Johne's disease is a condition that refers to chronic granulomatous enteritis in ruminants. It is believed that survival and replication of Mycobacterium (M.) paratuberculosis in mononuclear phagocytes plays an important role in the pathogenesis of Johne's disease. However, it is not clear how M. paratuberculosis survives for long time periods in mononuclear phagocytes, nor is it clear which factors trigger multiplication of these bacilli and result in the development of Johne's disease. Investigating the intracellular fate of M. paratuberculosis is challenging because of its very slow growth (more than two months to form visible colonies on media). Existing animal models also have limitations. Despite those obstacles, there has been progress in understanding the intracellular survival tactics of M. paratuberculosis and the host response against them. In this review, we compare known aspects of the intracellular survival tactics of M. paratuberculosis with those of other mycobacterial species, and consider possible mycobactericidal mechanisms of mononuclear phagocytes.
Animals ; Leukocytes, Mononuclear/*microbiology ; Mycobacterium avium subsp. paratuberculosis/*physiology ; Phagocytes/*microbiology

The influencing factors on cord blood storage after collection and mononuclear cell separation as well as cryopreservation were studied. The mononuclear cell are separated from blood after blood collection, then cryopreserved and washed after thawed. Results showed that the cord blood kept at 4 degrees C or room temperature less than 24 hours after blood collection, mononuclear cell separated by hydroxyethylstarch and 2 centrifugations, mononuclear cell cryopreserved with 50% DMSO and autoplasma from cord blood as protectives and washing the cells after thawing. In conclusion, the optimal project in this study can effectively preserve cord blood mononuclear cells.
Blood Preservation ; Cell Separation ; methods ; Cryopreservation ; Fetal Blood ; cytology ; Humans ; Leukocytes, Mononuclear ; physiology

CKLF-like MARVEL transmembrane domain containing member/chemokine-like factor super family member (CKLFSF/CMTM) is a novel tumor suppressor gene. CMTM3 is broadly expressed in normal human tissues and evolutionary conserved,especially in testis,spleen,and some cells of peripheral blood mononuclear cells. However,its expression is undetectable or down-regulated in most carcinoma cell lines and tissues. Restoration of CMTM3 may inhibit the proliferation,migration,and invasion of carcinoma cells. Although the exact mechanism of its anti-tumor activity remains unclear,CKLFSF3/CMTM3 is closely connected with immune system and associated with sex during tumorigenesis. The study advances of CKLFSF3/CMTM3 are elaborated in this review as CMTM3 may be a new target in the gene therapies for tumors,especially genitourinary tumors,while further studies on CMTM3 and its anti-tumor mechanisms are warranted.
Cell Transformation, Neoplastic ; Chemokines ; genetics ; physiology ; Down-Regulation ; Humans ; Leukocytes, Mononuclear ; MARVEL Domain-Containing Proteins ; genetics ; physiology ; Male ; Neoplasms ; pathology

For in vitro studies, both CD34+ selected cell and mononuclear cell (MNC) can be used to expand hematopoietic stem/progenitor cells. To investigate the expansion characteristics of mononuclear cells (MNC) and CD34+ selected cells the two cell fractions were cultured in the medium containing cytokine cocktails of SCF + IL-3 + IL-6 + FL + Tpo. It was found that the CD34+ selected cells had presented a high proliferation potential. The expansion of CD34+ selected cells could be maintained for 8 weeks while that of MNCs declined after 4 weeks. During the culture period, the maximum expansion of total cells in CD34+ selected cell culture achieved 31,270.9 +/- 8640.5 times, while that of MNC reached 50.9 +/- 8.2 times only. In the culture of MNCs, the colony density and the proportion of CD34+ cells increased from day 0 to day 7. However, in the culture of CD34+ selected cells, both the colony density and the proportion of CD34+ cells declined continuously during the whole culture period. During the ex vivo culture of CD34+ selected cells, the maximum expansion of CFU-GM and CD34+ cells achieved 185.7 +/- 14.1 fold and 191.7 +/- 188.8 fold, respectively. They are much higher than that of MNC, which were 12.4 +/- 3.2 fold and 50.6 +/- 33.2 fold only. While the BFU-E of both cell fractions only expanded by few times, which were 7.2 +/- 5.2 and 10.1 +/- 3.4 times, respectively. The results showed that the CD34+ selected cells culture could obtain more CFU-GM cells and CD34+ cells during the whole culture period.
Antigens, CD34 ; analysis ; Cell Count ; Cell Separation ; Cells, Cultured ; Fetal Blood ; cytology ; Hematopoietic Stem Cells ; physiology ; Leukocytes, Mononuclear ; cytology

