Identification of subclusters and prognostic genes based on glycolysis/gluconeogenesis in hepatocellular carcinoma.

Background: This study aimed to examine glycolysis/gluconeogenesis-related genes in hepatocellular carcinoma (HCC) and evaluate their potential roles in HCC progression and immunotherapy response. Methods: Data analyzed in this study were collected from GSE14520, GSE76427, GSE174570, The Cancer Genome Atlas (TCGA), PXD006512, and GSE149614 datasets, metabolic pathways were collected from MSigDB database. Differentially expressed genes (DEGs) were identified between HCC and controls. Differentially expressed glycolysis/gluconeogenesis-related genes (candidate genes) were obtained and consensus clustering was performed based on the expression of candidate genes. Bioinformatics analysis was used to evaluate candidate genes and screen prognostic genes. Finally, the key results were tested in HCC patients. Results: Thirteen differentially expressed glycolysis/gluconeogenesis-related genes were validated in additional datasets. Consensus clustering analysis identified two distinct patient clusters (C1 and C2) with different prognoses and immune microenvironments. Immune score and tumor purity were significantly higher in C1 than in C2, and CD4+ memory activated T cell, Tfh, Tregs, and macrophage M0 were higher infiltrated in HCC and C1 group. The study also identified five intersecting DEGs from candidate genes in TCGA, GSE14520, and GSE141198 as prognostic genes, which had a protective role in HCC patient prognosis. Compared with the control group, the prognostic genes all showed decreased expression in HCC patients in RT-qPCR and Western blot analyses. Flow cytometry verified the abnormal infiltration level of immune cells in HCC patients. Conclusion: Results showed that glycolysis/gluconeogenesis-related genes were associated with patient prognosis, immune microenvironment, and response to immunotherapy in HCC. It suggests that the model based on five prognostic genes may valuable for predicting the prognosis and immunotherapy response of HCC patients.

YTHDF3as a prognostic predictive biomarker of thyroid cancer and its correlation with immune infiltration.

PURPOSE: Thyroid cancer (TC) is one of the most common endocrine malignancies, and its morbidity continues to rise. N6-methyladenosine (m6A) RNA methylation, an epigenetic modification, is an important regulator of gene expression in TC. Therefore, it's worth finding the characteristics and predictive value of the m6A RNA methylation regulators in thyroid cancer (TC). METHOD: RNA-seq data of TC was downloaded from the Cancer Genome Atlas (TCGA) database to screen out the differential expressed regulators. The absolute contraction selection operator (Lasso) Cox regression was used to construct the risk model of m6A methylation regulators. The predictive value of the risk scoring model was evaluated by Kaplan Meier (K-M) analysis and receiver operating characteristic (ROC) curves. The underlying mechanism of m6A methylation regulators in TC was predicted by gene set enrichment analysis (GSEA). Further validation was performed by using immunohistochemistry (IHC) and q-PCR. The correlation between risk-related gene and immune infiltration was evaluated by Tumour Immune Estimation Resource (TIMER). RESULTS: IGF2BP2, YTHDF1 and YTHDF3 were screened out as strong independent prognostic factors of TC. Then a risk score model was established to further screen the predictors. Finally, according to the results of overall survival (OS) and clinical characteristics of TC, YTHDF3 was screened out as a potential predictor. Meanwhile, IHC and qPCR confirmed that YTHDF3 was expressed differential in TC. The expression of YTHDF3 was positively associated with the infiltration level of CD4+ T cells and macrophages. It was strongly correlated with a variety of immune markers in TC. CONCLUSION: We confirmed that YTHDF3 can be used as a potential prognostic biomarker of TC. It not only plays a decisive role in the initiation and development of TC, but also provides a new perspective for understanding the modification of m6A RNA in TC.

Study and Classification of Porosity Stress Sensitivity in Shale Gas Reservoirs Based on Experiments and Optimized Support Vector Machine Algorithm for the Silurian Longmaxi Shale in the Southern Sichuan Basin, China.

