OBJECTIVETo investigate the effect of necrotic cells on the secretion of inflammatory cytokines.

METHODSRAW264.7 macrophages and necrotic mouse thymocytes induced by heating were incubated for 18 h at a ratio of 5:1 in the absence or presence of lipopolysaccharide (LPS, 100 ng/ml). The supernatant of the cell culture was collected and the expression and secretion of the pro-inflammatory cytokines were measured using Bio-Plex suspension system.

RESULTSThe secretions of tumor necrosis factor-alpha (TNF-alpha) and interlukine-6 (IL-6) by macrophages co-cultured with the necrotic cells were significantly enhanced as compared with the control cells. The necrotic cells also significantly augmented the secretion of the pro-inflammatory cytokines induced by LPS.

CONCLUSIONNecrotic cells not only induces pro-inflammatory cytokine expression by themselves but also work synergistically with LPS to enhance the cytokine production, suggesting the important roles of necrotic cells to initiate and maintain the inflammatory responses.

Animals ; Cell Line ; Hot Temperature ; Inflammation ; etiology ; metabolism ; pathology ; Interleukin-6 ; secretion ; Lipopolysaccharides ; pharmacology ; Macrophages ; cytology ; drug effects ; metabolism ; Male ; Mice ; Necrosis ; complications ; Tumor Necrosis Factor-alpha ; secretion

OBJECTIVETo investigate the effects of apoptotic lymphocytes on the secretion of cytokines by hepatic sinusoidal endothelial cells (HSEC).

METHODSHuman HSEC cells were co-cultured for 16 h with allogenetic apoptotic lymphocytes induced by UVB irradiation. The supernatants were collected and the levels of interleukin-2, interferon-gamma, and tumor necrosis factor-alpha were detected by Luminex technique.

RESULTSAll the cytokines were down-regulated by about 50% in HSECs after co-culture with the apoptotic lymphocytes as compared with those in the control group (P<0.05).

CONCLUSIONSCo-culture with apoptotic lymphocytes can down-regulate the secretion of pro-inflammatory cytokines in HSECs, which may contribute to tolerogenic microenvironment in the liver.

Apoptosis ; physiology ; Cells, Cultured ; Coculture Techniques ; Cytokines ; secretion ; Down-Regulation ; Endothelial Cells ; cytology ; metabolism ; Humans ; Immune Tolerance ; Interferon-gamma ; secretion ; Interleukin-2 ; secretion ; Liver ; cytology ; Lymphocytes ; cytology ; Tumor Necrosis Factor-alpha ; secretion

OBJECTIVETo investigate the significance of PKC and p44/42 mitogen-activated protein kinase (MAPK) signal transduction in ischemic preconditioning (IP).

METHODSThrough liver cell IP models, PKC inhibitor and MEK inhibitor were utilized to analyze the phosphorylation level of p44/42 MAPK and cell viability was also observed. Rat liver IP models were established which were treated with various drugs. Then the phosphorylation level of p44/42 MAPK in vivo and serum alanine aminotransferase (ALT), aspartate aminotransferase (AST) concentrations were detected. And cellular structures were observed under light microscopy.

RESULTSSimilar results were obtained in vivo and in vitro IP models. Compared with the ischemia reperfusion (IR) group in vivo, the phosphorylation level of p44/42 MAPK was obviously increased in IP treated rats (q = 27.217, P < 0.01), and the cellular structure injured slightly. The concentrations of serum ALT and AST in IP group were significantly lower than those in IR group (281.0 U/L +/-35.6 U/L vs 762.8 U/L +/-130.5 U/L and 407.7 U/L +/-73.7 U/L vs 820.9 U/L +/-111.3 U/L, P < 0.01). However, opposite changes were found in PKC and MEK inhibited groups, when compared to IP group. The phosphorylation level of p44/42 MAPK was obviously decreased, the liver tissues injured evidently, and the concentrations of serum ALT and AST (645.61 U/L +/-90.4 U/L, 678.6 U/L +/-136.5U/L and 466.2 U/L +/-82.8 U/L, 732.9 U/L +/-91.1 U/L, respectively) were significantly greater than those in IP group.

CONCLUSIONThese results suggest that p44/42 MAPK pathway plays a vital role in the protection of hepatocytes in ischemic preconditioning.

