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The article discusses a study on the enhancing effect of Mekabu extract to immunoglobulin (Ig) production of mouse spleen lymphocytes. Topics discussed include fucoidan as a promising candidate for bioactive component contained in Mekabu which is described as a fucose-rich sulfated polysaccharides and sort of a water-soluble dietary fiber and findings that Mekabu extract has significant enhancing effect on the production of antibody by mouse spleen lymophocytes.

The article discusses the outcome of lactic acid bacteria (LAB) which was quarantined from eight samples of commercially available Nozawana-zuke, a traditional Japanese pickle, on cytokine formulation by mouse spleen cell cultures. In this study, the author concluded that some LAB heightened the cytokine expression in a mouse spleen cell culture.

The trichothecene mycotoxin deoxynivalenol (DON) targets the innate immune system and is of public health significance because of its frequent presence in human and animal food. DON-induced proinflammatory gene expression and apoptosis in the lymphoid tissue have been associated with a ribotoxic stress response (RSR) that involves rapid phosphorylation of mitogen-activated protein kinases (MAPKs). To better understand the relationship between protein phosphorylation and DON’s immunotoxic effects, stable isotope dimethyl labeling–based proteomics in conjunction with titanium dioxide chromatography was employed to quantitatively profile the immediate (≤ 30min) phosphoproteome changes in the spleens of mice orally exposed to 5mg/kg body weight DON. A total of 90 phosphoproteins indicative of novel phosphorylation events were significantly modulated by DON. In addition to critical branches and scaffolds of MAPK signaling being affected, DON exposure also altered phosphorylation of proteins that mediate phosphatidylinositol 3-kinase/AKT pathways. Gene ontology analysis revealed that DON exposure affected biological processes such as cytoskeleton organization, regulation of apoptosis, and lymphocyte activation and development, which likely contribute to immune dysregulation associated with DON-induced RSR. Consistent with these findings, DON impacted phosphorylation of proteins within diverse immune cell populations, including monocytes, macrophages, T cells, B cells, dendritic cells, and mast cells. Fuzzy c-means clustering analysis further indicated that DON evoked several distinctive temporal profiles of regulated phosphopeptides. Overall, the findings from this investigation can serve as a template for future focused exploration and modeling of cellular responses associated with the immunotoxicity evoked by DON and other ribotoxins. [ABSTRACT FROM PUBLISHER]

Mouse spleen cells were cultured for 4 days in RPMI 1640 medium with 5% fetal calf serum. The neutral proteinases trypsin and plasmin, and bacterial lipopolysaccharide LPS, all polyclonal B lymphocyte activators, stimulated the development of immunoglobulin producing cells as detected by the protein A plaque assay. At the same time, direct plaque forming cells reacting with mouse, human and rabbit IgG and the Fc fragment of human IgG were induced by the stimulants. The plaques could be inhibited by free IgG or Fc fragment. In the culture supernatants, IgM and IgM anti-IgG antibodies were detected by enzyme linked immunosorbant assays. Both general IgM and 1gM anti-IgG antibodies increased under the influence of the proteinases and of LPS. The results are discussed in relation to rheumatoid factor production during inflammatory diseases. [ABSTRACT FROM AUTHOR]

Mouse spleen cells produce haemagglutinating activity in culture which is species specific but not strain specific. The activity is produced by all strains of mice tested including nude mice. It is produced in cultures of spleen and lymph node but not of peritoneal exudate cells. Production of the factor is not affected by macrophage depletion of the spleen. This activity is not antibody mediated. [ABSTRACT FROM AUTHOR]

A null cell line (SCM1) was established by a culture of spleen cells (SC) from normal adult C57B1/6 mice with complete medium alone for 10 days and followed by weekly cultures with a 25% WEHI-3 cell culture supernatant. Phenotype analysis showed that the SCM1 cells were negative for CD3, Tbyl.2. B220. Mac-l,Gr-1, NK1.1 and MHC class II, but were positive for MHC class I. FcγRII/III. FcγRII. c-kit and the receptor against wheat germ agglutinin. These findings suggested that the SCM1 cells were mast cells. In an in vitro proliferation assay. SCM1 cells proliferated in the presence of either IL-3 or stem cell factor (SCF), but not in the presence of IL-4, whereas IL-4 showed an augmenting effect on their proliferation in the presence of either lL-3 or SCF. In analysing the mechanism by which such mast cells could be expanded from normal adult mouse SC, the addition of anti-IL-1 MoAb, but not anti-SCF MoAb, into the initial culture inhibited the subsequent expansion of either IL-3-or SCF-responding cells. The prior depletion of CD4+ T cells abrogated the capacity of the SC to enhance the expansion of SCF-responding cells, and this inability was restored by the addition of IL-3. Moreover, the culture supernatant of normal adult SC alone contained considerable levels of IL-3. Taken together, our findings suggest that, in an in vitro culture. CD4+ T cell-derived IL-3 therefore enhances the expansion of mast cells from the normal adult mouse spleen. [ABSTRACT FROM AUTHOR]