To understand the characteristics of variation in porosity and permeability, the physical properties of the shale reservoir under different stress conditions play an important role in guiding shale gas production. With the shale of the Wufeng-Longmaxi Formation in the south of the Sichuan Basin as the research object, stress-dependent porosity and permeability test, high-pressure mercury injection, and scanning electron microscope test were performed in this study to thoroughly analyze the variation in physical properties of different shale lithofacies with effective stress. Besides, the stress sensitivity of different lithofacies reservoirs was evaluated by using parameters such as pore compressibility coefficient (PCC) and porosity sensitivity exponent (PSE), while the optimized support vector machine (SVM) algorithm was adopted to predict the coefficient of reservoir porosity sensitivity. According to the research results, the porosity and permeability of shale reservoirs decline as a negative exponential function. When the effective stress falls below 15 MPa, the damage rate of permeability/porosity increases rapidly with the rise of effective stress. By contrast, the permeability curvature of the shale reservoirs plunges with the rise of effective stress. It was discovered that a higher siliceous content results in a higher permeability curvature of shale, indicating the greater stress sensitivity of the reservoir. The ratio of matrix porosity to microfracture porosity determines the PSE, which is relatively low, and low aspect ratio pores contribute to high porosity compressibility and stress sensitivity. Young's modulus shows a negative correlation with pore compressibility and a positive correlation with Poisson's ratio. High clay minerals have a large number of low aspect ratio pores and a low elastic modulus, which leads to both high PCC and low PSE. Based on the principal component analysis, a multiclassification SVM model was established to predict the PSE, revealing that the accuracy of the sigmoid, radial basis function (RBF), and linear kernel function is consistently above 70%. According to error analysis, the accuracy can exceed 80% with the RBF kernel function and appropriate penalty factor. The research results serve to advance the research on the parameters related to overburden pressure, porosity, and permeability. Moreover, the optimized SVM algorithm is applied to make a classification prediction, which provides a reference for shale reservoir exploration and development both in theory and practice.

Primary cilia-dependent signaling is involved in regulating mesenchymal stem cell proliferation and pluripotency maintenance.

Using a large-scale quantitative mesenchymal stem cells (MSCs) membrane proteomics analysis, we identified a large group of ciliary proteins in the MSCs membrane fraction, which prompted us to examine the cilia, intricate organelles that were originally discovered approximately 100 years ago. Here we characterize their primary structure and function in MSCs. We first characterized the primary cilia on undifferentiated human MSCs by immunostaining and verified these observation with scanning and 3D electronic microscopy. To investigate the function of the primary cilia of the MSCs we induced loss of function by means of siRNA knockdown (targeted to two known ciliary proteins: IFT172 and KIF3A). After either of these two proteins was knocked down by the respective siRNA, the MSCs showed fewer and shortened primary cilia. The MSCs proliferation assays showed increased cell proliferative activity under confluent conditions after the siRNA knockdown of IFT172 or KIF3A; among these MSCs, the proportion in S phase was increased in the IFT172 siRNA knockdown group. The expression of stem cell markers on the MSCs, namely, Oct4, Nanog and Sox2, were downregulated after the siRNA-induced knockdown of IFT172 or KIF3A, and the gene expression upregulation of ectoderm lineage markers was notable. Furthermore, manipulation of the primary cilia-dependent Shh pathway, using the Shh activator SAG (smoothened agonist), upregulated the gene expression of pluripotency markers, including Nanog and Oct4, and transcriptional target genes in the Shh pathway. This study confirms that MSCs have primary cilia and provides evidence that primary cilia-dependent signaling pathways play functional roles in MSCs proliferation and stemness maintenance.

Ultrasmall Oxygen-Deficient Bimetallic Oxide MnWOX Nanoparticles for Depletion of Endogenous GSH and Enhanced Sonodynamic Cancer Therapy.

