Effect of Echinococcus multilocularis infection on Tim3 expression in spleen natural killer cells of mice / 中国血吸虫病防治杂志

Objective To investigate the effect of Echinococcus multilocularis infection on Tim3 expression and its co-expression with immune checkpoint molecules 2B4 and LAG3 in spleen natural killer (NK) cells of mice. Methods C57BL/6 mice, each weighing (20 ± 2) g, were randomly divided into a high-dose infection group (15 mice), a low-dose infection group (13 mice), and a control group (11 mice). Mice in the high- and low-dose infection groups were inoculated with 2 000 and 50 Echinococcus multilocularis protoscolices via the hepatic portal vein, while animals in the control group was injected with an equivalent amount of physiological saline via the hepatic portal vein. Mouse spleen cells were harvested 12 and 24 weeks post-infection, and Tim3 expression and its co-expression with 2B4 and LAG3 in NK cells were detected using flow cytometry. Results There were significant differences in the proportions of Tim3 expression (F = 13.559, P < 0.001) and Tim3 and 2B4 co-expression (F = 12.465, P < 0.001) in mouse spleen NK cells among groups 12 weeks post-infection with E. multilocularis, and the proportion of Tim3 expression was significantly higher in mouse spleen NK cells in the low-dose infection group [(23.84 ± 2.28)%] than in the high-dose infection group [(15.72 ± 3.67)%] and the control group [(16.14 ± 3.83)%] (both P values < 0.01), while the proportion of Tim3 and 2B4 co-expression was significantly higher in mouse spleen NK cells in the low-dose infection group [(22.20 ± 2.13)%] than in the high-dose infection group [(14.17 ± 3.81)%] and the control group [(15.20 ± 3.77)%] (both P values < 0.01). There were significant differences in the proportions of Tim3 expression (F = 5.243, P < 0.05) and Tim3 and 2B4 co-expression (F = 4.659, P < 0.05) in mouse spleen NK cells among groups 24 weeks post-infection with E. multilocularis infection, and the proportions of Tim3 expression and Tim3 and 2B4 co-expression were significantly lower in mouse spleen NK cells in the high-dose infection group [(20.55 ± 7.04)% and (20.98 ± 7.12)%] than in the control group [(31.38 ± 3.19)% and (31.25 ± 3.06)%] (both P values < 0.05), and there were no significantly difference between the proportions of Tim3 expression and Tim3 and 2B4 co-expression in splenic NK cells in the low-dose infection group [(26.80 ± 6.47)% and (26.48 ± 6.48)%] and the control group (both P > 0.05). There were no significant differences in the proportions of Tim3 and LAG3 co-expression in mouse spleen NK cells among groups 12 (F = 2.283, P > 0.05) and 24 weeks post-infection (F = 0.375, P > 0.05). In the low-dose infection group, there were no significant differences in the proportions of Tim3 expression or Tim3 and 2B4 co-expression in mouse spleen NK cells 12 (t = −1.137, P > 0.05) or 24 weeks post-infection (t = −1.658, P > 0.05), and the proportion of Tim3 and LAG3 co-expression increased in mouse spleen NK cells 24 weeks post-infection relative to 12 weeks post-infection (t = −5.261, P < 0.01). In the highdose infection group, there was no significant difference in the proportion of Tim3 expression in mouse spleen NK cells 12 and 24 weeks post-infection (t = −1.546, P > 0.05); however, the proportions of Tim3 co-expression with 2B4 and LAG3 increased in mouse splenic NK cells 24 weeks post-infection relative to 12 weeks post-infection (t = −2.425 and −4.745, both P values < 0.05). Conclusions The Tim3 expression and Tim3 co-expression with LAG3 and 2B4 on spleen NK cells is affected by doses of E. multilocularis infection and disease stages, and present different phenotypes during the course of alveolar echinococcosis. NK cells tend to form an immunosuppressive phenotype with the progression of E. multilocularis infection, which facilitates immune escape and chronic parasitism of E. multilocularis.

