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ObjectiveThis study aims to investigate the effect of modified Baitouwengtang （MBTWD） on tumor growth and the number of tumor-associated macrophages （TAMs） in tumor tissue of MC38 cell tumor-bearing mice with colorectal cancer and explores whether MBTWD mediates the remodeling of TAM phenotype to play an immunologically antitumor effect. MethodFirstly， The C57BL/6 mouse tumor model grafted subcutaneously was established， and then model mice were classified into a model group， positive control group（3 mg·kg-1）， and MBTWD groups with high and low dosages（23.43、46.86 g·kg-1）， with 10 mice in each group. In addition， 10 healthy mice were set as the blank group， and the changes in body weight， tumor volume， and survival status of mice in each group were observed. Tumor tissue， spleen， and peripheral blood were collected to calculate the tumor volume change， tumor inhibition rate， and spleen mass. Hematoxylin-eosin （HE） staining was used to observe the morphological changes of tumor tissue， and an immunofluorescence assay was used to detect the expression levels of CD4， CD8， and CD206 in tumor tissues of tumor-bearing mice. The secretion levels of transforming growth factor （TGF）-β， interleukin （IL）-6， and chemokine （C-C Motif） ligand 2 （CCL2） in peripheral serum were measured by using enzyme-linked immunosorbent assay （ELISA）. Secondly， a co-culture model induced by IL-4 in vitro of MC38 cells and murine monocytic macrophage RAW264.7 cells was established. Cell proliferation and activity assay （CCK-8） was used to detect the inhibitory effect of MBTWD containing serum on cell proliferation. A transwell experiment was used to detect the effect of IL-4-induced M2 macrophages on the invasion of MC38 cells. Flow cytometry was used to detect the expression of CD86 on the membrane of M2 macrophages induced by IL-4 with MBTWD containing serum. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was used to detect the effect of MBTWD containing serum on the mRNA expression levels of M1 macrophage-related polarization factors CD86， nitric oxide synthase （iNOS）， and IL-12， as well as M2 macrophage-related polarization factors CD206， CD163， and IL-10 after co-cultivation. Finally， the protein expression levels of colony-stimulating factor 1 receptor （CSF1R）， stimulator of interferon genes （STING）， and TANK binding kinase 1 （TBK1） in tumor tissues of tumor-bearing mice were detected by Western blot. ResultIn vivo experimental results show that compared with the model group， the MBTWD can significantly inhibit the tumor growth of tumor-bearing mice. Immunofluorescence experiments show that the MBTWD can increase the number of CD8+ T cell infiltration in tumor tissue of tumor-bearing mice， reduce the number of CD206+ TAMs infiltration， and down-regulate the secretion levels of cytokines IL-6， TGF-β， and CCL2 in peripheral blood of tumor-bearing mice. The results of in vitro experiments show that the MBTWD containing serum has no obvious inhibitory effect on cell proliferation， but the cell supernatant after co-cultivation with RAW264.7 cells can inhibit the proliferation activity of MC38 cells， and the invasion ability of MC38 cells is enhanced by IL-4-induced M2 macrophages. However， this effect can be inhibited in a concentration-dependent manner by the MBTWD containing serum. At the same time， the results of Real-time PCR show that the MBTWD containing serum can up-regulate the mRNA expression levels of M1 macrophage-related polarization factors CD86， iNOS， and IL-12 and down-regulate those of M2 macrophage-related polarization factors CD206， CD163， and IL-10. Flow cytometry results also confirm that the MBTWD containing serum can increase the number of repolarized CD86+ M1 macrophages， indicating that MBTWD can induce M2 macrophages to repolarized M1 macrophages to play an anti-tumor growth role. Finally， Western blot results show that MBTWD can down-regulate the expression of CSF1R protein and up-regulate that of STING and TBK1 proteins in tumor tissue of tumor-bearing mice. ConclusionMBTWD can down-regulate the infiltration number of CD206+ TAMs and increase the infiltration of CD8+ T cells， thereby playing an immunologically antitumor effect on the growth inhibition of colorectal cancer， which may be related to regulating CSF1R signaling and then activating STING/TBK1 signaling pathway to induce phenotypic remodeling of TAMs.