Alanine Transaminase ; blood ; Animals ; Aspartate Aminotransferases ; blood ; Hepatocytes ; cytology ; physiology ; Humans ; Ischemic Preconditioning ; Liver ; blood supply ; cytology ; Male ; Mitogen-Activated Protein Kinase 1 ; metabolism ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Phosphorylation ; Protein Kinase C ; metabolism ; Rats ; Rats, Sprague-Dawley ; Signal Transduction

This study was aimed to investigate the effects of decitabine (DAC) on proliferation and apoptosis of leukemia NB4 and K562 cells. The proliferation inhibition of DAC on NB4 and K562 cells was detected by Trypan blue staining. After treatment of DAC at different concentrations, the changes of cell cycle and CD11b expression was determined by flow cytometry. The cell morphological changes were observed by Wright's staining. The DNA ladder was used to detect cell apoptosis. The results indicated that DAC significantly inhibited the proliferation of NB4 and K562 cells in dose-and time-dependent manner. The median inhibitory concentration (IC50) of DAC-treated NB4 and K562 cells for 72 h was 0.113 µmol/L and 0.138 µmol/L, respectively. After treating these two cell lines with DAC at different concentration for 72 h, the cell ratio in G0/G1 phase significantly increased, while the cell ratio in S phase obviously decreased in 0.15 µmol/L DAC group (P < 0.05). The expression levels of myeloid differentiation antigen CD11b of both cell lines significantly increased in contrast to the control group (P < 0.05). The cell morphology detected by Wright's staining displayed partial differentiation and apoptosis after treating NB4 and K562 cells with DAC for 48 h. Typical apoptotic DNA ladder was observed in 0.15 µmol/L DAC group at 48 h. It is concluded that DAC can inhibit NB4 and K562 cell proliferation, induce cell differentiation and apoptosis, but more obviously for NB4 cells.
Apoptosis ; drug effects ; Azacitidine ; analogs & derivatives ; pharmacology ; Cell Differentiation ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Humans ; K562 Cells

OBJECTIVETo track the location of the transfused apoptotic allogeneic lymphocytes and asses the process of their accumulation and phagocytosis removal as consequences on allograft tolerance in recipient mice.

METHODSDonor spleen lymphocytes were labeled by CFSE and induced to apoptosis by dexamethasone incubation. After purification by anti-annexin V-conjugated magnetic beads isolation, apoptotic lymphocytes were transfused into recipient mice through the tail veins. Tissue samples from various organs were taken at various time points to analyze the fates of the apoptotic allogeneic lymphocytes and the phagocytosis of them by organ resident APCs.

RESULTSUsing fluorescent microscopy and flow cytometry, after the apoptotic cells were recognized and uptaken, the largest amount of labeled cells were accumulated in the livers and disappeared within not more than 12 hours. Recipient liver APCs were highly efficacious in phagocytosis of apoptotic allogeneic lymphocytes; the removal was completed within 15 minutes after incubation. LSEC, KC and LDC all phagocytosized the apoptotic allogeneic lymphocytes but with significantly different rates. Considering the numbers of those cells in a normal liver, it could be calculated that LSEC and KC had greater effects in this activity.

CONCLUSIONSThe liver deserves foremost attention for study of the mechanism of allografts tolerance induced by pre-transfusion of apoptotic donor spleen lymphocytes. LSEC and KC are the main functional APCs to the alloantigens.

Animals ; Antigen-Presenting Cells ; cytology ; immunology ; Apoptosis ; physiology ; Liver ; cytology ; immunology ; Lymphocyte Transfusion ; Lymphocytes ; cytology ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Phagocytosis ; physiology ; Spleen ; cytology ; Transplantation, Homologous

AIMTo assess whether exposure to computers harms the semen quality of healthy young men.

METHODSA total of 178 subjects were recruited from two maternity and children healthcare centers in Shanghai, 91 with a history of exposure to computers (i.e., exposure for 20 h or more per week in the last 2 years) and 87 persons to act as control (no or little exposure to computers). Data on the history of exposure to computers and other characteristics were obtained by means of a structured questionnaire interview. Semen samples were collected by masturbation in the place where the semen samples were analyzed.

RESULTSNo differences in the distribution of the semen parameters (semen volume, sperm density, percentage of progressive sperm, sperm viability and percentage of normal form sperm) were found between the exposed group and the control group. Exposure to computers was not found to be a risk factor for inferior semen quality after adjusting for potential confounders, including abstinence days, testicle size, occupation, history of exposure to toxic substances.