Sonodynamic therapy (SDT) triggered by ultrasound (US) has attracted increasing attention owing to its abilities to overcome critical limitations including low tissue-penetration depth and phototoxicity in photodynamic therapy. Herein, the design of a new type of sonosensitizer is revealed, namely, ultrasmall oxygen-deficient bimetallic oxide MnWOX nanoparticles, for multimodal imaging-guided enhanced SDT against cancer. As-made MnWOX nanoparticles with poly(ethylene glycol) (PEG) modification show high physiological stability and biocompatibility. Interestingly, such MnWOX -PEG nanoparticles exhibit highly efficient US-triggered production of 1 O2 and •OH, higher than that of previously reported sonosensitizers (e.g., protoporphyrin IX and titanium dioxide), because the oxygen-deficient structure of MnWOX serves as an electron trap site to prevent electron-hole recombination. The glutathione depletion capability of MnWOX -PEG can also further favor SDT-triggered cancer cell killing. With efficient tumor homing as illustrated by computer tomography and magnetic resonance imaging, MnWOX -PEG enables effective destruction of mouse tumors under US stimulation. After accomplishing its therapeutic functions, MnWOX -PEG can be metabolized by the mouse body without any long-term toxicity. Herein, a new type of sono-sensitizing agent with high SDT efficacy, multimodal imaging functions, and rapid clearance is presented, an agent which is promising for noninvasive SDT cancer treatment.

Coffee consumption is positively related to insulin secretion in the Shanghai High-Risk Diabetic Screen (SHiDS) Study.

BACKGROUND: It has been proved that coffee consumption was associated with a lower risk of type 2 diabetes mellitus. But the benefit effect of coffee on hyperglycemia in Chinese population was largely unknown. Besides, the relationship of coffee intake and diabetic pathogenesis was still unclear. METHODS: The study population was selected from the Shanghai High-Risk Diabetic Screen (SHiDS) project. A total of 1328 individuals over 18 years of age who have the information of coffee intake were enrolled in the study from 2012 to 2016. Each participant finished a five-point 75 g oral glucose tolerance test and finished a standard questionnaire. Insulin resistance was evaluated by HOMA-IR and insulin secretion was evaluated by HOMA-ß, Stumvoll first phase and second phase indexes. RESULTS: Coffee consumption group had lower plasma glucose levels at 2-h and 3-h and higher insulin levels at fasting, 30-min and 1-h during OGTT after adjustment with age, fat%, BMI, waist, tea intake, smoking habit, alcohol intake, diabetes family history and educational status (P for PG2h = 0.002; P for PG3h = 0.010; P for FIN = 0.010; P for IN30min = 0.001; P for IN1h = 0.002). Both HOMA-ß and Stumvoll formula indexes were positively related to coffee consumption (P for HOMA-ß = 0.033; P for Stumvoll first phase = 0.003; P for Stumvoll second phase = 0.001). Logistic regression analysis further confirmed that coffee intake was independently associated with higher levels of HOMA-ß and Stumvoll insulin secretion indexes [OR (95% CI) for HOMA-ß = 2.270 (1.456-3.538); OR (95% CI) for Stumvoll first phase = 2.071 (1.352-3.173); OR (95% CI) for Stumvoll second phase = 1.914 (1.260-2.906)]. CONCLUSIONS: Coffee intake is independently and positively related to pancreatic beta cell function in a large high-risk diabetic Chinese population.

Serum is an indispensable factor in the maintenance of the biological characteristics of sweat gland cells.