Interleukin-2 (IL-2) augments natural killer (NK) activity as well as generating effector cells named lymphokine activated killer cells (LAK) which are capable of lysing a wide spectrum of target cells. A large body of evidence has been accumulated to evaluate the relationship between NK and LAK cells and conflicting results have been reported. Our study was addressed lo further analyse this relationship and in particular to investigate whether in a short incubation IL-2 is merely capable of augmenting the activity of pre-existing killer cells, or whether it can also promote the differentiation of precursor cells. Eighteen-hour culture of mouse spleen cells in human recombinant IL-2 induced a DNA-synthesis-independent generation of cytotoxic cells bearing an NK phenotype (aGM-1+, Thy 1.2±, CD8-, CD4-). These were generated from precursor cells also bearingan NK phenotype, recovered either from low density Percoll fractions enriched in lytic cells with LGL morphology as well as from high density fractions devoid of LGL and cytotoxic activity. [ABSTRACT FROM AUTHOR]

An experimental system is described, based on the use of 51Cr-labelled L cells, permitting the measurement of cell detachment and of 51Cr release by cells. This system is used to quantify detachment of, and 51 release by L cells subjected to normal mouse spleen cells. A detachment of L cells is thus observed. It is mediated by (a) soluble factor(s). It may be produced by normal non-immunized non-stimulated lymphocytes, but non-lymphoid nucleated cells may have the same effect. It is increased by heparin and Trypan blue. Possible implications of these observations for the interpretation of results of experiments on cell-mediated cytotoxicity are discussed. [ABSTRACT FROM AUTHOR]

In order to differentially test the function of lymphocytes in Tonoplasma gondii-incfected mice, the in vitro blastogenic response of spleen cell cultures to non-specific mitogens was studied. Phytohaemagglutinin (PHA) and concanavalin A (Con A) stimulation were used a stests of thymus-dependent lymphocyte (T cell) function and bacterial endotoxin lipopolysaccharide (LPS) was used as a probe of bursal equivalent lymphocyte (B cell) function. For the first 3 weeks following T. gondii infection, the uptake of tritiated thymidine ([2H]TdR) by spleen cells cultured with all three mitogens was markedly reduced to comparison to the uptake in spleen cells from uninfected control mice. Thereafter, the response to LPS returned to normal while stimulation by the T-cell mitogens (PHA and Con A) remained depressed. It is postulated that T. gondii infection either: (T) diluted out T cells in the spleen with unreactive cells: (2) modified T cells in such a way that they were less responsive to mitogens: (3) depleted the peripheral lymphoid tissues of T cells: (4) induced non-specific suppressor cells, which inhibited the T-cell function assays: or (5) activated macrophages which depressed T-cell function non-specifically. [ABSTRACT FROM AUTHOR]

The migration of specifically sensitized mouse spleen cells following exposure to allogeneic cells (antigen) in vitro was studied. The migration inhibition recorded when sensitized cells were admixed with allogeneic cells in capillary tubes (mix method) was compared to the inhibition observed when allogeneic cells were suspended in the culture chamber media (non-mix method). Specificity as well as a higher degree of migration inhibition were obtained using the mix method, suggesting that this method is superior to the non-mix method. [ABSTRACT FROM AUTHOR]