CONCLUSIONThe present study did not find that healthy men exposed to computers had inferior semen quality.

Adult ; Case-Control Studies ; China ; Computers ; Electromagnetic Fields ; adverse effects ; Humans ; Male ; Semen ; Surveys and Questionnaires

OBJECTIVETo establish an assay method for detecting the migration of transferred apoptotic cells into the recipient using flow cytometry.

METHODSSpleen lymphocytes were isolated and labeled with an intracellular amine dye, carboxyfluorescein diacetate succinimidyl ester (CFSE), to allow discrimination. The labeled cells were induced with dexamethasone to undergo apoptosis and transferred into recipient mice via tail venous transfusion. Flow cytometry and histological examination of different tissues were performed at different time points. The stability of CFSE labeling for apoptotic cells was also tested.

RESULTSThe CFSE-labeled apoptotic cells were highly fluorescent with a positive labeling rate of (98.0+/-1.9)%. The stability of CFSE-labeling was testified, and the CFSE-labeled apoptotic cells entering different tissues at different time points were detected by flow cytometry and verified by histological examination.

CONCLUSIONFlow cytometry using CFSE labeling is reliable, sensitive, precise and convenient for apoptotic cell tracing in vivo and in vitro.

Adoptive Transfer ; methods ; Animals ; Apoptosis ; Dexamethasone ; pharmacology ; Female ; Flow Cytometry ; methods ; Fluoresceins ; chemistry ; pharmacokinetics ; Fluorescent Dyes ; chemistry ; pharmacokinetics ; Lymphocytes ; chemistry ; cytology ; drug effects ; Mice ; Mice, Inbred BALB C ; Reproducibility of Results ; Spleen ; cytology ; Succinimides ; chemistry ; pharmacokinetics

Recently, phage display technology has been announced as the recipient of Nobel Prize in Chemistry 2018. Phage display technique allows high affinity target-binding peptides to be selected from a complex mixture pool of billions of displayed peptides on phage in a combinatorial library and could be further enriched through the biopanning process; proving to be a powerful technique in the screening of peptide with high affinity and selectivity. In this review, we will first discuss the modifications in phage display techniques used to isolate various cancer-specific ligands by in situ, in vitro, in vivo, and ex vivo screening methods. We will then discuss prominent examples of solid tumor targeting-peptides; namely peptide targeting tumor vasculature, tumor microenvironment (TME) and over-expressed receptors on cancer cells identified through phage display screening. We will also discuss the current challenges and future outlook for targeting peptide-based therapeutics in the clinics.

OBJECTIVETo study the BRCA1 mutations in patients with early-onset breast cancer and their affected relatives in Guangdong province and explore the relationship between BRCA1 mutation and the expressions of estrogen receptor(ER), progesterone receptor(PR), HER2 and ALN.

METHODSFrom 58 patients with early-onset breast cancer and their affected relatives, the genomic DNA was extracted from the peripheral blood mononuclear cells and the coding regions of the BRCA1 gene was amplified using polymerase chain reaction. BRCA1 gene mutations were screened by denaturing high performance liquid chromatography (DHPLC) and subsequent direct DNA sequencing. The expression of ER, PR, HER2 and ALN were detected with immunohistochemistry and their relations with the gene mutation were analyzed.

RESULTSDisease-related BRCA1 mutations were detected in 2 of the 58 patients, who were younger than 35 years old, including 1 with a novel splice-site mutation (IVS5-1 G-->A). No association was found between this novel mutation and the expressions of ER, PR, HER2 and ALN.

CONCLUSIONThe incidence of BRCA1 mutation is significantly lower in patients with early-onset breast cancer and their affected relatives in Guangdong province than in the Western populations. The novel mutation identified in BRCA1 gene may represent a mutation characteristic of the patients in Guangdong province. BRCA1 gene mutations may not have any relation with the expression of ER, PR, HER2 and ALN.

Adult ; Age of Onset ; Base Sequence ; Breast Neoplasms ; genetics ; China ; DNA Mutational Analysis ; Female ; Genes, BRCA1 ; Genotype ; Humans ; Molecular Sequence Data ; Mutation ; Receptor, ErbB-2 ; genetics ; Receptors, Estrogen ; genetics ; Receptors, Progesterone ; genetics