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1.
Journal of Experimental Hematology ; (6): 1522-1527, 2021.
Article in Chinese | WPRIM | ID: wpr-922289

ABSTRACT

OBJECTIVE@#To investigate the expression of cell division cycle protein 37 (Cdc37) in multiple myeloma (MM) and its effect on MM cell proliferation.@*METHODS@#The expression of Cdc37 mRNA in CD138@*RESULTS@#Cdc37 was highly expressed in newly diagnosed CD138@*CONCLUSION@#Cdc37 is highly expressed in newly diagnosed MM patients. Inhibition of Cdc37 results in decreased proliferation activity and G


Subject(s)
Animals , Apoptosis , Cell Cycle Proteins , Cell Proliferation , Chaperonins , Humans , Mice , Mice, Inbred NOD , Mice, SCID , Multiple Myeloma
2.
Journal of Experimental Hematology ; (6): 1429-1435, 2021.
Article in Chinese | WPRIM | ID: wpr-922276

ABSTRACT

OBJECTIVE@#To establish the in vivo traceable acute myeloid leukemia mice model with Luciferase-Expressing KG1a Cells.@*METHODS@#KG1a cells with stable luciferase gene expression (called as KG1a-Luc cells) were constructed by lentivirus transfection, then sifted out by puromycin. Eighteen male NOD-SCID-IL2rg@*RESULTS@#KG1a cells expressing luciferase stably were successfully obtained. The tumor luminescence wildly spread at day 17 captured by in vivo imaging. The KG1a-Luc tumor cells could be detected in the peripheral blood of the mice, with the average percentage of (16.27±6.66)%. The morphology and pathology result showed that KG1a-Luc cells infiltrate was detected in bone marrow, spleens and livers. The survival time of the KG1a-Luc mice was notably shorter as compared with those in the control group, the median survival time was 30.5 days (95%CI: 0.008-0.260).@*CONCLUSION@#The acute myeloid leukemia NOD-SCID-IL2rg


Subject(s)
Animals , Disease Models, Animal , Interleukin Receptor Common gamma Subunit , Leukemia, Myeloid, Acute , Luciferases/genetics , Male , Mice , Mice, Inbred NOD , Mice, SCID
3.
Biol. Res ; 53: 13, 2020. tab, graf
Article in English | LILACS | ID: biblio-1100919

ABSTRACT

BACKGROUND: Gallbladder cancer (GBC) is the most common tumor of the biliary tract. The incidence of GBC shows a large geographic variability, being particularly frequent in Native American populations. In Chile, GBC represents the second cause of cancer-related death among women. We describe here the establishment of three novel cell lines derived from the ascitic fluid of a Chilean GBC patient, who presented 46% European, 36% Mapuche, 12% Aymara and 6% African ancestry. RESULTS: After immunocytochemical staining of the primary cell culture, we isolated and comprehensively characterized three independent clones (PUC-GBC1, PUC-GBC2 and PUC-GBC3) by short tandem repeat DNA profiling and RNA sequencing as well as karyotype, doubling time, chemosensitivity, in vitro migration capability and in vivo tumorigenicity assay. Primary culture cells showed high expression of CK7, CK19, CA 19-9, MUC1 and MUC16, and negative expression of mesothelial markers. The three isolated clones displayed an epithelial phenotype and an abnormal structure and number of chromosomes. RNA sequencing confirmed the increased expression of cytokeratin and mucin genes, and also of TP53 and ERBB2 with some differences among the three cells lines, and revealed a novel exonic mutation in NF1. The PUC-GBC3 clone was the most aggressive according to histopathological features and the tumorigenic capacity in NSG mice. CONCLUSIONS: The first cell lines established from a Chilean GBC patient represent a new model for studying GBC in patients of Native American descent.


Subject(s)
Humans , Animals , Male , Middle Aged , Antigens, Tumor-Associated, Carbohydrate/genetics , Indians, South American/genetics , Gallbladder Neoplasms/genetics , Ascitic Fluid/metabolism , Tumor Cells, Cultured , Carcinogenicity Tests , Chile , DNA Fingerprinting , Tumor Suppressor Protein p53/genetics , Cisplatin/pharmacology , Mice, Inbred NOD , Clone Cells/drug effects , Clone Cells/metabolism , Sequence Analysis, RNA , Receptor, ErbB-2/genetics , Genes, erbB-2/genetics , Gene Expression Profiling , Cell Line, Tumor/drug effects , Cell Line, Tumor/metabolism , Deoxycytidine/analogs & derivatives , Deoxycytidine/pharmacology , Epithelial Cells/metabolism , Keratin-19/genetics , Keratin-7/genetics , Carcinogenesis/genetics , Gallbladder Neoplasms/metabolism , Antineoplastic Agents/pharmacology
4.
Article in Chinese | WPRIM | ID: wpr-826376

ABSTRACT

To obtain ultrasound and thermal tomography images of breast cancer during its growth and to assess the value of thermal tomography in detecting breast cancer. Breast cancer models were established with NOD/SCID mice and SD rats. These animal models were examined by thermal tomography,plain ultrasound,and contrast-enhanced ultrasound. Tumor tissues were stained with CD34 to explore the relationship between tumor heat production and vascular pathology. Thermal tomography detected breast cancer 2-4 days earlier than ultrasound. The expression of CD34 in tumor tissues was increased,along with thickened,increased,and irregular blood vessels. Thermal tomography can detect early breast cancer and is a promising tool for screening breast cancer.