In order to study three-dimensional fine architecture of the red pulp, especially the relationships between the splenic sinuses and the cordal capillaries, mouse spleens were perfusion-fixed, loading normal arterial and venous hydrostatic pressures via the abdominal aorta and the portal vein. The fixed splenic tissue were alcohol-freeze-fractured and specimens were processed routinely for scanning electron microscopy (SEM). The normal red pulp of the mouse spleen consists, like spleens of many other mammals, of the splenic cords formed by a three-dimensional network of reticular tissue and of the splenic sinuses separated by areas of the splenic cords. The inner surface of the sinus wall is covered with a lining of irregularly shaped flat cells with scattered nuclear protuberances. The outer surface of the sinus wall is covered with attenuated reticular cell processes supporting the sinus. The wall of the splenic sinus is perforated with round or elliptical stomata 1 to 3 μm in diameter through which the sinus lumen communicates with the cordal labyrinthine space. In comparison with other mammals such as human, dog and rat, stomata are rather few and distributed irregularly. It was observed that a large number of cells, most likely macrophages, are present fixed to the network of the cordal reticulum. The inner surface of the cordal capillary is lined by a relatively smooth endothelial sheet with a few short and small granular microvilli. The outside of the capillary is also covered with the reticular cell process. In the wall of the cordal capillary approaching the terminal are found a few openings through which the capillary lumen communicates with the cordal space. It was observed in the terminal that these openings become larger rather suddenly and the wall transforms into the network of the cordal reticular tissue. No image which may lead us to the conclusion that the cordal capillary connects directly with the splenic sinus was recognized. These findings strongly support the concept of “open blood circulation” in the red pulp of the mouse spleen. [ABSTRACT FROM PUBLISHER]

This article reports that the formation of antibodies represents a special case of induced cell differentiation. It has been recently reported that the complex series of events triggered by the injection of antigen includes dramatic changes in activities of several enzymes of nucleotide metabolism in the spleen. It is informed that a specific immune response can be suppressed by passive transfer of the corresponding antibody. To get additional information on the mechanism of this inhibition, researchers studied effects of the passive transfer of antibodies upon changes in activity of these enzymes induced by different doses of antigen. The study conducted showed that the specific activity of uridine kinase and thymidine kinase increased in the particle-free extract of mouse spleen after primary and secondary injections of sheep red blood cells.

This article focuses on the specific activity of several enzymes of nucleotide metabolism in mouse spleen after immunization. After the injection of antigen into the mammalian host. a complex series of events begins which culminates in the synthesis of antibody. These events include the activation of latent information stored in the genomic of the organism for new RNA synthesis as well as an increase in the rate of DNA synthesis. To learn more of the early events alter exposure of the precursor cells to antigen, researchers determined the specific activities of several enzymes involved in nucleotide metabolism found after immunization in the spleens of mice.

Low-virulent Orientia tsutsugamushi (Ot) were successfully isolated from scrub typhus patients in Shandong, China, and the isolates were similar to the Kawasaki type identified by nested polymerase chain reaction (PCR). To identify the morphological characterization of the low-virulent Ot, and elucidate the pathological changes on host cells, mouse spleen tissue infected with the Ot isolated from Shandong was used for the ultrastructural study. Transmission electron microscopy showed that the Ot parasitized in the spleen were different in size, shape and electron density and many significant changes occurred in cytoplasmic organelles of the inoculated mouse spleen cells. Swollen perinuclear cisterna was observed in the nuclear membranes of mononuclear cells and a multivesicular body was found in the intracytoplasm of the macrophage. In the phagosome of the macrophage, many Ot enveloped with an additional membrane were found to push the phagosomal membrane outward from inside. The results indicated that the low-virulent Ot and the spleen cells suffered various damages. [ABSTRACT FROM AUTHOR]

BALB/c spleen cells were stimulated with C57B1/6 cells, keyhole limpet haemocyanin (KLH), pokeweed mitogen (PWM) or phytohaemagglutinin (PHA). By the immunofluorescence method the development of transformed cells with immunoglobulin-like receptor molecules or theta antigen was investigated. Transformed Ig-receptor cells were predominant after stimulation with KLH, followed by PWM, PHA and allogeneic cells in decreasing order. Transformed cells with theta-antigen were clearly increased after stimulation with allogeneic cells. With all systems, more transformed cells with Ig-receptors were found in the stimulated cultures than in the control cultures, indicating that all stimulants have an effect on both cell types. The implications of the results on the immunologic interpretation of results obtained by in vitro stimulation of lymphocytes with the different stimulants are discussed. [ABSTRACT FROM AUTHOR]

