ABSTRACT
Objective: To evaluate the anti-tumor activity of mouse multi-subtype heat shock protein/peptide (mHSP/P) vaccine in combination with a programmed death ligand 1 (PD-L1) inhibitor in mouse sarcoma. Methods: Immunohistochemical staining and en-zyme-linked immunosorbent assay (Elisa) was used to quantitatively identify the expression of heat shock proteins (HSP70, HSP90, Grp94) in the sarcoma cell line MCA207. From the protein suspension prepared, mHSP/P and Grp94/peptide (Grp94/P) sarcoma vac-cines were isolated using chromatography and were identified by Western blot (WB). Flow cytometry was used to determine their cy-totoxic effects. The levels of interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) produced upon mHSP/P and Grp94/P stimulation were measured by Elisa. The effect of sarcoma vaccines on the growth and survival of sarcoma was evaluated in mice. The expression of PD-L1 on the surface of MCA207 sarcoma cells was evaluated by immunofluorescent staining. The effect of IFN-γ treatment on the expression of PD-L1 was determined by WB. Animal experiments explored the effects of PD-L1 inhibitor in combination with mHSP/P treatment on tumors. Results: Tumor tissue carries a variety of HSP subtypes (HSP70, HSP90, Grp94). We successfully isolated sarco-ma tissue-derived mHSP/P and Grp94/P tumor vaccines, which were identified by WB; flow cytometry analysis demonstrated their cy-totoxicity. The levels of IFN-γ and TNF-α cytokines upon mHSP/P stimulation were significantly higher than that observed upon Grp94/P stimulation (P<0.05). The expression of PD-L1 on the surface of sarcoma cells increased with IFN-γ treatment. Animal experiments demonstrated that PD-L1 inhibitor in combination with mHSP/P significantly increased the immune response against tumor (P<0.05). Conclusions: Tumor-derived mHSP/P and Grp94/P can be used as tumor vaccines in animal models. The mHSP/P can elicit a stronger anti-tumor immune response than Grp94/P. IFN-γ stimulates the expression of PD-L1 in sarcoma cells, which results in immune eva-sion. The PD-L1 inhibitor in combination with mHSP/P increased the anti-tumor effect in the tumor microenvironment.
ABSTRACT
Objective To discussion the value of palate standard section in prenatal ultrasound diagnosis of isolate cleft palate,and summarize the two-dimensional ultrasound characteristics of isolate cleft palate.Methods Two-dimensional ultrasound was performed in 1 8 073 fetuses during 1 8 to 40 gestational weeks of pregnancy.Fetal palate structure were scanned through the oral cleft,the cheek,the neck part,and the standard sagittal section,transverse section and coronal section were obtained to diagnosis cleft palate. The results were compared with labour or postpartum findings.Results In 1 8 073 cases,20 cases of isolate cleft palate,1 9 cases were diagnosed as fetal isolate cleft palate by prenatal ultrasonography,of which 1 8 cases were correctly diagnosed,1 case was misdiagnosed,1 case was missed diagnosed.The sensitivity, specificity,accuracy of prenatal ultrasound detection were 95%,100%,95%,respectively.The ultrasonic characteristics of isolate cleft palate:lack of crack were located in the midline,soft cleft palate was shown as the midline of soft palate interrupted,short of crack was“><”,“‖”,“/\”shape,hard cleft palate was shown as the midline of hard palate interrupted,short of crack was open back“V”or“U”shape.Conclusions The palate standard section has high value in prenatal ultrasound diagnosis of cleft palate,which can assess fracture morphology,length,width,direction and involving range.
ABSTRACT
@#Objective To investigate the antitumor immunity induced by tumor derived mixed heat shock protein/peptide(mHSPs),interleukin-12(IL-12)and Cyclophosphamide(Cy).MethodsPurified mixed HSP was prepared from tumor by S180 protein extraction and purification,SDS-PAGE,Western blot and animal experiment were applied for mixed HSPs analysis.ResultsThe proliferation of cytotoxic T lymphocyte(CTL)cocultured in the mHSPs+Cy+IL-12 group was significantly remarkable and the content of CD8+ CTLs was significantly more in comparison with the other groups(P<0.01).To the tumor bearing mice,mHSPs+Cy+IL-12 group showed partial therapeutic efficacy,the averaged survival period was over 60 d,and 90% of the mice in this group got long period tumor free survival(>90d),obvious difference(P<0.05)from the other groups.ConclusionTumor derived mixed HSPs can induce powerful antitumor immune efficacy and show favorable therapeutic efficacy.