Study on the characteristics of 18F-flurodeoxyglucose uptake in the microenvironment of hepatic alveolar echinococcosis / 中华肝胆外科杂志

Objective:To investigate characteristics of the 18F-flurodeoxyglucose ( 18F-FDG) uptake intensity and ranges in distinct hepatic alveolar echinococcosis lesions. Methods:The clinical data of 39 patients with position emission tomography during Jan 2017 to Dec 2019 in the First Affiliated Hospital of Xinjiang Medical University were enrolled. Among them, there were 17 males and 22 females, aging from 15 to 65 years (median 34 years). Lesions were classified into six groups based on heterogenic scales of calcification and liquefaction: A. non-calcified and non-liquefied ( n=7); B. obvious calcified and non-liquefied ( n=7); C. partial calcified and partial liquefied( n=10); D. obvious calcified and partial liquefied ( n=5); E. partial calcified and subtotal liquefied ( n=5); F. obvious calcified and subtotal liquefied ( n=5). Tumor to background ratio (TBR) and width (W) of lesion infiltrative boundary were measured and calculated. Statistical comparison using Mann-Whitney U test as well as correlation analysis was performed. Results:TBR values [ M( Q1, Q3)] for each group were 4.40(3.66, 7.03), 2.55(1.69, 3.60), 3.73(3.37, 5.21), 2.90(2.75, 3.60), 3.80(3.49, 6.36), 2.49(2.21, 3.97), among which A>B, A>D, A>F, C>B, E>B ( U=3.0, 4.0, 4.5, 11.0, 5.0, all P<0.05); From the perspective of the calcification in each group, it was found that the lighter the calcification was, the greater the TBR value was. W values [ M( Q1, Q3)] for each group were [12.5(10.0, 19.5), 11.2(10.5, 12.5), 12.2(10.9, 13.2), 7.8(7.3, 9.3), 10.0(7.3, 13.4), 7.3(6.8, 7.6)] mm, among which A>D, A>F, B>D, B>F, C>D, C>F (all U=0, all P<0.05); According to the degree of calcification and liquefaction of lesions in each group, the lighter the calcification was, the greater the W value was; The heavier the liquefaction was, the smaller the W value was. A mild strength linear correlation has been observed between the TBR value and W value ( r=0.4136, P<0.05). Conclusions:Less calcification and liquefaction implicated higher 18F-FDG uptake intensity and wider range. Radical resection margins and tissue sampling should be individualized based on different lesion features in surgical treatment.

Current progress on lesion classification of hepatic alveolar echinococcosis / 中华肝胆外科杂志

Human alveolar echinococcosis is a chronic infectious disease caused by Echinococcus multilocularis infection. It predominantly injuries the liver and grows like the malignant tumor. The therapeutic options and prognosis depend on types of human alveolar echinococcosis, clinical stages, biological activity, vascular invasion, pathological characteristics, and patient's immune status. However, despite of multiple classification methods, there are still lacking of comprehensive typing schemes. which leads to inappropriate diagnosis and therapy. This research systematically reviewed the recent studies on human alveolar echinococcosis at home and abroad and analyzed the classifications based on ultrasound, computer tomography, magnetic resonance imaging, positron emission computed tomography, serology and pathology, and some novel technologies and summarized the individual advantage and disadvantage for each classification Relationships and their advantages plus disadvantages have been assessed comprehensively. Meanwhile, the possible reference factors or theoretical basis for optimized future classification are proposed, in order to establish a unified classification system to provide guidance for clinical diagnosis and treatment.

Treatment of primary hepatic carcinoma by transcatheter artery combined with portal vein chemoembolization / 实用放射学杂志

Objective To compare the clinical efficacy and postoperative liver function in patients with primary hepatic carcinoma treated by transcatheter arterial chemoembolization(TACE) or TACE combined with portal vein chemoembolization.Methods 48 patients with primary hepatic carcinoma, randomly divided into 2 groups (hepatic artery group in 25 cases and dual interventional group in 23 cases),underwent interventional treatment.The hepatic artery group underwent conventional hepatic artery interventional therapy, while the dual interventional group underwent hepatic artery and portal vein interventional treatment.The postoperative clinical efficiency, liver volume and liver function between the two groups'' patients were compared.Results To the endpoint of observation,the clinical efficacy and tumor reduction degree of dual interventional group were better than that of hepatic artery group.Compared with hepatic artery group, the postoperative ALT, AST and TBIL of dual interventional group were higher on the first and third days.On the seventh and fourteenth days, the statistical difference was not significant.The volume of non-embolization part in dual interventional group was larger than that in preoperative volume to different degrees.The most obvious change of liver volume happened in the 4th weeks after treatment.There was no treatment-related death or severe adverse reaction in two groups.Conclusion The treatment of TACE combined with portal vein chemoembolization is a safe and effective method, which may effectively inhibit the growth and reduce the volume of tumor, and result in compensatory hypertrophy of non-embolization part.