Early after infection, mice inoculated with coxsackievirus B3 showed consistent reduction of antibody responsiveness. Beginning one week after infection they also evidenced progressive spleen atrophy. The cellular basis of the reduced antibody response exhibited in vitro by spleen cells of infected mice at the onset of atrophy was investigated by the use of different antigens, by supplementation with different subpopulations of immunocompetent cells from normal donors, and by cross-recombination with normal lymphoid cells. Whereas B cell functions appeared to be preserved, a deficit of macrophage accessory functions was clearly evident, possibly at the level of antigen presentation. In infected spleens, nonspecific suppressor T cells were also observed, but their activity did not correlate strictly with the degree of immunodepression. Becauseno evidence for a direct effect of virus on, or for viral replication in, immunocompetent cells was found, these alterations were tentatively ascribed to activation of the host's suppressor system. Such changes may have implications in the pathogenesis of coxsackievirus infections. [ABSTRACT FROM PUBLISHER]

Intratumour heterogeneity has been shown to play a role in the malignant progression of cancer. The clonal evolution in primary cancer has been well studied, however, that in metastatic tumorigenesis is not fully understood. In this study, we established human colon cancer-derived organoids and investigated clonal dynamics during liver metastasis development by tracking barcode-labelled subclones. Long-term subclone co-cultures showed clonal drift, with a single subclone becoming dominant in the cell population. Interestingly, the selected subclones were not always the same, suggesting that clonal selection was not based on cell intrinsic properties. Furthermore, liver tumours developed by co-transplantation of organoid subclones into the immunodeficient mouse spleen showed a progressive drastic reduction in clonal diversity, and only one or two subclones predominated in the majority of large metastatic tumours. Importantly, selections were not limited to particular subclones but appeared to be random. A trend towards a reduction in clonal diversity was also found in liver metastases of multiple colour-labelled organoids of mouse intestinal tumours. Based on these results, we propose a novel mechanism of metastasis development, i.e. a subclone population of the disseminated tumour cells in the liver is selected by neutral selection during colonization and constitutes large metastatic tumours. [ABSTRACT FROM AUTHOR]

Objectives The aim was to prepare novel Ganoderma lucidum polysaccharide nanoparticles and to evaluate the physicochemical properties and anti-tumour activity in in-vitro cytotoxicity studies using HepG2, HeLa and A549 cancer cell lines, and growth promotion effects on mouse spleen cells. Methods Chitosan nanoparticles loaded with G. lucidum polysaccharide were prepared using the ion-revulsion method. The diameter distribution of the particles and the surface charge were measured using a zetasizer analyser. The entrapment efficiency and drug loading capacity were examined by the diethylaminoethanol weak anion exchange method. The cytotoxic effects of nanoparticles on tumour cells and the growth promotion effects on mouse spleen cells were tested using the MTT assay. Key findings Nanoparticles loaded with G. lucidum polysaccharide at 6 μg/ml and chitosan/sodium tripolyphosphate (mass) ratio of 5.5 had significantly greater cytotoxic effects on tumour cells and growth promotion effects on mouse spleen cells than empty nanoparticles. Conclusions G. lucidum polysaccharide nanoparticles showed significant anti-tumour efficacy, having both cytotoxic effects on tumour cells and growth promotion effects on spleen cells, making it a promising candidate in the clinical setting. [ABSTRACT FROM AUTHOR]

Graft versus host disease (GvHD) is a major clinical problem with a significant unmet medical need. We examined the role of cytotoxic T lymphocyte antigen‐4 (CTLA‐4) in a xenogenic GvHD (xeno‐GvHD) model induced by injection of human peripheral mononuclear cells (hPBMC) into irradiated non‐obese diabetic (NOD) SCID gamma (NSG) mice. Targeting the CTLA‐4 pathway by treatment with CTLA‐4 immunoglobulin (Ig) prevented xeno‐GvHD, while anti‐CTLA‐4 antibody treatment exacerbated the lethality and morbidity associated with GvHD. Xeno‐GvHD is associated with infiltration of hPBMCs into the lungs, spleen, stomach, liver and colon and an increase in human proinflammatory cytokines, including interferon (IFN)‐γ, tumor necrosis factor (TNF)‐α and interleukin (IL)‐5. Infiltration of donor cells and increases in cytokines were attenuated by treatment with CTLA‐4 Ig, but remained either unaffected or enhanced by anti‐CTLA‐4 antibody. Further, splenic human T cell phenotyping showed that CTLA‐4 Ig treatment prevented the engraftment of human CD45+ cells, while anti‐CTLA‐4 antibody enhanced donor T cell expansion, particularly CD4+ (CD45RO+) subsets, including T box transcription factor TBX21 (Tbet)+ CXCR3+ and CD25+ forkhead box protein 3 (FoxP3) cells. Comprehensive analysis of transcriptional profiling of human cells isolated from mouse spleen identified a set of 417 differentially expressed genes (DEGs) by CTLA‐4 Ig treatment and 13 DEGs by anti‐CTLA‐4 antibody treatment. The CTLA‐4 Ig regulated DEGs mapped to down‐regulated apoptosis, inflammasome, T helper type 17 (Th17) and regulatory T cell (Treg) pathways and enhanced Toll‐like receptor (TLR) receptor signaling, TNF family signaling, complement system and epigenetic and transcriptional regulation, whereas anti‐CTLA‐4 antibody produced minimal to no impact on these gene pathways. Our results show an important role of co‐inhibitory CTLA‐4 signaling in xeno‐GvHD and suggest the therapeutic utility of other immune checkpoint co‐inhibitory pathways in the treatment of immune‐mediated diseases driven by hyperactive T cells. [ABSTRACT FROM AUTHOR]