Subject(s)
Animals , Breast Neoplasms , Diagnostic Imaging , Early Diagnosis , Mice , Mice, Inbred NOD , Mice, SCID , Neoplasms, Experimental , Diagnostic Imaging , Rats , Rats, Sprague-Dawley , Tomography , Ultrasonography, Mammary
5.
Article in Chinese | WPRIM | ID: wpr-880764

ABSTRACT

OBJECTIVE@#To investigate the antitumor effect of ponatinib on the growth of cholangiocarcinoma xenograft derived from a clinical patient in a mouse model expressing FGFR2-CCDC6 fusion protein.@*METHODS@#Lung metastatic tumor tissue was collected from a patient with advanced intrahepatic cholangiocarcinoma and implanted subcutaneously a NOD/SCID/ Il2rg-knockout (NSG) mouse. The tumor tissues were harvested and transplanted in nude mice to establish mouse models bearing patient-derived xenograft (PDX) of cholangiocarcinoma expressing FGFR2-CCDC6 fusion protein. The PDX mouse models were divided into 4 groups for treatment with citrate buffer (control group), intragastric administration of 20 mg/kg ponatinib dissolved in citrate buffer (ponatinib group), weekly intraperitoneal injections of 50 mg/kg gemcitabine and 2.5 mg/ kg cisplatin (gemcitabine group), or ponatinib combined with gemcitabine and cisplatin at the same doses (10 mice in each group, and 9 mice were evaluated in ponatinib group). The expressions of p-FGFR, p-FRS2, p-AKT, p-ERK, CD31, and Ki-67 in the xenografts were evaluated with immunohistochemistry, and cell apoptosis was analyzed with cleaved caspase-3 (CC3) staining and TUNEL staining. Western blotting was used to detect the expressions of FGFR2, p-FGFR, AKT, p-AKT, ERK, p-ERK, FRS2 and p-FRS2 in the tumor tissues.@*RESULTS@#Compared with those in the control group, the mice in ponatinib group showed a significantly reduced tumor volume (@*CONCLUSIONS@#Ponatinib can regulate FGFR signaling to inhibit the proliferation and induce apoptosis of tumor cells in mice bearing patient-derived cholangiocarcinoma xenograft with FGFR2 fusion. FGFR inhibitor can serve as a treatment option for patients with cholangiocarcinoma with FGFR2 fusion.


Subject(s)
Animals , Bile Duct Neoplasms/genetics , Cell Line, Tumor , Cell Proliferation , Cholangiocarcinoma/genetics , Cytoskeletal Proteins , Heterografts , Humans , Imidazoles , Mice , Mice, Inbred NOD , Mice, Nude , Mice, SCID , Pyridazines , Receptor, Fibroblast Growth Factor, Type 2 , Xenograft Model Antitumor Assays
6.
Article in Chinese | WPRIM | ID: wpr-772338

ABSTRACT

BACKGROUND@#Small cell lung cancer (SCLC) is characterized by poor differentiation, high malignancy and rapid growth fast, short double time, early and extensive metastatic malignancy. In clinical, chemotherapy is the main treatment method, while resistance to multiple chemotherapy drugs in six to nine months has been a major clinical challenge in SCLC treatment. Therefore, It has important clinical value to building SCLC aninimal model which is similar to patients with SCLC. Animal model of xenotransplantation (PDX) from the patients with small cell lung cancer can well retain the characteristics of primary tumor and is an ideal preclinical animal model. The study is aimed to establish SCLC PDX animal model and induce the chemoresistance model to help to study the mechanism of chemoresistance and individual treatment.@*METHODS@#Fresh surgical excision or puncture specimens from SCLC patients were transplanted into B-NSGTM mice subcutaneous tissues with severe immunodeficiency in one hour after operation the B-NSGTM mice subcutaneous in 1 hour, and inject chemotherapy drugs intraperitoneally after its tumor growed to 400 mm³ with EP which is cisplatin 8 mg/kg eight days and etoposide 5 mg/kg every two days until 8 cycles. Measure the tumor volum and mice weights regularly, then re-engrafted the largest tumor and continue chemotherapy.@*RESULTS@#Nine cases were conducted for B-NSG mice modeling. Three of nine cases could be engrafted to new B-NSG mice at least two generation. The SCLC PDX animal models have been established successfully. After adopting chemotherapy drugs, the chemoresistance PDX models have been established. High homogeneity was found between xenograft tumor and patient's tumor in histopathology, immunohistochemical phenotype (Syn, CD56, Ki67).@*CONCLUSIONS@#The SCLC PDX animal model and the chemoresistance PDX animal model have been successfully constructed, the success rate is 33%, which provides a platform for the clinical research, seeking for biological markers and choosing individual treatment methods of SCLC.


Subject(s)
Animals , Antineoplastic Combined Chemotherapy Protocols , Pharmacology , Cisplatin , Disease Models, Animal , Drug Resistance, Neoplasm , Etoposide , Female , Humans , Interleukin Receptor Common gamma Subunit , Genetics , Lung Neoplasms , Drug Therapy , Metabolism , Pathology , Mice, Inbred BALB C , Mice, Inbred NOD , Mice, Knockout , Mice, SCID , Small Cell Lung Carcinoma , Drug Therapy , Metabolism , Pathology , Transplantation, Heterologous , Methods , Xenograft Model Antitumor Assays
7.
Journal of Experimental Hematology ; (6): 1522-1529, 2019.
Article in Chinese | WPRIM | ID: wpr-775690

ABSTRACT

OBJECTIVE@#To investigate the tumorigenicity of several multiple myeloma (MM) cell lines transplanted in mice without γ-ray irradiation and to construct the MM disease model to facilitate in vivo experiments.@*METHODS@#NOD/SCID or NSG mice were subcutaneously or caudally transplanted with MM cell lines (LP-1, OPM2, RPMI 8226 and MOLP8), or cell lines with luciferase (RPMI-Luc-Puro, RPMI-Luc-mCherry and MOLP8-Luc-Puro). Tumor growth was observed by measuring the tumor size with a caliper. CD138 tumor cells in peripheral blood were detected by flow cytometry. The free light chain in mouse serum was detected by immunofixation electrophoresis. Tumor type was identified by immunohistochemistry.@*RESULTS@#Twenty one NOD/SCID mice were subcutaneously transplanted with LP-1 cells or OPM2 cells respectively, and no tumors formed till 7 weeks after transplantation. Fifteen NOD/SCID mice subcutaneously transplanted with RPMI 8226 cells showed tumor formation one week later. As of 7 weeks, the rate of tumorigenesis was 80% (12/15). Serum λ light chain was detected and no CD138 tumor cells were detected in peripheral blood. Two NOD/SCID mice each were subcutaneously transplanted with RPMI-Luc-Puro, RPMI-Luc-mcherry and MOLP8-Luc-Puro cells respectively. No tumor signal was detected through IVIS in RPMI-Luc-mcherry cells-transplanted mice. There was tumor signal at 1 week in RPMI-Luc-Puro and MOLP8-Luc-Puro cells-transplanted mice, the former disappeared at 2 weeks and the latter persisted more than 3 weeks. NSG mice subcutaneously transplanted with both cells persistently displayed the tumor signal. Neither NOD/SCID nor NSG mice transplanted with RPMI 8226, RPMI-Luc-Puro, RPMI-Luc-mcherry or MOLP8-Luc-Puro cells through tail vein developed the tumor signal. Only one NSG mice transplanted with MOLP8-Luc-Puro cells appeared transient tumor signal.@*CONCLUSION@#Unirradiated mice transplanted with MM cell lines tended to develop local tumor, and failed to develop disseminated tumor. The tumorigenicity of different cell lines is quite different and the vector transfection can reduce the tumorigenic ability. NSG mice with more severe immunodeficiency are more suitable for tumor growth.