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The acupuncture method of Shugan-Tiaoshen is proposed by the famous old Chinese medicine Professor Shan Qiuhua in Shandong Province in combination with Traditional Chinese Medicine theory and clinical experience. Shugan-Tiaoshen acupuncture method is widely used in clinic and has curative effect. It can treat mental diseases, nervous system diseases, digestive system diseases, endocrine system diseases, urinary system diseases, diseases caused by physical and chemical factors and other internal diseases, as well as surgical diseases such as skin trauma, facial symptoms and breast diseases. It can also treat gynecological diseases. It can also improve the brain function of stress disorder. The role of soothing liver and regulating spirit in clinic is never underestimated. In clinical practice, doctors should try their best to combine regulating spirit with treating spirit. Shugan-Tiaoshen acupuncture provides a good idea for clinical treatment, but there are still some problems in the current researches, such as the lack of multi-center, large sample trials, or well-design mechanism researches.
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Objective@#To search for biomarkers for human familial prion disease.@*Methods@#Two-dimensional differential gel electrophoresis (2D-DIGE) proteomic analysis has been performed in frontal lobe tissues of 3 patients suffering from human familial prion disease (PrP) and 3 age-and sex-matched patients suffering from sudden death due to heart failure without neurological disease.@*Results@#The maps revealed 14 polypeptide chains differentially modulated in the PrP samples, among those, 7 could be identified upon digestion and MALDI-TOF/MS analysis, of which 6 appeared to be up-regulated, 1 being down-regulated.@*Conclusions@#We highlight Galectin-1(Gal-1), ryanodine receptor 2 (RyR2), ubiquitin, Rab-interacting lysosomes protein-like protein 1 (RILPL-1) profillin 2 (PFN2), in the differential map. These proteins are related to neurogenesis, the clearance of misfolded proteins, stasis of calium channel, myoclonus and so on. These proteins are potential biomarkers or targets for treatment of prion disease.
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Objective To investigate the effect of stress-related hyperglycemia on myocardial function of patients with cardiac disorder after non-cardiac surgery.Methods The clinical data of 211 patients having undergone thoracic or abdominal operations in Department of Critical Care Medicine of Sun Yat-sen University Cancer Center were retrospectively analyzed. According to the postoperative average blood glucose level in the following 3 days after surgery and the patients' history of cardiac disorder, they were divided into four groups: without hyperglycemia (blood glucose ≤ 10 mmol/L) and cardiac disorder group (HG0CV0 group), without hyperglycemia but with cardiac disorder group (HG0CV1 group), with hyperglycemia (blood glucose > 10 mmol/L) but without cardiac disorder group (HG1CV0 group) and with hyperglycemia and cardiac disorder group (HG1CV1 group). The correlations between the blood glucose and each level of the following items: high-sensitivity cardiac troponin I (hs-cTnI), brain natriuretic peptide (BNP), myocardial zymogram aspartate aminotransferase (AST), creatine kinase (CK), MB isoenzyme of creatine kinase (CK-MB), lactic dehydrogenase (LDH), lactic dehydrogenase isoenzyme (LDH-1), myoglobin (MYO), α- hydroxybutyrate dehydrogenase (HBDH) in each group were analyzed.Results The postoperative blood glucose levels of all 211 patients were significantly higher than those before operation (mmol/L: 8.7±0.2 vs. 5.7±0.2,P 0.05). In HG0CV1 group, the postoperative blood glucose level was negatively correlated with hs-cTnI level (r = -0.609, 95%CI = -0.810 to -0.264,P = 0.001). There were no correlations between postoperative blood glucose level and the levels of BNP, MYO, AST, LDH, LDH-1, HBDH, CK and CK-MB (allP > 0.05). In HG1CV0 group, there were no correlations between postoperative blood glucoselevel and hs-cTnI, BNP, MYO, AST, LDH, LDH-1, HBDH, CK and CK-MB (allP > 0.05). In HG1CV1 group, the postoperative blood glucose level was positively correlated with hs-TnI level (r = 0.837, 95%CI = 0.476 - 0.984,P = 0.001). There were no correlations between postoperative blood glucose level and the levels of BNP, MYO, AST, LDH, LDH-1, HBDH, CK and CK-MB (allP > 0.05).Conclusion The early stress-related hyperglycemia after non-cardiac surgery may have a protective effect on myocardial function of patients with cardiac disorder.