ABSTRACT
BACKGROUND: Elimination of antigenic substances from natural extracellular matrix with the integrity of the tissue structure retained renders the matrix to possess better biocompatibility and provides a cell culture environment close to conditions of the internal environment. Such materials are the primary choice for cell culture scaffold in tissue engineering.OBJECTIVE: To prepare human articular cartilage acellular matrix so as to provide a methodological basis for further study of articular cartilage acellular matrix as cell scaffold materials.DESIGN: A single sample study of bone tissues.SETTING: The experiment was performed in Institute of Orthopedics, General Hospital of PLA, between January and May in 2004. The specimens were obtained from patients requiring joint replacement for femoral neck fracture.MATERIAIS: The experiment was conducted in the Department of Orthopedics, General Hospital of PLA from January to May in 2004. Human articular cartilage specimens were obtained from the femoral head of patients with total hip arthroplasty for femoral neck fracture.METHODS: Totally 10 specimens of fresh articular cartilage(3.5 mm × 4. 5 mm × 2.0 mm) were obtained and freeze-dried for 12 hours. Cartilage acellular matrix was prepared using Triton X-100, Dnase and Rnase and identified by means of hematoxylin-eosin(HE) and safranine O staining and immunohistochemical staining for cartilage proteoglycan.MAIN OUTCOME MEASURES: Histological observation of the articular cartilage acellular matrix and immunohistochemical staining of cartilage proteoglycan.RESULTS: HE and safranine O staining both showed no cellular structure in the matrix with only recesses left by the removed cells. Immunohistochemical staining for cartilage proteoglycan yielded positive results, suggesting the presence of cartilage proteoglycan in the acellular matrix.CONCLUSION: Human articular cartilage acellular matrix can be prepared using the modified four-step procedures with detergent and enzymatic extraction with lyophilization, and the preserved cartilage proteoglycan in the material may retain good pressure resistance.
ABSTRACT
<p><b>OBJECTIVE</b>To search novel method for diagnosis and therapy of B-lymphoma, specific small molecular peptide ligands against binding site of tumor cells were screened and its effects on signal transduction and cell apoptosis were tested.</p><p><b>METHODS</b>Specific peptide ligands were screened by binding with site of human B lymphoma cell (OC1LY8) using peptide-bead libraries. The identified peptides were characterized with responsible cells by rebinding test. The role of tyrosine phosphorylation of peptide ligand was tested by Western blot; and its apoptosispromoting role was observed by confocal fluorescent microscope.</p><p><b>RESULTS</b>Specific peptide ligand was able to bind specifically to site on cell surface and enter into cytoplasm. Tetrameric peptide ligand was able to strongly trigger signal transduction resulting in tyrosine phosphorylation and cellular apoptosis in OC1LY8 cell line.</p><p><b>CONCLUSION</b>Screened peptide ligand can effectively bind with OC1LY8 cell, stimulate cellular tyrosine phosphorylation and induce cellular apoptosis.</p>
Subject(s)
Humans , Amino Acid Sequence , Apoptosis , Cell Line, Tumor , Ligands , Lymphoma, B-Cell , Metabolism , Pathology , Oligopeptides , Chemistry , Pharmacology , Peptide Library , Phosphorylation , Signal Transduction , Tyrosine , MetabolismABSTRACT
e: Objective:To establish a useable animal model for the purification and immunotherapy of HSP with the observation of the growth rule of S180 sarcome tumor tissue and the immunohistochemical expression of different HSP in mice. Methods:The tumor incidence?the role of growth?the survival time and immunohistochemical expression of different HSP were observed after the S180 sarcome cells were inoculated at Balb/C mice back subcutaneous in different dose. Results:Sarcoma were formed in 100%.There are significant difference between vary dose in survival time, The expression was positive in HSP60?70?90? and grp94, and HSP70 expression was enhanced whereas HSP90? expression was decreased after heat shock treated. Conclusion:The use of S180 sarcome in mice is a good model to observe the expression of different HSP and the tumor tissue is suitable to purify. The model can be also used in the library study of HSP tumor vaccine.