Expression of β-tubulin Ⅲ in colon adenocarcinoma tissue and its clinical significance / 吉林大学学报(医学版)

Objective:To detect the expression and clinical significance of β-tubulin Ⅲ in cancer tissue of the patients with colon adenocarcinoma,and to explore its clinical significance.Methods:A total of 111 colon adenocarcinoma tissue samples were obtained.According to the location of β-tubulin Ⅲ positive cells, all patients were divided into front group (n=72) and non-front group (n=39).The positive expression rate of β-tubulin Ⅲ in the patients with colon adenocarcinoma was detected with immunohistochemistry.The correlations among the expression of β-tubulin Ⅲ and gender,age,tumor differentiation, clinical stage, lymph node metastasis, recurrence and death were analyzed.Results: The expression levels of β-tubulin Ⅲ had no significant differences between the patients with different gerder,age,lymph node metastasis,clinical stages,death and recurrence.The positive expression rates of β-tubulin in cancer tissue of the patients had significant difference between front and non-front groups (χ2=8.76, P=0.01).Lowly-to-moderately differentiated tissue was more common in front group, and highly-differentiated tissue was more common in non-front group(χ2=6.88, P=0.03).There were significant differences in the expression levels of β-tubulin Ⅲ between cancer tissues with different differentiation degrees (χ2=5.74, P=0.04).In non-front group, lymph node metastasis was closely correlated with the expression of β-tubulin Ⅲ (χ2=6.02,P=0.05).The results of immunohistochemical staining showed that the β-tubulin Ⅲ positive-expressing cells were colored brown-yellow.The number of cells with positive β-tubulin Ⅲ expression was significantly increased in highly differentiated tissue compared with low-differentiated tissue.Conclusion:The expression of β-tubulin Ⅲ is closely related to tumor differentiation in colon adenocarcinoma tissue.The highly differentiated colon adenocarcinoma tissue is more common in non-front group in which the expression of β-tubulin Ⅲ is related to lymph node metastasis.

Molecular Cloning and Characterization of a P38-Like Mitogen-Activated Protein Kinase from Echinococcus granulosus

Cystic echinococcosis (CE) treatment urgently requires a novel drug. The p38 mitogen-activated protein kinases (MAPKs) are a family of Ser/Thr protein kinases, but still have to be characterized in Echinococcus granulosus. We identified a 1,107 bp cDNA encoding a 368 amino acid MAPK protein (Egp38) in E. granulosus. Egp38 exhibits 2 distinguishing features of p38-like kinases: a highly conserved T-X-Y motif and an activation loop segment. Structural homology modeling indicated a conserved structure among Egp38, EmMPK2, and H. sapiens p38α, implying a common binding mechanism for the ligand domain and downstream signal transduction processing similar to that described for p38α. Egp38 and its phosphorylated form are expressed in the E. granulosus larval stages vesicle and protoscolices during intermediate host infection of an intermediate host. Treatment of in vitro cultivated protoscolices with the p38-MAPK inhibitor ML3403 effectively suppressed Egp38 activity and led to significant protoscolices death within 5 days. Treatment of in vitro-cultivated protoscolices with TGF-β1 effectively induced Egp38 phosphorylation. In summary, the MAPK, Egp38, was identified in E. granulosus, as an anti-CE drug target and participates in the interplay between the host and E. granulosus via human TGF-β1.

Autofluorescence and spectrum characteristics of Echinococcus granulosus culturing in vitro / 中国人兽共患病学报

The objective of this study was to illustrate the autofluorescence phenomenon and scanning confocal λcharacteris‐tics of Echinococcus granulosus (Eg) cultured in vitro ,which can not only rich the spectroscopy and biological information of Eg ,but also lay a foundation for study the immunofluorescence and medical photonics .Protoscoleces (PSC) of Eg were aspi‐rated and pooled from sheep liver hydatid cysts collected from a slaughterhouse .The PSC viability was determined by Methyl‐ene blue staining .The autofluorescence of endocyst were observed through the confocal microscope excited respectively by 405 nm ,488 nm ,514 nm ,and 561 nm .The autofluorescence curve was measured by thermo Varioskan Flash respectively by 405 nm ,488 nm ,514 nm ,561 nm ,and 633 nm .Theλ scanning analysis was executed by confocal respectively by 405 nm ,488 nm ,514 nm ,561 nm ,and 633 nm to measure the autofluorescence curve .The Eg excited by different excitation light through confocal showed different colors autofluorescence .Under the same excitation intensity and detector voltage condition ,blue fluo‐rescence showed the best fluorescence effect and clearer structure which excited by 405 nm laser .Scanning respectively by five different excitation lights ,the corresponding sensitive emission wave lengths scope were 490‐520 nm ,around 520 nm and 580 nm ,580‐600 nm ,610‐630 nm ,and around 650 nm ,respectively .The 405 nm ,488 nm and 561 nm excitation effect were bet‐ter than that of the 514 nm and 633 nm .The autofluorescence curve detected by Microplate Reader were basically as same as that by confocal ,although the intensity detected by Microplate Reader were lower .The results indicated that the best excita‐tion light to detect the autofluorescence of Eg is 405 nm ,which showed blue fluorescence . The autofluorescence spectrum of Eg is wide ,mainly in 490‐520 nm and 610‐630 nm .The better excitation wave lengths are 405 nm ,488 nm and 561 nm .