Subject(s)
Animals , Carcinogenesis , Cell Line , Mice , Mice, Inbred NOD , Mice, SCID , Multiple Myeloma
8.
Neuroscience Bulletin ; (6): 216-224, 2019.
Article in English | WPRIM | ID: wpr-775435

ABSTRACT

Diffuse intrinsic pontine glioma (DIPG) is the main cause of brain tumor-related death among children. Until now, there is still a lack of effective therapy with prolonged overall survival for this disease. A typical strategy for preclinical cancer research is to find out the molecular differences between tumor tissue and para-tumor normal tissue, in order to identify potential therapeutic targets. Unfortunately, it is impossible to obtain normal tissue for DIPG because of the vital functions of the pons. Here we report the human fetal hindbrain-derived neural progenitor cells (pontine progenitor cells, PPCs) as normal control cells for DIPG. The PPCs not only harbored similar cell biological and molecular signatures as DIPG glioma stem cells, but also had the potential to be immortalized by the DIPG-specific mutation H3K27M in vitro. These findings provide researchers with a candidate normal control and a potential medicine carrier for preclinical research on DIPG.


Subject(s)
Animals , Brain Stem Neoplasms , Genetics , Metabolism , Pathology , Cell Line, Tumor , Cellular Senescence , Female , Glioma , Genetics , Metabolism , Pathology , Histones , Genetics , Humans , Mice, Inbred NOD , Mice, SCID , Neoplasm Transplantation , Neoplastic Stem Cells , Metabolism , Pathology , Neural Stem Cells , Metabolism , Pathology , Pons , Embryology , Metabolism , Pathology , Primary Cell Culture
9.
Frontiers of Medicine ; (4): 57-68, 2019.
Article in English | WPRIM | ID: wpr-771260

ABSTRACT

Lung cancer is the most common incident cancer and the leading cause of cancer death. In recent years, the development of tumor immunotherapy especially chimeric antigen receptor T (CAR-T) cell has shown a promising future. Epidermal growth factor receptor variant III (EGFRvIII) is a tumor-specific mutation expressed in various types of tumors and has been detected in non-small cell lung cancer with a mutation rate of 10%. Thus, EGFRvIII is a potential antigen for targeted lung cancer therapy. In this study, CAR vectors were constructed and transfected into virus-packaging cells. Then, activated T cells were infected with retrovirus harvested from stable virus-producing single clone cell lines. CAR expression on the surfaces of the T cells was detected by flow cytometry and Western blot. The function of CAR-T targeting EGFRvIII was then evaluated. The EGFRvIII-CAR vector was successfully constructed and confirmed by DNA sequencing. A stable virus-producing cell line was produced from a single clone by limited dilution. The culture conditions for the cell line, including cell density, temperature, and culture medium were optimized. After infection with retrovirus, CAR was expressed on more than 90% of the T cells. The proliferation of CAR-T cells were induced by cytokine and specific antigen in vitro. More importantly, EGFRvIII-CART specifically and efficiently recognized and killed A549-EGFRvIII cells with an effector/target ratio of 10:1 by expressing and releasing cytokines, including perforin, granzyme B, IFN-γ, and TNF-α. The in vivo study indicated that the metastasis of A549-EGFRvIII cells in mice were inhibited by EGFRvIII-CART cells, and the survival of the mice was significantly prolonged with no serious side effects. EGFRvIII-CART showed significantly efficient antitumor activity against lung cancer cells expressing EGFRvIII in vivo and in vitro. Therefore, CAR-T targeting EGFRvIII is a potential therapeutic strategy in preventing recurrence and metastasis of lung cancer after surgery.


Subject(s)
Animals , Carcinoma, Non-Small-Cell Lung , Allergy and Immunology , Therapeutics , Cell Line, Tumor , ErbB Receptors , Allergy and Immunology , Metabolism , Female , Humans , Immunotherapy, Adoptive , Methods , Lung Neoplasms , Allergy and Immunology , Therapeutics , Mice , Mice, Inbred NOD , Receptors, Chimeric Antigen , Allergy and Immunology , T-Lymphocytes , Allergy and Immunology , Xenograft Model Antitumor Assays
10.
Article in English | WPRIM | ID: wpr-773993

ABSTRACT

OBJECTIVE@#To investigate the protective effect of Zengye Decoction (, ZYD) on the submandibular glands (SMGs) in nonobese diabetic (NOD) mice.@*METHODS@#Twenty-seven female NOD mice were randomly equally divided into 3 groups: the model group, the hydroxychloroquine (HCQ) group, and the ZYD group. Nine C57/B6 mice served as the normal group. After 1-week acclimation, the HCQ and ZYD groups were intragastrically administered with HCQ and ZYD, respectively, and the normal and model groups were administered with normal saline. Changes in the salivary flow rate were observed. Mice from all 4 groups were sacrificed at the age of 20 weeks. The serum and SMGs were collected. Serum cytokines gamma-interferon (IFN-γ), interleukin-10 (IL-10) were detected by enzyme-linked immunosorbent assay. Histological changes in the submandibular glands were examined by hematoxylin and eosin staining. The mRNA expression of IFN-γ, IL-10 and vasoactive intestinal peptide (VIP) in the submandibular glands were measured by real-time polymerase chain reaction.@*RESULTS@#Compared with the model group, the salivary flow of the ZYD group significantly increased (P<0.05), the extent of the histological changes was ameliorated (P<0.05), and the Th1/Th2 cytokine imbalance was remedied (P<0.05). In the ZYD-treated mice, the VIP mRNA was up-regulated (P<0.05).@*CONCLUSIONS@#ZYD is beneficial in protecting structure and function of SMGs in NOD mice. The mechanism may be associated with the correction of the Th1/Th2 cytokine imbalance, and with the prevention of a progressive decline of the VIP level.