The tolerance of sweat gland cells for in vitro amplification and subcultivation is low as they are somatic cells. The present study aimed to formulate an optimal medium for the culture of human eccrine sweat gland cells (HESGCs) and to establish a method for induction of HESGCs proliferation, whilst maintaining the characteristics of sweat gland cells. HESGCs cultured in sweat gland (SG):keratinocyte growth medium­2 (KGM­2) (1:1) medium had a higher proliferation rate and a stable morphology compared with cells cultured in SG and KGM­2 medium only. Reverse transcription­quantitative polymerase chain reaction indicated that cells cultured in the SG:KGM­2 (1:1) medium exhibited higher expression levels of α­smooth muscle actin, keratin (K)77, carcinoembryonic antigen, K8, K18, ectodysplasin A receptor, c­Myc, Kruppel­like factor 4 and octamer­binding transcription factor 4 compared with cells cultured in SG only or KGM­2 only medium. Three­dimensional culture analysis revealed that HESGCs cultured in SG:KGM­2 1:1 medium differentiated into sweat gland­like structures, whereas cells cultured in KGM­2 only medium underwent cornification. The present study also determined that the maintenance of the biological characteristics of HESGCs occurred due to the presence of fetal bovine serum (FBS). Cells cultured in medium without FBS differentiated into keratinocytes. Therefore, the SG:KGM­2 (1:1) medium may be a suitable culture medium for HESGCs. In conclusion, this mixed medium is a valuable compound and should be considered to be a potential supplemental medium for HESGCs.

Plasma Level of Placenta-Derived Macrophage-Stimulating Protein -Chain in Preeclampsia before 20 Weeks of Pregnancy.

OBJECT: This study aimed to investigate the diagnostic value of placenta-derived macrophage-stimulating protein α-chain (MSP-α) before the 20th week of gestation for the early diagnosis of preeclampsia (PE). METHODS AND MATERIALS: Two parts of this nested case-control study were simultaneously executed, and 1500 pregnant women were recruited. A total of 124 pregnant women were included in the plasma analysis part of this study. The MSP-α plasma level was measured before the 20th week of gestation, and the participants were followed until delivery. A case group of 62 women with PE and a control group of 62 women matched by gestational age, maternal age, and pre-pregnancy BMI (with normotensive pregnancies) were evaluated. In the placenta analysis part of this nested case-control study, the placentas of 34 pregnant women were randomly obtained. The placental levels of MSP were measured in 17 individuals with PE (case group) and in 17 women with a normotensive pregnancy matched by gestational age and maternal age (control group). RESULTS: The plasma level of MSP-α was higher in the PE group than in the control group before the 20th week of gestation (p < 0.001). In addition, compared to the women with severe features in the PE group, those without severe features had a significantly higher plasma MSP-α level before the 20th week of gestation (p < 0.001). The area under the receiver operating characteristic curve (AUC) of MSP-α before the 20th week of gestation was 0.905 (95% CI, 0.811-0.962) for the women with early-onset PE without severe features. With regard to the placenta, the PE group (accumulated optical density, IOD [SUM] = 8862.37 ± 2064.42) exhibited increased MSP staining (more intense MSP staining or more extensive staining) compared with the control group (normal pregnancies (IOD [SUM] = 447.92 ± 114.72, P < 0.001). Furthermore, increased MSP staining was detected among the women without severe features compared with those with severe features in the PE group (IOD [SUM]: 12192.65 ± 5325.56 vs. 4104.83 ± 2383.06, P = 0.021). CONCLUSION: According to the findings of this study, the plasma level of MSP-α may be associated with PE, and MSP-α may be considered a candidate protein for further analysis in studies of PE. Multicenter studies with larger sample sizes must be performed in the future to obtain accurate results regarding the predictive value of MSP-α in combination with other protein factors for the early diagnosis of PE.

Direct Reprogramming of Human Amniotic Fluid Stem Cells by OCT4 and Application in Repairing of Cerebral Ischemia Damage.

Amniotic fluid stem cells (AFSCs) are a type of fetal stem cell whose stemness encompasses both embryonic and adult stem cells, suggesting that they may be easily and efficiently reprogrammed into induced pluripotent stem cells (iPSCs). To further simplify the reprogramming process, the creation of AFSC-derived iPSCs using a single factor is desirable. Here we report the generation of one-factor human AFSC-iPSCs (AiPSCs) from human AFSCs by ectopic expression of the transcription factor OCT4. Just like human embryonic stem cells, AiPSCs exhibited similar epigenetic status, global gene expression profiles, teratoma formation and in vitro & in vivo pluripotency. Our results indicate that the OCT4 is necessary and sufficient to directly reprogram human AFSCs into pluripotent AiPSCs. Moreover, reflecting the similar memory characteristics of AFSCs and neural stem cells, we show that AiPSC membrane-derived vesicles (MVs) repair cerebral ischemia damage. We anticipate that the successful generation of one-factor AiPSCs will facilitate the creation of patient-specific pluripotent stem cells without the need for transgenic expression of oncogenes. Moreover, MVs from tissue-specific AiPSCs have potential in tissue repair, representing a novel application of iPSCs.