Expression of tumor CD90 in hepatocellular carcinoma and its relationship with epithelial mesenchymal transition / 中华消化杂志

Objective To investigate the expression of stem cell associated protein CD90 in primary hepatocellular carcinoma (HCC) and its relationship with clinical pathological characters ;to analyze the relation between CD90 and epithelial mesenchymal transition (EMT) markers E‐cadherin (E‐cad) and N‐cadherin (N‐cad) .Methods The expressions of CD90 ,E‐cad and N‐cad in 53 tissues of HCC were detected by immunohistochemistry streptavidin‐perosidase (SP ) method , and 10 surrounding of liver benign lesions were studied as control .The expression of E‐cad and N‐cad in CD90 positive cells of two HCC cell lines (M HCC97‐L and HCCLM‐3 ) was measured by flow cytometry .Chi square test and Spearman rank correlation analysis were performed for count data .Independent sample t test was used for measurement data comparison .Results There was no CD90 expression in control liver tissue . The positive rate of CD90 expression in HCC tissue was (34% ,18/53) ,and the difference between two groups was statistically significant (χ2 = 4 .755 , P< 0 .05) .The expression of CD90 in HCC was positively correlated with portal vein cancer embolus (r= 0 .378 , P< 0 .05) and was negatively correlated with histological differentiation degree and capsular infiltration (r= -0 .398 and -0 .519 ,both P<0 .05) .The expression of CD90 was negatively correlated with the expression of E‐cad (r= -0 .341 , P<0 .05) ,however was positively correlated with the expression of N‐cad (r=0 .441 ,P<0 .05) .The positive expression rate of E‐cad in CD90 positive HCCLM‐3 cells was lower than that of MHCC97‐L cells ((2 .56 ± 0 .29)% and (4 .31 ± 0 .18)% ,t= 8 .757 ,P< 0 .05) ,while the positive expression rate of N‐cad was higher than that of MHCC97‐L cells ((8 .10 ± 1 .45)% and (5 .51 ± 0 .44)% ,t= -9 .667 ,P<0 .05) .Conclusions The expression of stem cell associated protein CD90 is correlated with the EMT of HCC .Their interaction promote tumor infiltration metastasis w hich may be a new target of HCC treatment .

The Role of CXCR7 on Proliferation, Adhesion and Invasion of Gastric Cancer / 天津医药

Objective To investigate the expression of CXCR7 in gastric carcinoma tissues and the role of CXCR7 in tumor growth, adhesion and invasion of gastric cancer. Methods The expression levels of CXCR7, Survivin, CD44v6 and matrix metalloproteinase (MMP)-3 were detected by immunnohistochemistry method in gastric cancer specimens came from 160 patients (gastric cancer group) and 30 specimens of normal gastric tissues (control group). Results The expression of CXCR7 protein was significantly higher in gastric cancer group than that of control group (P<0.05). The positive CXCR7 staining was significantly different with tumor size, depth of invasion, lymph node metastasis and clinical staging ( P<0.05). CXCR7 expression was positively correlated with Survivin, CD44v6 and MMP-3. Conclusion CXCR7 might take part in progression of gastric cancer by anti-apoptosis (Survivin). CXCR7 might take part in the lymph node metastasis of gastric cancer and mediate the adhesion and invasion by CD44v6 and MMP-3.

The expression of TLR2,4,7 mRNA on peripheral blood mononuclear cells in patients with chronic cystic echinococcosis infection / 中华微生物学和免疫学杂志