Subject(s)
Animals , Cytokines , Blood , Drugs, Chinese Herbal , Pharmacology , Female , Mice , Mice, Inbred C57BL , Mice, Inbred NOD , Salivation , Sjogren's Syndrome , Drug Therapy , Allergy and Immunology , Submandibular Gland , Pathology , Th1 Cells , Allergy and Immunology , Th2 Cells , Allergy and Immunology , Vasoactive Intestinal Peptide , Genetics
11.
Chinese Medical Journal ; (24): 1174-1184, 2018.
Article in English | WPRIM | ID: wpr-688148

ABSTRACT

<p><b>Background</b>The incidence of cancer, diabetes, and autoimmune diseases has been increasing. Furthermore, there are more and more patients with solid organ transplants. The survival rate of these immunocompromised individuals is extremely low when they are severely hit-on. In this study, we established cardiac arrest cardiopulmonary resuscitation (CPR) model in severe combined immunodeficient (SCID) mice, analyzed the expression and activation of mitochondrial autophagy and NLRP3 inflammasome/caspase-1, and explored mitochondrial repair and inflammatory injury in immunodeficiency individual during systemic ischemia-reperfusion injury.</p><p><b>Methods</b>A potassium chloride-induced cardiac arrest model was established in C57BL/6 and nonobese diabetic/SCID (NOD/SCID) mice. One hundred male C57BL/6 mice and 100 male NOD/SCID mice were randomly divided into five groups (control, 2 h post-CPR, 12 h post-CPR, 24 h post-CPR, and 48 h post-CPR). A temporal dynamic view of alveolar epithelial cells, macrophages, and neutrophils from bronchoalveolar lavage fluid (BALF) was obtained using Giemsa staining. Spatial characterization of phenotypic analysis of macrophages in the lung interstitial tissue was analyzed by flow cytometry. The morphological changes of mitochondria 48 h after CPR were studied by transmission electron microscopy and quantified according to the Flameng grading system. Western blotting analysis was used to detect the expression and activation of the markers of mitochondrial autophagy, NLRP3 inflammasome, and caspase-1.</p><p><b>Results</b>(1) In NOD/SCID mice, macrophages were disintegrated in BALF, and many alveolar epithelial cells were shed at 48 h after resuscitation. Compared with C57BL/6 mice, the ratio of macrophages/total cells peaked at 12 h and was significantly higher in NOD/SCID mice (31.17 ± 4.13 vs. 49.69 ± 2.43, t = 14.46, P = 0.001). After 24 h, the results showed a downward trend. Furthermore, a large number of macrophages were disintegrated in the BALF. (2) Mitochondrial autophagy was present in both C57BL/6 and NOD/SCID mice after CPR, but it began late in the NOD/SCID mice. Compared with C57BL/6 mice, phos-ULK1 (Ser) expression was significantly lower at 2 h and 12 h after CPR (2 h after CPR: 1.88 ± 0.36 vs. 1.12 ± 0.11, t = -1.36, P < 0.01 and 12 h after CPR: 1.52 ± 0.16 vs. 1.05 ± 0.12, t = -0.33, P < 0.01), whereas phos-ULK1 (Ser) expression was significantly higher at 2 h and 12 h after CPR in NOD/SCID mice (2 h after CPR: 1.28 ± 0.12 vs. 1.69 ± 0.14, t = 1.7, P < 0.01 and 12 h after CPR: 1.33 ± 0.10 vs. 1.94 ± 0.13, t = 2.75, P < 0.01). (3) Furthermore, NLRP3 inflammasome/caspase-1 activation in the pulmonary tissues occurred early and for only a short time in C57BL/6 mice, but this phenomenon was sustained in NOD/SCID mice. The expression of the NLRP3 inflammasome increased modestly in the C57 mice, but the increase was higher in the NOD/SCID mice than in the C57BL/6 mice, especially at 12, 24, 48 h after CPR (48 h after CPR: 1.46 ± 0.13 vs. 2.97 ± 0.19, t = 5.34, P = 0.001). The expression of caspase-1-20 generally followed the same pattern as the NLRP3 inflammasome.</p><p><b>Conclusions</b>There is a regulatory relationship between the NLRP3 inflammasome and mitochondrial autophagy after CPR in the healthy mice. This regulatory relationship was disturbed in the NOD/SCID mice because the signals for mitochondrial autophagy occurred late, and NLRP3 inflammasome- and caspase-1-dependent cell injury was sustained.</p>


Subject(s)
Animals , Autophagy , Physiology , Heart Arrest , Metabolism , Inflammasomes , Metabolism , Lung , Metabolism , Macrophages , Metabolism , Physiology , Male , Mice , Mice, Inbred C57BL , Mice, Inbred NOD , Mice, SCID , Mitochondria , Metabolism , NLR Family, Pyrin Domain-Containing 3 Protein , Metabolism
12.
Chinese Medical Journal ; (24): 1533-1540, 2018.
Article in English | WPRIM | ID: wpr-688080