Suspended, Shrinkage-Free, Electrospun PLGA Nanofibrous Scaffold for Skin Tissue Engineering.

Electrospinning is a technique for creating continuous nanofibrous networks that can architecturally be similar to the structure of extracellular matrix (ECM). However, the shrinkage of electrospun mats is unfavorable for the triggering of cell adhesion and further growth. In this work, electrospun PLGA nanofiber assemblies are utilized to create a scaffold. Aided by a polypropylene auxiliary supporter, the scaffold is able to maintain long-term integrity without dimensional shrinkage. This scaffold is also able to suspend in cell culture medium; hence, keratinocyte cells seeded on the scaffold are exposed to air as required in skin tissue engineering. Experiments also show that human skin keratinocytes can proliferate on the scaffold and infiltrate into the scaffold.

Children Diagnosed as Mixed-Phenotype Acute Leukemia Didn't Benefit from the CCLG-2008 Protocol, Retrospective Analysis from Single Center.

Mixed phenotype acute leukemia (MPAL) is a rare type of acute leukemia with a poor clinical outcome which lacks specific therapy. To evaluate the therapeutic efficiency of CCLG-2008 protocol used for acute lymphoblastic leukemia (ALL) in China on MPAL children who were initially diagnosed as ALL by morphology, we reviewed patients' database diagnosed as ALL and MPAL according to WHO classification and compared their outcomes from July 2008 to June 2012. Total newly enrolled ALL in this study were 309 cases by morphology, in which ten cases were identified as MPAL mainly by immunophenotyping: B+ myeloid (3/10), T+ myeloid (2/10), B + T (4/10), trilineage (1/10). Two cases were classified as intermediate risk (IR) and 8 cases were high risk (HR) according to the CCLG-2008 criteria. Only one case of IR survived and others died due to primary resistance of chemotherapy and relapse. Compared with MPAL, ALL children in IR and HR had a longer survival (28.1 vs 9.5 months, p < 0.0001) and lower relapse (16.3 vs 85.7 %, p = 0.0002). In a summary, our result indicated that MPAL in children is a poor-risk disease which needs personalized therapy to improve outcome.

Prevalence of type 2 diabetes among high-risk adults in Shanghai from 2002 to 2012.

OBJECTIVE: The objective of this study was to evaluate the trend and prevalence of prediabetes and diabetes among high-risk adults in Shanghai from 2002 to 2012. METHODS: From 2002 to 2012, 10043 subjects with known risk factors for diabetes participated in the diabetes-screening project at the Shanghai Sixth People's Hospital of Shanghai Jiao Tong University. All participants were asked to complete a nurse-administered standard questionnaire concerning age, sex, smoking status, and personal and family histories of diabetes, cardiovascular disease, stroke, hypertension and other diseases. The participants' body mass index scores, blood pressures and blood glucose levels at 0, 30, 60, 120 and 180 min were measured in response to a 75 g oral glucose tolerance test. RESULTS: The overall prevalence of diabetes increased from 27.93% to 34.78% between 2002 and 2012 in high-risk subjects. The study also showed that the prevalence increased much faster in male compared to female subjects. Specifically, an increased rate was seen in middle-aged men, with no change observed in middle-aged females over the eleven-year period. CONCLUSION: This study showed that sex, age, parental diabetic history, and being overweight were associated with an increased risk for diabetes in high-risk people. Therefore, as prediabetes and diabetes are highly prevalent in people with multiple diabetes risk factors in Shanghai, screening programs targeting these individuals may be beneficial.

Homology modeling and molecular dynamics studies of Wilms' tumor gene 1 frameshift mutations in exon 7.