The purpose of this study was to observe whether human peripheral dervied monouncleas cells (hMNCs) could participate in the regeneration process of the ischemic hearts in the way of differentiating into cardiomyocytes, vascular endothelial cells and smooth muscle cells. hMNCs were transplanted into the bodies of the mice with myocardial infarction through the tail vein injection. Hearts were harvested 2-12 weeks after injection then sliced up into frozen sections of 5 micron thickness. Double immunofluorescence staining was used to test the differentiation of the grafted cells into cardiomyocytes, smooth muscle cells and vascular endothelial cells which revealed that cells expressing both HLA and TNT, HLA and alpha-SMA, HLA and vWF existed in the hearts of the mice. According to the study, it is probable that hMNCs could participate in the regeneration process of the infarcted hearts in the way of differentiation.
Animals ; Cell Differentiation ; physiology ; Humans ; Leukocytes, Mononuclear ; transplantation ; Mice ; Mice, Nude ; Myocardial Infarction ; pathology ; therapy ; Myocytes, Cardiac ; cytology ; Transplantation, Heterologous

OBJECTIVEThe abnormality of hemopoietic inductive microenvironment (HIM) is involved in the pathophysiology of aplastic anemia (AA). Mesenchymal stem cells (MSC) are main source of bone marrow stromal cells which constitute the bone marrow HIM. Thus, the bone marrow failure in AA may be related to the function of MSC. The aim of the study was to investigate the hematopoiesis support function of MSC in children with AA in vitro.

METHODSBone marrow samples were collected from 24 children with AA at diagnosis and 19 children with idiopathic thrombocytopenic purpura (ITP), infectious mononucleosis or lymphadenitis (controls). MSCs from bone marrow samples were isolated, cultured and expanded. Morphology, proliferation activity and colony forming unit-fibroblast (CFU-F) were measured. The ability of bone marrow MSC to adhere hemopoietic cells was assayed by MTT. The concentration of stem cell factor (SCF) released from MSC was tested using ELISA. Mononuclear cells (MNC) of bone marrow were plated onto a feeder layer formed by MSC. Cells count and BFU-E, CFU-GM, CFU-GMME productions were measured.

RESULTSThe first and third passage time of MSC in children with AA was longer than that in the controls. The number of CFU-F in children with AA (15.70+/-5.78) was less than that in the controls (21.73+/-5.74) (P<0.05). The concentration of SCF in MSC supernatants in children with AA (30.69+/-16.82 pg/mL) was significantly lower than the controls (50.74+/-14.83 pg/mL) (P<0.01). The total MNC count and the number of BFU-E, CFU-GM and CFU-GMME colonies in the support of MSC in children with AA were significantly lower than those in the controls (P<0.01).

CONCLUSIONSThe hematopoiesis support function of MSC was significantly reduced in children with AA in vitro. The decreased hematopoiesis support function of MSC may be related its decreased proliferation capacity and SCF release activity.

Adolescent ; Anemia, Aplastic ; physiopathology ; Cell Adhesion ; Child ; Child, Preschool ; Female ; Hematopoiesis ; Humans ; Leukocytes, Mononuclear ; physiology ; Male ; Mesenchymal Stromal Cells ; physiology ; Stem Cell Factor ; physiology

Regulation of P2X7 receptor expression is of interest because activation of this receptor by extracellular ATP triggers a wide variety of cell functions in leukocytes. However, its expression and modulation in human peripheral blood mononuclear cells (PBMC) and monocytes remain unclear. RT-PCR was used to detect the constitutive level of P2X7 receptor and the levels upon stimulation with bacteria, bacterial product, mitogen and various cytokines in human PBMC and monocytes. P2X7 receptor mRNA was detected in PBMC and monocytes. P2X7 receptor expression in PBMC was up-regulated by interleukin-2, -4, -6 (IL-2, IL-4, IL-6) tumour necrosis factor-alpha (TNF-alpha), lipopolysaccharide (LPS) and heat-inactivated Staphylococcus aureus Cowan strain I (SAC). However, interferon-gamma (IFN-gamma), granulocyte-macrophage colony-stimulating factor (GM-CSF), macrophage colony-stimulating factor (M-CSF) and phytohemagglutinin-M (PHA-M) had little effect on the expression of P2X7 receptor. Furthermore, LPS and M-CSF could up-regulate P2X7 receptor expression in monocytes, while IFN-gamma, TNF-alpha and GM-CSF had weak effects, but pretreatment with these inducers could not further enhance LPS-stimulated P2X7 receptor expression in monocytes. The results obtained demonstrate that inflammatory stimuli drive P2X7 expression, thus supporting the hypothesis that P2X7 receptor may play a role in the inflammatory responses against bacteria infection, which need further verification.
Humans ; Interleukin-2 ; physiology ; Interleukin-4 ; physiology ; Leukocytes, Mononuclear ; drug effects ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Receptors, Purinergic P2X7 ; genetics ; metabolism ; Tumor Necrosis Factor-alpha ; physiology

OBJECTIVEThe pathogenesis of bronchiolitis has not been fully identified. Immune function abnormality following virus infection may be associated with the pathogenesis. CD40 and CD40 ligand (CD40L) is a pair of co-stimulatory molecules in immunoreaction. They might play an important role in the development of bronchiolitis. This study aimed to investigate the expression of CD40 and CD40L in peripheral blood mononuclear cells (PBMCs) in children with bronchiolitis and explore their possible roles in the disease.