ABSTRACT

<p><b>Background</b>The previous study showed that mycophenolic acid (MPA) synergizing with lipopolysaccharide (LPS) promoted interleukin (IL)-1β release, but the mechanism is unclear. This study aimed to investigate the mechanism of MPA synergizing with LPS to induce IL-1β release.</p><p><b>Methods</b>Undiluted human blood cells, THP-1 human myeloid leukemia mononuclear cells (THP-1) cells, or monocytes were stimulated with LPS and treated with or without MPA, and the supernatant IL-1β was detected by enzyme-linked immunosorbent assay. The mRNA levels of IL-1β were detected by real-time quantitative polymerase chain reaction. The intracellular protein levels of nuclear factor kappa B (NF-κB) phospho-p65 (p-p65), precursor interleukin-1β (pro-IL-1β), NOD-like receptor pyrin domain containing-3 (NLRP3), and cysteine aspartic acid-specific protease-1 (caspase-1) p20 in THP-1 cell were measured by Western blot.</p><p><b>Results</b>The MPA alone failed to induce IL-1β, whereas MPA synergized with LPS to increase IL-1β in a dose-dependent manner (685.00 ± 20.00 pg/ml in LPS + 5 μmol/L MPA group, P = 0.035; 742.00 ± 31.58 pg/ml in LPS + 25 μmol/L MPA group, P = 0.017; 1000.00 ± 65.59 pg/ml in LPS + 75 μmol/L MPA group, P = 0.024; versus 408.00 ± 35.50 pg/ml in LPS group). MPA alone has no effect on the IL-1β mRNA expression, LPS induced the expression of IL-1β mRNA 2761 fold, and LPS + MPA increased the IL-1β expression 3018 fold, which had the same effect with LPS group (P = 0.834). MPA did not affect the intracellular NF-κB p-p65 and pro-IL-1β protein levels but activated NLRP3 inflammasome. Ac-YVAD-cmk blocked the activation of caspase-1 and subsequently attenuated IL-1β secretion (181.00 ± 45.24 pg/ml in LPS + MPA + YVAD group vs. 588.00 ± 41.99 pg/ml in LPS + MPA group, P = 0.014).</p><p><b>Conclusions</b>Taken together, MPA synergized with LPS to induce IL-1β release via the activation of caspase-1, rather than the enhanced production of pro-IL-1β. These findings suggested that patients immunosuppressed with mycophenolate mofetil may have overly activated caspase-1 during infection, which might contribute to a more sensitive host defense response to invading germs.</p>


Subject(s)
Animals , Caspase 1 , Metabolism , Cells, Cultured , Humans , Inflammasomes , Interleukin-1beta , Metabolism , Lipopolysaccharides , Pharmacology , Mice , Mice, Inbred NOD , Mycophenolic Acid , Pharmacology , NLR Family, Pyrin Domain-Containing 3 Protein
13.
Chinese Journal of Applied Physiology ; (6): 309-312 339, 2018.
Article in Chinese | WPRIM | ID: wpr-773753

ABSTRACT

OBJECTIVE@#To study the effect of ursolic acid on cardiomyopathy in mice with diabetes induced by high-fat diet combined with low dose streptozotocin, and to explore its possible mechanism.@*METHODS@#Thirty male ICR mice were randomly divided into control group (=10) and moulding group (=20), the mice in the two groups were fed with regular diet and high-fat diet respectively for 6 weeks, and then the mice in the moulding group were injected with streptozotocin (30 mg/kg) for 5 successive days to induce diabetes mellitus (DM). Fasting blood glucose (FBG) was measured after 9 days. Mice with FBG over 11.1 mmol/L were regarded as DM. Twenty DM mice were randomly divided into model group and ursolic acid group (=10). Mice in each group were continuously administrated ursolic acid (100 mg/kg) or corresponding solvent intragastrically for 8 weeks. After that, FBG was measured, body weight (BW), heart weight and left ventricular weight were weighed in order to calculate the heart mass index (HMI) and left ventricular mass index (LVMI). Levels of creatine kinase (CK), lactate dehydrogenase (LDH) in serum and the level of superoxide dismutase (SOD), malondialdehyde (MDA) in myocardial tissue were detected. HE staining was used to observe pathological changes of myocardial tissue. Immunohistochemistry was employed to determine the expression of NOD-like receptor protein 3 (NLRP3) and interleukin 1β (IL-1β).@*RESULTS@#Compared with the control group, HMI, LVMI were apparently enlarged, levels of FBG, CK, LDH in serum and MDA in myocardial tissue were extremely increased, while the activity of SOD in myocardial tissue were extraordinary decreased in diabetic group. HE staining of myocardium showed that arrangement disorder of myocardial fibers, edema and hypertrophy in myocardial cell, as well as inflammatory cell infiltration in model group. Immunohistochemistry showed that the expression of NLRP3 and IL-1β in myocardial tissue increased obviously in model group, the above changes inursolic acid group were significantly ameliorated.@*CONCLUSIONS@#Ursolic acid has a obvious protective effect on myocardial injury in mice with diabetes induced by high-fat diet combined with low dose streptozotocin, and its mechanism may be associated with inhibiting NLRP3 inflammasome activation, reducing IL-1β generation and alleviating myocardial inflammatory injury.


Subject(s)
Animals , Cardiomyopathies , Diabetes Mellitus, Experimental , Male , Mice , Mice, Inbred ICR , Mice, Inbred NOD , Myocardium , NLR Family, Pyrin Domain-Containing 3 Protein , Triterpenes , Pharmacology
14.
Journal of Experimental Hematology ; (6): 1616-1620, 2018.
Article in Chinese | WPRIM | ID: wpr-773047