As a transcription factor, the Wilms' tumor 1 (WT1) gene plays an important role in leukemogenesis. The impact of WT1 gene mutations has been reported in acute myeloid leukemia (AML). However, the number of available studies on the spatial configuration changes following WT1 mutation is limited. In this study, we sequenced the mutation in exon 7 of the WT1 gene in 60 children with newly diagnosed AML and the spatial configuration of WT1 with frameshift mutations in exon 7 was evaluated using the software for homology modeling and optimization of molecular dynamics. Three cases with frameshift mutations in exon 7 were identified (3/60; mutation rate, 5%). One case had a mutation that had been previously described, whereas the remaining two mutations were first described in our study. Of the three cases, one case presented with antecedent myelodysplastic syndrome (MDS) and the remaining two cases exhibited primary resistance to induction chemotherapy. The spatial configuration analysis demonstrated that the three mutations affected the spatial structure of exon 7 and even affected exon 8 compared to its wild-type. This study demonstrated that the frameshift mutation in exon 7 of the WT1 gene is a poor prognostic factor for children with AML, partly through the spatial configuration changes following frameshift mutations of WT1, which highlights the structure-based function analysis and may facilitate the elucidation of the pathogenesis underlying WT1 gene mutations.

[Effects of Brucella phosphoglucomutase on inducing infection of trophoblastic cells].

OBJECTIVE: We studied the biological function of Brucella phosphoglucomutase (pgm) gene, and detected the changes of human trophoblast cell invaded by the Brucella pgm mutant and PGM protein. METHODS: Human trophoblast cells were infected by the pgm mutant and PGM protein. The changes of cytokines were detected by enzyme-linked immunosorbent assay, and morphology of cells was identified. RESULTS: PGM protein was purified, and pgm mutant was constructed. The sera of mice immunized by pgm mutant were negative by agglutination test and Standard Tube Agglutination Test for Brucellosis. The cellular morphology of human trophoblast cells infected pgm mutant or PGM protein changed. The adhesion and infection of the pgm mutant reduced more than Brucella vaccine strain M5-90, and human trophoblast cells partially cracked off. The activity of IL-6, TNF-alpha or lactic dehydrogenae increased in human trophoblast cells infected by the pgm mutant more than Brucella vaccine strain M5-90 (P < 0.01), but not for IL-10. Lactic dehydrogenae in human trophoblast cells infected by the PGM Protein increased more than sodium phosphate buffer (P < 0.01), whereas IL-6 and TNF-alpha decreased in human trophoblast cells less than sodium phosphate buffer (P < 0.05). CONCLUSION: The results suggest that the pgm mutant of brucella and PGM protein had the cytotoxic effect for human trophoblast cells with cellular morphology and changes of cytokines.

Function and expression of insulin-like growth factor-binding protein 7 (IGFBP7) gene in childhood acute myeloid leukemia.

Insulin-like growth factor-binding protein 7 (IGFBP7) has been identified as a tumor suppressor in solid tumors. In acute leukemia, the role of IGFBP7 is largely unknown. The authors used quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) to investigate the expression level of IGFBP7 gene in bone marrow (BM) specimen from 66 children with acute myeloid leukemia (AML) at different stages and in 30 nonleukemia patients as control. Furthermore, U937 cells were transfected with siRNA-2 of IGFBP7 (as U937R) for 24 hours. Coculture experiment was performed to explore the impact of IGFBP7 gene in U937 cell adhesion, invasion, and migration in existing ECV304 cells, which mimicked the interaction between AML cells and endothelial cells. IGFBP7 expression at the initial diagnosed stage and relapse of AML was significantly higher than that of control (P < .001). The viable cell percentage in transfected cell was significantly decreased by 42% compared with control groups (P < .01). The percentage for U937R cells adherent to ECV304 cells was significantly lower than the control groups (P < .01). Matrigel study to quantify the invasive potential showed that U937R migrated to the lower chamber were significantly less than those in the parental control groups (P < .01). In summary, IGFBP7 aberrantly overexpressed in majority of AML at diagnosis and upon relapsed, but not at remission stage. IGFBP7 plays a positive contributing role in the interaction between leukemia cells and microenvironment, which may promote the leukemic cells' adhesion, invasion, and migration.