Human peripheral blood leucocytes, cultured for 4-6 days with the appropriate concentration of protein or synthetic polypeptide antigen, yield helper cells. These cells, after a further 24 hr incubation with the appropriate antigen release helper factors, which are of two types, antigen specific and non-specific; a similar situation to that described in the mouse. These factors are assayed by their effects on the plaque-forming cell response of mouse spleen cells cultures in vitro. The potential applications of this new assay for human T cell function are discussed. [ABSTRACT FROM AUTHOR]

The luminol-enhanced chemiluminescence (CL) assay was used to examine the effects of antifungal agents tested at concentrations above and below therapeutically achievable levels on the CL response of mouse spleen cells. Reduction in the CL response of phagocytic cells may be indicative of an inhibition of the cellular immune response. Concomitantly, an increase in the CL response of phagocytic cells may indicate an enhancement of the immune capacity of these cells. The effects of four antifungal agents, bifonazole, fluconazole (UK-49,858), itraconazole, and terbinafine were studied. Changes in the CL response were assessed in terms of peak intensity, time to peak intensity, and area under the intensity-time curve compared with appropriate diluent controls for each drug. Both bifonazole and itraconazole caused significant reduction in peak CL intensity only at the highest level assayed (20 mg/l). Fluconazole had no significant effect on the CL response of mouse spleen cells at levels up to 20 mg/l, inclusive. Although terbinafine had no significant effect on peak CL intensity, it did cause a significant decrease in time to peak response at levels above 5 mg/l. This decrease in time to peak response may be indicative of an enhancement in the immune capacity of the mouse spleen cells; the clinical significance of this observation remains to be determined. [ABSTRACT FROM AUTHOR]

C57BL/6 were infected intravenously with 107 Mycobacterium lepraemurium (MLM). At increasing time intervals after infection different isolated splenic cell subpopulations were tested for their ability to suppress the mixed lymphocyte reaction (MLR) of normal syngeneic mouse splenocytes. During the first 6 months after infection neither T depleted nor plastic adherent spleen cells from infected mice exerted a suppressive activity on the normal mouse allogeneic proliferative response. Conversely, splenic T cells from MLM infected mice exhibited suppressive activity as early as 2 months after infection. Attempts to characterize the Lyt phenotype of splenic suppressor T cells from 6 months infected mice showed that both Lyt 1 +2- and Lyt 2+ enriched cell subsets possessed the ability to suppress the MLR of the normal mouse spleen cells and Lyt 1 +2- T cells were shown to be more efficient suppressors than Lyt 2+ cells. [ABSTRACT FROM AUTHOR]