ABSTRACT

OBJECTIVE@#To investigate the effect of Quercetin on cell cycle and adhesive molecules of NOD.SCID mice with acule B lymphocytic leuaemia(B-ALL).@*METHODS@#5×10 Nalm-6(B-ALL cell line) cells were injected into the tail vein of 48 NOD/SCID mice to establish the NOD/SCID mice with B-ALL. After 15 day, the NOD/SCID mice with B-ALL were randomly divided into 3 groups: salive group as control (injection with saline of 0.2 ml/mouse), cyclophos-phamid group (injection with cyclophosphamide of 100µg/kg) and quercetin group(injection with quercetin of 3 mg/kg). After treatment for 21 d, the perecntage of Nalm-6 cells in G1, G2, M and S phases was detected by flow cytonetry; the B lymphocytes Nalm-6 cells, neutrophils and WBC in while blood were counted before and after treatment; the expression of intercellalar. Adhesion molecole-1(FCAIU-1), vascular cell adhesion molecule-1(VCAM-1) and P-selectin was detected by double autibody soundwich method.@*RESULTS@#Compared with level before treatment, the expression of ICAM-1, VCAM-1 and P-selectin decreased after treatment with guercetin, The hemogram showed that the peripheral blood nentrophil level obviously increased, while the levels of B lymphocytes, Nalm-6 cells and WBC count decreased obviously after treatment with guercetin. The cell proliferatim rario in G0/G1 phase decreased, yet the cell proliferation ratio in S and G2/M phases increased after treatment with guercetin.@*CONCLUSION@#The guercetin can decrease the intercellular adhesion through inhibition of ICAM-1 expression, and arrests Nalm-6 cells in S and G2/M phases. The guercetin has obviously inhibitory effect on B-ALL cells.


Subject(s)
Animals , Cell Adhesion , Cell Adhesion Molecules , Intercellular Adhesion Molecule-1 , Leukemia, B-Cell , Mice , Mice, Inbred NOD , Mice, SCID , Quercetin , Vascular Cell Adhesion Molecule-1
15.
Article in Chinese | WPRIM | ID: wpr-813149

ABSTRACT

To observe the protective effect of alpha-mangostin (α-MG) on focal segmental glomurular sclerosis (FSGS) induced by adriamycin.
 Methods: Adriamycin nephropathy (AN) model was induced by adriamycin (10 mg/kg) via a tail vein. Then the mice were treated with α-MG (12.5 mg/kg) or normal salin once daily for 6 weeks. At the end of 6 weeks, the mice were sacrificed, and the kidneys and blood samples were collected. Histopathology of the kidneys were analyzed under the optical microscope. The serum levels of biochemical indicators, such as serum creatinine (SCr), blood urea nitrogen (BUN) and cholesterol were determined. The levels of superoxide anion, malondialdehyde (MDA), and glutathione (GSH), the activity of superoxide dismutase (SOD) and catalase (CAT) in kidney tissues were determined. Serum levels of IL-1β, IL-18, IL-10 and adiponectin were determined. The levels of TGF-β1, collagen I (Col I), α-SMA, silent information regulator 1 (Sirt1) and the nucleotide-binding domain (NOD)-like receptor protein 3 (NLRP3) in kidney tissues were determined using immunohistochemical staining, Western blot, and RT-PCR.
 Results: The levels of SCr, proteinuria, urine protein to creatinine ratio and serum cholesterol were attenuated in AN mice after α-MG treatment, while creatinine clearance rate and serum albumin were upregulated (P<0.05). α-MG treatment alleviated the glomerular and interstitial fibrosis, downregulated the expression of fibrosis markers, such as Col I and α-SMA (P<0.05). α-MG treatment reduced the production of superoxide anion, the levels of MDA and GSH, and increased the activity of CAT and SOD (P<0.05). α-MG treatment inhibitd the generation of pro-inflammatory cytokines, such as IL-1β and IL-18 and promoted the production of anti-inflammatory cytokines, such as the IL-10 and adiponectin (P<0.05); α-MG treatment promoted the expression of Sirt1, inhibitd the expression of NLRP3 in kidney tissues (P<0.05).
 Conclusion: α-MG could attenuates FSGS of mice induced by adriamycin ameliorate and improve renal function. α-MG exerts its anti-inflammatory and anti-oxidative effects by up-regulation the expression of Sirt1 and suppression of NLRP3.


Subject(s)
Animals , Disease Models, Animal , Doxorubicin , Glomerulosclerosis, Focal Segmental , Kidney , Mice , Mice, Inbred NOD , Protein Kinase Inhibitors , Pharmacology , Therapeutic Uses , Xanthones , Pharmacology , Therapeutic Uses
16.
São Paulo; s.n; 2017. 135 p. ^c2.24 Mbytesilus PDF, tab, graf.
Thesis in Portuguese | LILACS | ID: biblio-846877

ABSTRACT

Sepse é uma resposta sistêmica e deletéria do indivíduo a uma infecção, sendo um importante problema de saúde pública. Pacientes diabéticos são bastante afetados representando cerca de 22% de todos os pacientes sépticos. A suscetibilidade para o desenvolvimento de sepse no diabetes, bem como a ação da insulina em modular alguns parâmetros imunológicos necessitam de maiores esclarecimentos O objetivo deste estudo foi avaliar os efeitos do tratamento com insulina em um modelo murino de diabetes e sepse. Camundongos C57BL/6 foram tornados diabéticos por administração de aloxana. Os seguintes parâmetros foram analisados vinte e quatro horas após a ligadura cecal e punção (CLP): (a) interleukine (IL)-6, IL-10, chemokine (C -C motif) ligand 2 (CCL2) e tumor necrosis fator (TNF ) -α no soro; (b) os níveis de IL-1ß, IL-6, TNF-α, IL-10, chemokine (C -X-C motif) ligand (CXCL)-1 e CXCL2 no lavado peritoneal (LPe) e broncoalveolar (LBA), bem como nos rins e fígado; (c) contagens celulares totais e diferenciais em LPe e LBA; (d) capacidade endocítica de neutrófilos e produção de espécies reactivas de oxigénio (ERO); (e) níveis de apoptose e necrose no baço e níveis relativos de células CD4+ e CD8+; (f) resultados histopatológicos de pulmão, rim e fígado; e (g) níveis de translocação nuclear de NF-κB p65. Camundongos diabéticos-CLP exibiram concentrações séricas aumentadas de TNF-α, IL-6, CCL2, IL-1, IL-6, CXCL1, CXCL2 e IL-10 e contagens de neutrófilos em LPe. A capacidade endocítica dos neutrófilos e a produção de ERO apresentavam-se reduzidas em animais CLP-diabéticos e os níveis de IL-6, TNF-α, CXCL1 e CXCL2 em LBA e IL-1ß, IL-6, CXCL1 e CXCL2 nos homogenados renais aumentaram diabéticos -CLP. O tratamento destes com insulina reduziu os nívies de citocinas séricas, aumentou a concentração de citocinas e a migração celular para o Lpe, restaurou a capacidade endocítica e a produção de ERO e reduziu a translocação nuclear NF-κB p65 no tecido renal. Estes dados sugerem que a insulina modula a produção/libertação de citocinas, regula a migração celular, a apoptose, a necrose e a translocação nuclear de NF-κB p65 na sepse induzida por CLP em camundongos diabéticos.