ABSTRACT
<p><b>OBJECTIVE</b>To assess the effects of nuclear translocation of tissue transglutaminase (TTG) and the release of cytochrome C on hepatocyte apoptosis and to reveal the mechanism of signal transduction of early apoptosis in injured hepatocytes.</p><p><b>METHODS</b>Hepatocytes isolated from tissue transglutaminase gene knock-out mice and rats were stimulated with ethanol. Proteins from whole cell, cytoplasm and nuclei were extracted for determination of TTG activity by (14)C-putrescine incorporation. Distribution of TTG throughout the entire cell, as well as just nucleus was observed under a confocal scanning microscope. The amount of cytochrome C released from mitochondria was determined by ELISA. Cell apoptosis was observed by fluorescent cytochemistry.</p><p><b>RESULTS</b>TTG activity in whole cells and nuclei was significantly increased after the hepatocytes were treated with ethanol. Cytochrome C release was remarkably increased in the cells isolated from rat and wild-type mouse after treatment with ethanol but not in TTG gene knock-out mice. Cellular apoptosis appeared in hepatocytes isolated from rats and wild-type mice but not in the hepatocytes from TTG gene knock-out mice after stimulation with ethanol.</p><p><b>CONCLUSIONS</b>Increased TTG in hepatocytes can be translocated into the nucleus and promote release of mitochondrial cytochrome C into the cytoplasm. Passing through a series of signal pathways, hepatocyte apoptosis is induced eventually.</p>
Subject(s)
Animals , Male , Mice , Rats , Apoptosis , Cell Nucleus , Metabolism , Cytochrome c Group , Metabolism , Hepatocytes , Cell Biology , Metabolism , Signal Transduction , Transglutaminases , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To investigate the role of apoptosis in radiation-induced mouse thymus lymphocyte damage and repair and provide the basis for understanding the molecular mechanism of radiation-induced lymphocyte damage and repair as well as the prevention and treatment of acute radiation sickness.</p><p><b>METHODS</b>We studied the dynamic changes of apoptosis of mouse thymus lymphocytes and the expression of bax and bcl-2 gene products after 2, 4, 6 and 8 Gy of whole body gamma-irradiation using in situ terminal labeling, DNA electrophoresis and immunohistochemical techniques.</p><p><b>RESULTS</b>At the early stage after irradiation, the percentage of apoptotic lymphocytes increased rapidly in accordance with the increasing of radiation doses, while the counts of the thymus and peripheral lymphocytes decreased sharply, showing an opposite change to lymphocyte apoptosis. After 6 Gy gamma-irradiation, typical morphological characteristics of thymus apoptotic lymphocytes in early, middle and late stages were found by transmission electron microscopy. The thymus lymphocytes displayed characteristic DNA ladders 4 hr and 8 hr after 2-6 Gy gamma-irradiation,using DNA gel electrophoresis techniques. Abnormal expression of bcl-2 and bax gene products were shown in irradiated lymphocytes.</p><p><b>CONCLUSIONS</b>Apoptosis plays an important role in the process of radiation-induced mouse thymus lymphocyte damage and repair. Bcl-2 and Bax proteins may regulate the process of lymphocyte apoptosis.</p>
Subject(s)
Animals , Male , Mice , Apoptosis , Radiation Effects , Dose-Response Relationship, Radiation , Gamma Rays , Lymphocytes , Physiology , Radiation Effects , Proto-Oncogene Proteins , Proto-Oncogene Proteins c-bcl-2 , Thymus Gland , Pathology , Radiation Effects , Time Factors , bcl-2-Associated X ProteinABSTRACT
AIM:To examine the role and dynamic changes of mast cell(MC) and its subpopulations in simple and irradiated-wound of rats.METHODS:MC and its subpopulations were estimated using alcian blue-safranin (ABS)double staining RESULTS:(1)The total number of MC in two groups decreased coincidently on day 2 after wounding, and the total MC increased rapidly and reached maximal gradually on day 5 and 7 after wounding, the increment of MC remained consistently 28 day after wounding (2)Both mucosal MC( MMC) and Mix MC decreased obviously on day 2 after wounding, hereafter,they remained the low level all the time However, the CTMC kept in the high level after wounding (3)The Mix MC on day 5 and the total MC during day 5-15 after wounding were lower in irradiated group than in simple wound group CONCLUSION: MC and its subpopulations could delay the healing process of simple and irradiated wound