METHODSThirty children with bronchiolitis, 26 children with bronchopneumonia and 30 healthy children (control) were enrolled. Flow cytometry was used to detect CD40 and CD40L expression in PBMCs. Total serum IgE level was measured using ELISA.

RESULTSCompared with the control group, CD40L expression significantly increased in the bronchiolitis and bronchopneumonia groups (P< 0.05). The CD40L expression in the bronchiolitis group was significantly higher than that in the bronchopneumonia group (P< 0.05). A significantly increased CD40 expression was also found in the bronchiolitis group when compared with the bronchopneumonia and the control group (P< 0.01). Total serum IgE level in the bronchiolitis group was significantly higher than the bronchopneumonia and the control groups (P< 0.01). CD40 and CD40L expression was positively correlated with serum IgE level in the bronchiolitis group (r=0.607, r=0.819, respectively; P< 0.01).

CONCLUSIONSCD40 and CD40L expression in PBMCs and serum IgE level increased and there is a positive correlation between CD40 and CD40L expression and serum IgE level in children with bronchiolitis. Over-expression of CD40 and CD40L may play an important role in the development of bronchiolitis.

Bronchiolitis ; etiology ; immunology ; CD40 Antigens ; blood ; physiology ; CD40 Ligand ; blood ; physiology ; Female ; Humans ; Immunoglobulin E ; blood ; Infant ; Leukocytes, Mononuclear ; chemistry ; Male

To study and compare the immunomodulatory functions of human bone marrow mesenchymal stem cell (MSC) and cord blood mononuclear cell (CBMNC) in vitro, human bone marrow MSC were separated with Percoll (1.073 g/L) and cultured in low-glucose DMEM, and cord blood mononuclear cells were isolated with Ficoll (1.077 g/L). These two kinds of cells were added to mixed lymphocyte cultures and PHA induction transformation cultures with various concentrations. The proliferation of lymphocytes was measured by MTT method, effects of MSC and CBMNC on mixed lymphocyte response and PHA-induced lymphocyte transformation were investigated. The results showed that both 5 x 10(4), 1 x 10(4) MSC and 2 x 10(5) CBMNC could inhibit the mixed lymphocyte response (MLR) and PHA induced transformation. But with lower concentrations (MSC < or = 1 x 10(3), CBMNC < or = 1 x 10(5)), the inhibition effects of MSC and CBMNC were less consistent. 1 x 10(2) MSC and 1 x 10(4) CBMNC mainly increased the lymphocyte activation. In addition, the inhibition ratio of 5 x 10(4) MSC (MLC 65.3%, PHA 79.1%) was higher than that of 2 x 10(5) CBMNC (MLC 8.6%, PHA 37.3%). It is concluded that larger numbers of both MSC and CBMNC showed negative immunomodulatory functions and inhibited the mixed lymphocyte response and induction of transformation in vitro. Moreover, the inhibitory effect of MSC was much stronger than that of CBMNC.
Bone Marrow Cells ; Fetal Blood ; cytology ; Humans ; Leukocytes, Mononuclear ; physiology ; Lymphocyte Activation ; Lymphocyte Culture Test, Mixed ; Mesenchymal Stromal Cells ; physiology ; Phytohemagglutinins ; pharmacology

Recent studies have shown that diet can affect the body's immunity. Roughage of dairy cows consists of a variety of plant materials which make different contributions to health. This study investigated the effect of different roughages on the immunity of dairy cows. Serum, peripheral blood mononuclear cells (PBMCs), and milk samples were collected from 20 multiparous mid-lactation cows fed mixed forage (MF)- or corn straw (CS)-based diets. Expression profile analysis was used to detect the differentially expressed genes (DEGs) from PBMCs. The results showed that milk protein in the MF group increased to 3.22 g/100 ml, while that of the CS group milk was 2.96 g/100 ml; by RNA sequencing, it was found that 1615 genes were differentially expressed between the CS group and the MF group among the 24 027 analyzed probes. Gene ontology (GO) and pathway analysis of DEGs suggested that these genes (especially genes coding cytokines, chemokine and its receptors) are involved in the immune response. Results were confirmed at the protein level via detecting the levels of interleukin-2 (IL-2), IL-6, IL-10, IL-12, leptin (LEP), interferon-γ (IFN-γ), transforming growth factor-β1 (TGF-β1), and tumor necrosis factor-α (TNF-α) in peripheral blood by enzyme-linked immunosorbent assay (ELISA) and radioimmunoassay analysis. Our data supported the conclusions that the protein content in milk of the MF group was higher than that of the CS group, the CS-based diets induced more release of cytokines than the MF-based diets in dairy cows' PBMCs, and milk protein content may be affected by cytokines.
Animals ; Cattle/immunology* ; Cytokines/physiology* ; Diet ; Female ; Gene Ontology ; Leukocytes, Mononuclear/immunology* ; Milk/chemistry* ; Transforming Growth Factor beta/physiology* ; Zea mays