Sepsis is a systemic and harmful response of the individual to infection and is an important public health problem. Diabetic patients are greatly affected representing about 22% of all septic patients. The susceptibility to sepsis development in diabetic individuals and insulin action in modulating some immunological parameters require further clarification. The aim of this study was to evaluate the effects of insulin treatment in a mouse model of diabetes and sepsis. C57BL/6 mice were rendered diabetic by alloxan administration. The following parameters were analyzed twenty-four hours after a cecal ligation and puncture (CLP): (a) interleukin (IL)-6, IL-10, chemokine (C-C motif) ligand 2 (CCL2), and tumor necrosis factor (TNF) - α levels in serum; (b) IL-1ß, IL-6, TNF-α, IL-10, chemokine (C-X-C motif) ligand (CXCL)1 and CXCL2 levels in peritoneal lavage (PeL) and bronchoalveolar lavage (BAL) fluid, as well as in the kidneys and liver; (c) total and differential cell counts in PeL and BAL fluid; (d) neutrophil endocytic capacity and reactive oxygen species (ROS) production; (e) spleen cell apoptosis and necrosis levels and relative CD4+ and CD8+ T cell levels; (f) lung, kidney, and liver histopathological results; and (g) NF-kB p65 nuclear translocation levels. Diabetic-CLP mice exhibited increased serum TNF-α, IL-6, CCL2, IL-1, IL-6, CXCL1, CXCL2 and IL-10 concentrations and neutrophil counts in PeL fluid. Neutrophil endocytic capacity and ROS production were decreased in diabetic-CLP mice, and IL-6, TNF- α, CXCL1 and CXCL2 leves in BAL fluid and IL-1ß, IL-6, CXCL1 and CXCL2 levels in kidney homogenates were increased in diabetic-CLP mice. Treatment of these mice with insulin reduced serum cytokine levels increased cytokine and cell migration into PeL fluid, and restored neutrophil endocytic capacity and ROS production and NF-kB p65 nuclear translocation in the kidney. These data suggest that insulin modulates cytokine production/release, regulates cellular migration, apoptosis, necrosis and NF-kB p65 nuclear translocation in CLP-induced sepsis in diabetic mice.


Subject(s)
Animals , Male , Female , Mice , Cell Death/drug effects , Insulin/analysis , Mice, Inbred NOD/genetics , Sepsis/complications , Acute Kidney Injury/complications , Cytokines/administration & dosage
17.
Pesqui. vet. bras ; 36(7): 617-624, jul. 2016. tab, graf, ilus
Article in English | LILACS, VETINDEX | ID: lil-794778

ABSTRACT

Chronic cutaneous lesions affect 15% of diabetic human patients and represent a risk 15 to 46 times larger of limb amputations compared to people with normal glycemia. It is assumed that half of these amputations could be prevented by early treatment of wounds, for example, with proper cell therapy. Objectives: In this study, the action of the autologous transplant of mesenchymal stem-cells (MSC) was evaluated compared to the treatment with autologous platelet rich plasma (PRP) in the cicatrization of cutaneous lesions induced in diabetic mice. These animals were previously treated with streptozootocin to induce diabetes mellitus and round wounds of 1.5cm in diameter were created in the posterior region. Diameters of the wounds and healing time were evaluated during 30 days and the results were submitted to variance analysis and Tukey's test average. It was noticed that the animals treated with MSC presented a more accelerated cicatrization of the cutaneous lesion than the animals treated with PRP. However, the treatment with PRP presented better results than just the daily asepsis of the lesions with saline or covering them with semi-permeable bandage. Besides, the use of semi-permeable bandage kept the cutaneous lesions of diabetic mice did not interfere negatively with cicatrization, proved to be harmless to use, but kept the cutaneous lesions more hydrated than the ones exposed to the environment.(AU)


Lesões cutâneas crônicas afetam 15% dos pacientes diabéticos e humanos representam um risco 15 a 46 vezes maior de amputações de membros em comparação com as pessoas com a glicemia normal. Supõe-se que a metade destas amputações poderia ser evitada por meio do tratamento precoce das feridas cutâneas com, por exemplo, uma adequada terapia celular. Objetivos: Neste estudo, a ação do transplante autólogo de células estaminais mesenquimais (MSC) foi avaliada em comparação com o tratamento com plasma rico em plaquetas autólogo (PRP) na cicatrização de lesões cutâneas induzidas em camundongos diabéticos. Estes animais foram previamente tratados com estreptozotocina para induzir diabetes mellitus e feridas redondas de 1,5 cm de diâmetro foram criadas na região posterior. Os diâmetros dos ferimentos e tempo de cicatrização foram avaliados durante 30 dias e os resultados foram submetidos à análise de variância e média pelo teste de Tukey. Verificou-se que os animais tratados com MSC apresentam uma cicatrização mais acelerada da lesão cutânea que do que os animais tratados com PRP. No entanto, o tratamento com PRP apresentou melhores resultados do que apenas a assepsia das lesões diariamente com solução salina ou cobrindo-os com atadura semi-permeável. Além disso, a utilização de atadura semi-permeável mantidas as lesões cutâneas de camundongos diabéticos não interfere negativamente com a cicatrização, provou ser inofensiva para usar, mas manteve as lesões cutâneas hidratadas mais do que os expostos ao meio ambiente.(AU)