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<p><b>OBJECTIVE</b>Fatal familial insomnia (FFI) is an autosomal dominant prion disease characterized clinically by inattention, sleep loss, dysautonomia, and motor signs. This study is aimed to investigate clinical and familial characteristics of ten Chinese Patients with FFI.</p><p><b>METHODS</b>We identified ten FFI cases from the surveillance network for Creutafeldt-Jakob disease (CJD) in China. Final diagnosis of FFI cases was made in accordance with the WHO criteria for CJD. The main clinical features and family histories of these ten FFI cases were analyzed.</p><p><b>RESULTS</b>The median age of ten cases at onset was 38 years (from 19 to 55). The foremost symptoms seemed to be various, including sleep disturbances, vision disorder, dizziness and anorexia. Sleep disturbances appeared in all cases and lasted in the whole clinical courses. Progressive sympathetic symptoms, memory loss, movement disturbances, myoclonus and hypertension were also frequently observed. The median duration of the disease was 9.5 months. EEG and MRI did not figure out special abnormality. 14-3-3 protein in CSF was positive in five out of eight tested patients. Clear family histories were identified in 8 patients.</p><p><b>CONCLUSION</b>The data from our study confirm that the Chinese FFI cases have similar clinical characteristics as that of the Caucasian cases. Compared with other genetic CJD associated mutations, the genetic frequencies of D178N in PRNP are apparently high among the Chinese cases.</p>
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Adult , Female , Humans , Male , Middle Aged , Young Adult , Asian People , Insomnia, Fatal Familial , PathologyABSTRACT
<p><b>OBJECTIVE</b>To break immune tolerance to prion (PrP) proteins using DNA vaccines.</p><p><b>METHODS</b>Four different human prion DNA vaccine candidates were constructed based on the pcDNA3.1 vector: PrP-WT expressing wild-type PrP, Ubiq-PrP expressing PrP fused to ubiquitin, PrP-LII expressing PrP fused to the lysosomal integral membrane protein type II lysosome-targeting signal, and PrP-ER expressing PrP locating the ER. Using a prime-boost strategy, three-doses of DNA vaccine were injected intramuscularly into Balb/c mice, followed by two doses of PrP protein. Two weeks after the last immunization, sera and spleens were collected and PrP-specific humoral and cellular immune responses evaluated by ELISA and ELISPOT tests.</p><p><b>RESULTS</b>Higher levels of serum PrP antibodies were detected in mice vaccinated using the strategy of DNA priming followed by protein boosting. Of these, WT-PrP, Ubiq-PrP, and PrP-LII induced significantly higher humoral responses. ELISPOT tests showed markedly increased numbers of IFN-γ-secreting T cells in mice vaccinated using the strategy of DNA priming followed by protein boosting after stimulation with recombinant PrP23-90 and PrP23-231. PrP-ER induced the strongest T-cell response.</p><p><b>CONCLUSION</b>Prion vaccines can break tolerance to PrP proteins and induce PrP-specific humoral and cellular immune responses.</p>
Subject(s)
Animals , Cricetinae , Female , Humans , Mice , Antibodies , Allergy and Immunology , CHO Cells , COS Cells , Chlorocebus aethiops , Cricetulus , Enzyme-Linked Immunosorbent Assay , HeLa Cells , Immune Tolerance , Interferon-gamma , Allergy and Immunology , Lysosomal-Associated Membrane Protein 2 , Genetics , Allergy and Immunology , Mice, Inbred BALB C , Peptide Fragments , Allergy and Immunology , Prions , Genetics , Allergy and Immunology , Receptors, Peptide , Genetics , Allergy and Immunology , Recombinant Fusion Proteins , Genetics , Allergy and Immunology , Recombinant Proteins , Allergy and Immunology , Transfection , Ubiquitin , Genetics , Allergy and Immunology , Vaccines, DNAABSTRACT
<p><b>BACKGROUND</b>An amino acid polymorphism for Met to Val has been identified at PrP codon 129 from different human races. In this study,the characteristics of polymorphism of PRNP 129th amino acid in Han and Uighur Chinese have been investigated.</p><p><b>METHODS</b>Human DNAs were extracted from peripheral lymphocytes and PrP gene fragments were amplified with a specific PCR protocol. The distribution of 129th amino acid in PRNP was determined by a PCR-RFLP and the results were analyzed with software SAS for Windows 6.12.</p><p><b>RESULTS</b>The frequencies of the allele 129 Met and 129 Val were 97.0% and 3.0% in Han Chinese, whereas 91.4% and 8.6% in Uighur Chinese. The frequency of 129 M/M phenotypes in Han Chinese was significantly higher than that in Uighur Chinese (P=0.0490). Comparing the phenotype distributions of codon 129 of Han Chinese with that of Japanese and Caucasian, there was significant difference with Caucasian (P=0.0005),but there was no difference with Japanese (P=0.5040).</p><p><b>CONCLUSIONS</b>The polymorphism of 129th amino acid in PRNP of Han Chinese is similar to Japanese, but different from Uighur Chinese.</p>