Clinical study on the treatment of knee osteoarthritis of Shen-Sui insufficiency syndrome type by electroacupuncture.

OBJECTIVE: To study the clinical effificacy of electroacupuncture (EA) on treating knee osteoarthritis (KOA) of Shen ()-Sui () insuffificiency (SSI) syndrome type. METHODS: A total of 245 patients (279 knees) of KOA-SSI were randomly assigned to two groups by lottery: 141 knees in the treatment group and 138 knees in the control group. The treatment group was managed with EA at the dominant points of Neixiyan (Ex-LE4) and Waixiyan (Ex-LE5) as well as the conjugate points of Xuanzhong (GB39) and Taixi (KI3) for 30 min, once a day, with 15 days as one course; 2 courses were applied with a 5-day interval in between. The control group was treated with intra-articular injection of 2 mL hyaluronic acid into the affected joint every 7 days for 5 times in total. The clinical effects on the patients in different stages were observed, and their symptom scores of knee and contents of cytokines, including interleukin-1 (IL-1), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), prostaglandin E(2alpha) (PGE(2alpha)) and matrix metalloproteinases-3 (MMP-3), in the knee joint fluid were measured before and after treatment. RESULTS: The study was completed in 235 patients (263 knees); four patients (7 knees) in the treatment group and six patients (9 knees) in the control group dropped out. Comparison of therapeutic effects (excellent and effective rates) between the two groups showed insignificant differences (P>0.05). Symptom scores of knee and contents of cytokines in the knee flfluid after treatment were lowered signifificantly in the patients of stage I-III in both groups (P<0.05 or P<0.01). However, the lowering of the total symptom score of knee in the patients of stage III in the treatment group was more signifificant (P<0.05). CONCLUSIONS: EA could effectively alleviate the clinical symptoms in KOA patients of stage III, showing an effect superior to that of hyaluronic acid. EA also shows action in suppressing the secretion of IL-1, IL-6, TNF-alpha, PGE(2alpha) and MMP-3 in the knee flfluid.

[Effect of shock lymph on the expression of inflammatory mediators in pulmonary micro-vascular endothelial cells of rats].

OBJECTIVE: To observe the effect of lymph collected during shock on free radical and expressions of nitric oxide (NO), tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) mRNA of pulmonary micro-vascular endothelial cells (PMVECs) of rats in order to explore the mechanisms of damaging effect of lymph collected during shock to the PMVECs. METHODS: PMVECs were isolated and cultured, and used at passage 3. The model of serious hemorrhagic shock was reproduced by maintaining the arterial blood pressure of rats at 40 mm Hg (1 mm Hg=0.133 kPa) for 90 minutes by exsanguination under aseptic condition. Mesentery lymph and portal vein blood were obtained from both shock rats and normal rats. PMVECs were respectively incubated in them for 6 hours, and at the same time, fetal bovine serum (FBS) and serum-free DMEM were used as culture media for comparison. The expressions of inducible nitric oxide synthase (iNOS), TNF-alpha and IL-6 mRNA were detected by the method of reverse transcription-polymerase chain reaction (RT-PCR), and the content of malondialdehyde (MDA), NO, TNF-alpha and IL-6 in culture supernatants were determined. RESULTS: After the PMVECs was treated by shock lymph at a final concentration of 4% for 6 hours, the expressions of iNOS, TNF-alpha and IL-6 mRNA in PMVECs and the contents of MDA, NO, TNF-alpha and IL-6 in culture supernatant fluids in shock lymph group were significantly increased compared with those of FBS group, normal lymph group, shock plasma group, normal plasma group and DMEM group. At the same time, the expressions of iNOS, TNF-alpha and IL-6 mRNA in PMVECs and the contents of NO in culture supernatant fluid of shock plasma group were significantly increased compared with those of FBS group, normal lymph group, normal plasma group and DMEM group (P<0.05 or P<0.01). CONCLUSION: The results demonstrate that the shock lymph in final concentration of 4% could enhance the expressions of iNOS, TNF-alpha and IL-6 of PMVECs, reduce the free radical, and as a result, induce damage to PMVECs.