Subject(s)
Animals , Male , Guinea Pigs , Mice , Platelet-Rich Plasma/physiology , Stem Cells/physiology , Transplantation, Autologous/rehabilitation , Wound Healing/physiology , Diabetes Mellitus/veterinary , Mice, Inbred NOD/physiology , Wounds and Injuries/veterinary
18.
Article in Chinese | WPRIM | ID: wpr-272478

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effects of oxygen concentration and reactive oxygen species (ROS) on the biological characteristics of hematopoietic stem cells (HSC) and to analyzed the relationship among the oxygen concentration, ROS and the biological characteristics of mouse HSC through simulation of oxygen environment experienced by PB HSC during transplantation.</p><p><b>METHODS</b>The detection of reactive oxygen species (ROS), in vitro amplification, directional differentiation (BFU-E, CFU-GM, CFU-Mix), homing of adhesion molecules (CXCR4, CD44, VLA4, VLA5, P-selectin), migration rate, CFU-S of NOD/SCID mice irradiated with sublethal dose were performed to study the effect of oxgen concentration and reactive oxygen species on the biological characteristics of mouse BM-HSC and the relationship among them.</p><p><b>RESULTS</b>The oxygen concentrations lower than normal oxygen concentration (especially hypoxic oxygen environment) could reduce ROS level and amplify more Lin(-) c-kit(+) Sca-1(+) BM HSC, which was more helpful to the growth of various colonies (BFU-E, CFU-GM, CFU-Mix) and to maintain the migratory ability of HSC, thus promoting CFU-S growth significantly after the transplantation of HSC in NOD/SCID mice irradiated by a sublethal dose. BM HSC exposed to oxygen environments of normal, inconstant oxygen level and strenuously thanging of oxygen concentration could result in higher level of ROS, at the same time, the above-mentioned features and functional indicators were relatively lower.</p><p><b>CONCLUSION</b>The ROS levels of BM HSC in PB HSCT are closely related to the concentrations and stability of oxygen surrounding the cells. High oxygen concentration results in an high level of ROS, which is not helpful to maintain the biological characteristics of BM HSC. Before transplantation and in vitro amplification, the application of antioxidancs and constant oxygen level environments may be beneficial for transplantation of BMMSC.</p>


Subject(s)
Animals , Cell Differentiation , Culture Media , Chemistry , Erythroid Precursor Cells , Cell Biology , Granulocyte-Macrophage Progenitor Cells , Cell Biology , Hematopoietic Stem Cells , Cell Biology , Metabolism , Mice , Mice, Inbred NOD , Mice, SCID , Oxygen , Chemistry , Reactive Oxygen Species , Metabolism
19.
Chinese Journal of Biotechnology ; (12): 1685-1693, 2016.
Article in Chinese | WPRIM | ID: wpr-243689

ABSTRACT

Type 1 diabetes (T1D), the most prevalent human autoimmune disease, occurs in genetically susceptible individuals. Regulatory T cells (Tregs) are defective in T1D setting. Therefore, efforts to repair or restore Tregs in T1D may prevent or reverse this autoimmune disease. Here, we studied the potential role of rgp96 in preventing T1D, using non-obese diabetic (NOD) mice as an animal model. High-dose rgp96 immunization elicited efficient protection of mice against T1D, as evidenced by stable blood glucose, decreased disease incidence. Significantly increased CD4⁺ CD25⁺ Foxp3⁺ Tregs were observed in immunized mice. In vitro co-culture experiments demonstrated that rgp96 stimulation enhanced Treg proliferation and suppressive function by up-regulation of Foxp3 and IL-10. Our work shows that activation of Tregs by high-dose rgp96 immunization protects against T1D via inducing regulatory T cells and provides preventive and therapeutic potential for the development of an rgp96-based vaccine against T1D.


Subject(s)
Animals , Antigens, Neoplasm , Allergy and Immunology , Coculture Techniques , Diabetes Mellitus, Type 1 , Therapeutics , Forkhead Transcription Factors , Heat-Shock Proteins , Allergy and Immunology , Interleukin-10 , Allergy and Immunology , Mice , Mice, Inbred NOD , T-Lymphocytes, Regulatory , Allergy and Immunology , Up-Regulation , Vaccination
20.
Rev. chil. nutr ; 42(2): 151-156, jun. 2015. tab
Article in Spanish | LILACS | ID: lil-755552

ABSTRACT

The relationship between a high consumption of fructose-sweetened foods and obesity and its co-morbidities remains controversial. In this study the effects of three isocaloric and isolipidic diets containing different carbohydrates - fructose, sucrose and starch (AIN 93) - on biomass, abdominal fat depots, blood and liver lipid profile and hepatic histopathology in adult male IIMb/obese and diabetic rats were evaluated. Plasma cholesterol and triacylglycerol were significantly higher in fructose and sucrose groups, while liver lipids showed higher levels in the starch-fed group. There were no differences in hepatic histology in the three groups. The findings of this study support the hypothesis that sugar-rich diets -with fructose or sucrose - replacing starch in equivalent amounts produce similar effects in plasma glucose-lipid profile.


La controversia acerca de si el alto consumo de fructosa ha sido determinante en la prevalencia actual de la obesidad y sus comorbilidades, o si sus efectos son semejantes a los de un consumo equivalente de otros azúcares continúa vigente. En este trabajo se evaluaron los efectos de tres dietas isocalóricas e isolipídicas con fructosa, sacarosa o almidón -como control-, sobre la biomasa, el depósito adiposo abdominal, el perfil glucolipídico sanguíneo y los lípidos y la histología hepáticos en ratas adultas IIMb/β, obesas y diabéticas. Los valores de colesterol y triacilgliceroles plasmáticos fueron mayores en los grupos alimentados con fructosa y sacarosa, mientras que los lípidos hepáticos resultaron mayores en el grupo que consumió almidón. La histología hepática mostró leve esteatosis micro-macro vacuolar en los tres grupos. Los efectos de fructosa y sacarosa sobre las variables evaluadas no fueron diferentes.


Subject(s)
Rats/physiology , Sucrose , Mice, Inbred NOD , Fructose , Metabolism
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