Objective To explore the expression of TLR2,4,7 mRNA on peripheral blood mono-nuclear cells (PBMCs) in patients with chronic cystic echinococcosis(CE) infection, and the level of serum IL-10. Methods The expression level of TLR2,4,7 mRNA on peripheral blood mononuclear were tested in 42 chronic CE cases and 28 normal controls (NC) by real-time fluorescent quantitative reverse transcription polymerase chain reaction (FQ-RT-PCR) method. GAPDH was selected as the internal control. The level of serum IL-10 was determined in ELISA. The subjects were determined by t test. The correlations between TLR2, TLR4, TLR7 and IL-10 were determined by differences of expression of TLR2, TLR4, TLR7 on PBMCs and serum IL-10 in two groups of study linear correlation test. Results The expressions of TLR2, TLR4,TLR7 mRNA in chronic CE group were higher than those of in NC group. Compared with the NC group, the expressions of TLR2, TLR4 and TLR7 mRNA increased more than 7.3-, 3.6-, 3.6-fold, respectively. In chronic CE group, TLR2, TLR4 and TLR7 mRNA expressions were 1.0729 ±0.4006, 5.0976 ±1.6682, 0. 6481 ±0. 2574, respectively. TLR2, TLR4 and TLR7 mRNA expressions were 0. 1468 ± 0.0435, 1.4067 ±0. 3279, 0. 1804 ±0. 0568 in NC group, respectively. Compared with NC group, the differences of TLR2 and TLR4 mRNA expression were significant (P = 0.0287, 0. 033), while the expression of TLR7 mRNA was not difference (P =0.0862). Moreover, in chronic CE group, the level of serum IL-10 was higher than that of in NC group. In chronic CE group and NC group, the level of serum IL-10 was (17.6770±1.6298) pg/ml, (9.4898 ±0.7049) pg/ml. Compared with NC group, there was significant difference in chronic group (P<0.01). Significant positive correlation between TLR2 and TLR4 was found in chronic CE group, r = 0. 1135, P =0.036. Others were not correlations. Conclusion In the development of chronic CE, TLR2 and TLR4 participate in this progression. As the receptors of antigen of cystic echinococcus, TLR2 and TLR4 can regulate the immune response through interacting with different antigens from cystic echinococcus. Meanwhile, under the participation of TLR2, TLR4 and increased serum IL-10, they will approach to Th2 immune reaction, which play an important role in chronic CE that can induce immune evasion.

Diagnosis of small focal nodular lesions in patients with liver cirrhosis: comparison between contrast-enhanced ultrasound and contrast-enhanced helical CT / 中华超声影像学杂志

Objective To compare the diagnostic value of contrast-enhanced ultrasound (CEUS) and contrast-enhanced helical CT (CECT) for various small focal nodular lesions (≤2 cm) in patients with liver cirrhosis. Methods Eighty-one small hepatic space-occupying lesions in 72 patients with liver cirrhosis were detected with CEUS and CECT, respectively. The diagnostic performance was calculated by histological results obtained from biopsy or surgery, which was considered as the gold standard, Results Fifty-three of the 81 small nodules were hepatocellular carcinoma, 26 were regenerative nodules and 2 were hemangioma. On CEUS, 51 (96.2%,51/53) HCC were hypervascular during arterial phase. On CECT, 41 (77.4%, 41/53) HCC were hypervascular (P < 0.01).Nodules which appeared by contrast enhancement during the arterial phase and contrast wash-out during the portal/late phase on CEUS or CECT were considered as HCC. The sensitivity, specificity and accuracy were 86.8% (46/53) ,82.1% (23/28) ,and 85.2 % (69/81) in CEUS, and 73.6% (39/53), 92.9 % (26/28), and 80.2 % (65/81) in CECT, respectively. Overall, there was no significant difference between CEUS and CECT in the diagnostic confidence for small hepatic nodules (P >0.05).Conclusions CEUS is superior to CECT in the detection of arterial vascularization for small hepatocellular carcinoma with a diameter ≤2 cm. The ability of CEUS in the characterization of focal nodular lesions in cirrhotic livers is similar to that of CECT.

RNAi-mediated Stable Silencing of HGPRT Expression in Rabbit Fibroblasts and SCNT Embryo Production / 生物化学与生物物理进展

The hypoxanthine-guanine phosphoribosyl transferase (HGPRT) gene mutation is responsible for gouty arthritis, kidney stone, and Lesch-Nyhan Syndrome (LNS). It has been reported that the expression of HGPRT is decreased or even absent in these diseases. Rabbits are an ideal model for studying the pathology of these diseases. Therefore, the development of an HGPRT-knockdown rabbit model will be highly beneficial m such studies. Stable HGPRT-knoekdown transgenie fibroblast lines were generated by transfecting rabbit fibroblasts with RNA interference (RNAi) plasmids. Polymerase chain reaction (PCR) analyses indicated that the average positive rate was 83.3%. The mRNA and protein levels of HGPRT in the transgenic fibroblast lines were significantly lower than that in the control. Transgenic rabbit blastocysts were derived after performing nuclear transfer. The results show that RNAi can be used to stably knock down expression of the HGPRT in rabbit fibroblasts and further improvements in related technologies will facilitate the use of this method for the generation of HGPRT-knockdown rabbits.