Elimination of C5aR prevents intestinal mucosal damage and attenuates neutrophil infiltration in local and remote organs.

The complement C5a pathway has been shown to be an important mediator of inflammation and tissue injury. To further understand the role of C5a receptor (C5aR) pathway in ischemia/reperfusion (I/R) injury, and to evaluate the potential of antagonizing C5aR to protect from I/R injury, we tested the effect of eliminating C5aR using C5aR knockout (KO) mice and their wild-type (WT) littermates in a superior mesenteric artery occlusion (SMAO) intestinal I/R injury model. C5aR KO and WT mice were subjected to SMAO or sham for 45 min. After 3 h of reperfusion, the percentage of injured ileal villi was twice as high in WT mice subjected to SMAO as compared with the C5aR KO mice. In addition, the number of neutrophils was 34% higher in WT mice subjected to SMAO as compared with the C5aR KO mice. Moreover, ileum and lung myeloperoxidase activities after SMAO were significantly higher in WT than C5aR KO mice. Apoptotic cell death was induced after reperfusion in WT-SMAO and was reduced by more than 50% in C5aR KO mice. The plasma level of TNF-alpha was increased approximately 3.74-fold in WT subjected to SMAO compared with sham. In contrast, the level was increased only approximately 1.18-fold in the C5aR KO mice subjected to SMAO. In conclusion, this study demonstrates that elimination of the C5aR pathway protects the intestine from I/R injury and diminishes intestine-derived pulmonary neutrophil sequestration. Blocking C5aR may be considered as a potential therapeutic intervention for I/R injury.

[Effect of ligation of mesenteric lymph duct on inflammation response of liver in hemorrhagic shock in rats].

OBJECTIVE: To observe the effect of ligation of mesenteric lymph duct on changes in free radicals and pro-inflammatory mediators in the liver of rats with serious hemorrhagic shock at different periods, and explore the effect of blockage of intestinal lymphatic pathway on inflammation response of liver. METHODS: Seventy-eight male Wistar rats were randomly divided into the sham group (n=6), shock group (n=42), and ligation group (n=30). The model of serious hemorrhagic shock was reproduced in shock group and ligation group. Mesenteric lymph was blocked by ligating mesenteric lymph duct in ligation group after resuscitation. Six rats were sacrificed at 0, 1, 3, 6, 12, and 24 hours, and the livers were harvested and homogenized for the determination of malondialdehyde (MDA), nitric oxide (NO), nitric oxide synthase (NOS), superoxide dismutase (SOD), tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6) and myeloperoxidase (MPO) activity. The expression of inducible nitric oxide synthase (iNOS) mRNA in liver was detected by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: The contents of TNF-alpha, IL-6, NO, NOS, MDA, MPO and iNOS mRNA in liver homogenate of shock group were increased after transfusion and resuscitation, and their levels were higher at 6 and 12 hours. The values were significantly higher than those of the sham group, while the activity of SOD was significantly lower than that of sham group (P<0.05 or P<0.01). The contents of TNF-alpha, IL-6, NO, NOS, MDA, MPO and iNOS mRNA in liver homogenate were lower significantly after transfusion and 3, 6, 12 and 24 hours after resuscitation than those of shock group at each time points, and the SOD activity was higher (P<0.05 or P<0.01). CONCLUSION: The results demonstrate that the ligation of mesenteric lymph duct could reduce the polymorphonuclear leucocyte (PMN) detain, and its mechanism might relate to reduction of neutrophil aggregation, thus decreases the release of TNF-alpha and IL-6, reduces the NO and expression of iNOS mRNA, reduces the release of free radicals and consumption of SOD, as a result, it reduces the inflammation response of liver in serious hemorrhagic shock rats.