) ng/L vs 17.65(0.00, 26.10) ng/L, 29.53(21.47, 36.76) ng/L vs 22.79(1.19, 28.29) ng/L] (P < 0.05), but the expression of sPD-1 and sPD-L1 was not related with clinical characteristics of OLP. There were negative correlations between the levels of sPD-1 protein and CD4⁺ T cells or CD16⁺ + 56⁺ cells (r1 = -0.378, P1 = 0.007; r2 = -0.365, P2 = 0.009), while there was a positive correlation between the levels of sPD-1 and CD19⁺ B cells (r = 0.482, P = 0.000). There was a negative correlation between sPD-L1 expression level and CD4⁺ and a positive correlation between sPD-L1 expression level and IgG (r¹ = -0.286, P1 = 0.044; r² = 0.365, P₂= 0.029).

CONCLUSIONSIn patients with OLP, the cellular immune function is low with humoral immunity function disorder. PD-1/PD-L1 signaling pathway, which might be influenced by the involvement of sPD-1 and sPD-L1 proteins in a certain extent, may play an important role in the immune pathogenesis of OLP.

B-Lymphocytes ; B7-H1 Antigen ; blood ; Case-Control Studies ; Enzyme-Linked Immunosorbent Assay ; Flow Cytometry ; Humans ; Immunoglobulins ; blood ; Lichen Planus, Oral ; blood ; immunology ; Nephelometry and Turbidimetry ; methods ; Programmed Cell Death 1 Receptor ; blood ; Signal Transduction ; T-Lymphocytes

OBJECTIVEEmbryonic stem cells (ESCs) are derived from totipotent cells of early embryo and they are potential to differentiate to any kind of cells of tissues in the body. Some reports showed that ESCs had broad capabilities of differentiating to variety of hematopotietic cells, such as erythroid, granulocyte/macrophage, megakaryocyte, mast and lymphocyte precursors. However, it is very difficult to control the phase of differentiation for ESCs in vitro. There is few report about hematopotietic stem cells (HSCs) from ESCs. Therefore, this research was designed to establish a culture system for generation of CD(34)(+)/Sca-1(+) HSC from ESC in vitro.

METHODSSingle mouse E 14.1 cells were suspended in methylcellulose medium, containing 40 ng/ml stem cell factor (SCF) and 20 ng/ml vascular endothelial growth factor (VEGF) and incubated at 37 degrees C with 5% CO2. In order to ensure the viability of the primary differentiation cultures over an extended period of time, the cultures were fed on day 7 with a dilute methylcellulose medium containing VEGF, SCF, interleukin-3 (IL-3), IL-6 and erythropoietin (EPO), which promoted their primary differentiation into embryoid bodies (EBs) with more CD(34)(+)/Sca-1(+) cells. Then, EBs with peak level of CD(34)(+)/Sca-1(+) cells were dispersed into single cells and replanted either in methylcellulose medium or in bone marrow stromal cells differentiation system containing 15% fetal bovine serum (FBS), 160 ng/ml SCF, 20 ng/ml VEGF, 30 ng/ml IL-3, 30 ng/ml IL-6, 3 U/ml EPO and 20% BIT for HSC into second-step differentiation. The HSCs were characterized by flow cytometric analysis, colonogenic cell assay and Wright-Giemsa stains.

RESULTSVEGF had the strongest stimulatory effect on the enhancement of the CD(34)(+)/Sca-1(+) cells population when combined with SCF, IL-3, IL-6 and EPO. It could markedly accelerate mouse E14.1 cells to differentiate into EB with more CD(34)(+)/Sca-1(+) cells. Cell cytometric analysis showed CD(34)(+)/Sca-1(+) cells were up to (1.91 +/- 0.40)% by day 5 and (8.11 +/- 1.17)% by day 8, and the peak level of CD(34)(+)/Sca-1(+) cells was (13.72 +/- 1.92)% by day 12. However, CD(34)(+)/Sca-1(+) cells could not increase in number with the prolongation of differentiation. So renewal single cells suspension from EB by day 12 was dispersed into the second step differentiation. The results showed that HSC was slowly generated with a few hematopoietic colony formations in methylcellulose medium differentiation system. CD(34)(+)/Sca-1(+) cells got (2.74 +/- 0.80)% by day 5 and (11.37 +/- 1.84)% by day 10, and apex percentage of CD(34)(+)/Sca-1(+) cells was about (20.52 +/- 2.78)% by day 14. However, EBs generated quickly for HSC with increased hematopoietic cell population by co-culture on bone marrow stromal cells feeder. Flow cytometric analysis showed that the percentages of CD(34)(+)/Sca-1(+) cells was (7.33 +/- 1.61)% by day 5, (13.28 +/- 2.59)% by day 8, and (20.81 +/- 3.19)% by day 10. EB cells were induced after 12 days to reach the peak level of (34.60 +/- 3.71)%. Hematopoietic colony formation unit (CFU) analysis showed that CFU was sufficient from cells on bone marrow stromal cells differentiation system in the second step compared to that in methylcellulose medium differentiation system, and Wright-Giemsa stain could confirm its characteristics of hematopoietic progenitors.

CONCLUSIONUsing two-step differentiation, the investigators got a good way to control the phase of differentiation from ESC to HSC. The bone marrow stromal cell differentiation system combining with VEGF, SCF, IL-3, IL-6 and EPO was an optimal system for the generation of HSC with CD(34)(+)/Sca-1(+) surface marker from ESC differentiated in vitro. This study demonstrated that these cells could form more hemopoietic colonies.

Animals ; Antigens, CD34 ; Cell Culture Techniques ; Cell Differentiation ; drug effects ; Cell Survival ; Cells, Cultured ; Embryonic Stem Cells ; drug effects ; immunology ; Hematopoietic Stem Cells ; physiology ; Mice

OBJECTIVETo investigate the polymorphism of mitochondrial DNA region V in Bouyei people and Miao people living in Guizhou province of China.

METHODSPolymerase chain reaction and direct sequencing were used in the study.

RESULTSOnly two kinds of polymorphism were found in Bouyei and Miao people. One was standard pattern, the other short pattern. And the frequencies of short pattern(9 bp deletion) in Bouyei and Miao people were 31.1% and 33.3% respectively.

CONCLUSIONThe polymorphisms of mitochondrial DNA region V in Bouyei people and Miao people living in Guizhou province of China are similar, but they are different from those in other people.

Base Sequence ; China ; DNA, Mitochondrial ; chemistry ; genetics ; Female ; Gene Frequency ; Humans ; Male ; Polymorphism, Genetic ; genetics ; Sequence Analysis, DNA ; Sequence Deletion