In vitro studies were conducted in the non-obese diabetic (NOD) mouse, prone to Type 1 autoimmune diabetes, to investigate the mechanisms involved in cell-mediated rejection of pig islet xenografts. Our previous work concerning the mechanisms of proliferation of xenogeneic lymphocytes to pig islet cells (PIC) was not indicative of PIC impairment. Consequently, a test was developed based on perifusion analysis of the alteration of basal and stimulated insulin release from adult PIC incubated with mouse splenocytes or subsets. Compared with PIC incubation alone or with syngeneic pig splenocytes, co-incubation with mouse whole spleen cells resulted in a decrease of basal and stimulated insulin release (P < 0·001). Two components of this alteration were detected separately: PIC impairment was decreased (P < 0·01) after removal of plastic-adherent cells from spleen cells, but maintained (P < 0·01) when plastic-adherent cells alone were co-incubated with PIC. The increase of murine interleukin-1β when mouse plastic-adherent spleen cells were cultured with PIC (P < 0·04) was indicative of macrophage activation. Soluble factors produced during co-incubation of mouse splenocytes or plastic-adherent cells with PIC were involved in the impairment process, since supernatant fluids collected during previous PIC–mouse cell co-incubations directly altered (P < 0·01) insulin release from PIC. Moreover, impairment of PIC by mouse spleen cells was abolished (P < 0·01) by gadolinium chloride (which inhibits macrophages), but not by cyclosporin A. Another mechanism was apparent, since co-incubation of PIC with purified mouse T cells or CD4+ T cells, re-mixed with antigen-presenting cells, led to a decrease (P < 0·01) of insulin release. This model, based on the alteration of dynamic basal and stimulated insulin release, is indicative of in vitro cell-mediated alteration of PIC in the NOD mouse. The effect of whole spleen cells was rapid, and a crucial role was played by plastic-adherent cells. Two mechanisms were responsible for the behaviour of these cells: an early direct effect (at least in part via soluble products); and the indirect presentation of PIC xenoantigens (leading to impairment by CD4+ T lymphocytes). [ABSTRACT FROM AUTHOR]

Single-cell multi-omics integration enables joint analysis at the single-cell level of resolution to provide more accurate understanding of complex biological systems, while spatial multi-omics integration is benefit to the exploration of cell spatial heterogeneity to facilitate more comprehensive downstream analyses. Existing methods are mainly designed for single-cell multi-omics data with little consideration of spatial information and still have room for performance improvement. A reliable multi-omics integration method designed for both single-cell and spatially resolved data is necessary and significant. We propose a multi-omics integration method based on dual-path graph attention auto-encoder (SSGATE). It can construct the neighborhood graphs based on single-cell expression profiles or spatial coordinates, enabling it to process single-cell data and utilize spatial information from spatially resolved data. It can also perform self-supervised learning for integration through the graph attention auto-encoders from two paths. SSGATE is applied to integration of transcriptomics and proteomics, including single-cell and spatially resolved data of various tissues from different sequencing technologies. SSGATE shows better performance and stronger robustness than competitive methods and facilitates downstream analysis. [ABSTRACT FROM AUTHOR]

Heat-inactivated A. laidlawii (AL) was found to be a potent mitogen for mouse spleen cells. Spleen cells from homozygous nude mice and spleen cells treated with anti-theta serum and complement responded as well as their respective controls, indicating that AL represented a B-cell mitogen for mouse spleen cells. Spleen cells from LPS-unresponsive C3H/HeJ mice responded well to AL. The peripheral blood leucocytes from unselected human donors were also stimulated by AL, which appeared to represent a T- cell mitogen for human leucocytes. However, the possibility that it acted as a specific antigen could not be excluded. Attention was drawn to the possibility that the presence of mycoplasma might considerably affect the results of tests where tissue-culture cells or derivatives thereof are added to leucocyte cultures. [ABSTRACT FROM AUTHOR]

We have investigated the requirements of mitogenic response of mouse spleen cells for free and cell-bound concanavalin A (Con A) and inhibition of the response by egg lecithin. We also studied 125I-Con A binding with the mouse cells at different times after stimulation with the lectin. Our results suggested that mitogenic response to Con A by mouse spleen cells may be regulated by two receptor-mediated mechanisms acting in concert. Mitogenic response was inhibited during the first 2 hr following stimulation with Con A by removing free Con A from culture medium. But after 4 hr. the response became independent of free (on A and. concomitantly. (`on A binding by mouse cell increased by about 2 3-fold. It was further shown that the increase in Con A binding depends on prior reaction of the cells with the lectin hut it does not, on de nora protein synthesis. Treatment of mouse cells with glutaraldehyde may also increase Con A binding. These results suggested that the initial Con A binding with mouse cells may not be sufficient to initiate mitogenic response. It may however mobilize those Con A receptors which have remained hitherto cryptic hut occupancy of which by Con A may he essential for triggering of mitogenic response. The second mechanism affects mitogenic response at a time after receptor mobilization anti before cells are committed to proliferation. The nature of this mechanism remains obscure but it requires Con A binding and is sensitive to inhibition by the membrane active substance. egg lecithin. [ABSTRACT FROM AUTHOR]

The use of monoclonal antibodies for the detection of cellular biomarkers during carcinogenesis provides new strategies for cancer diagnosis or prognosis in patients. Loss of the Restrictive Element 1-Silencing Transcription (REST) factor has been observed in previous molecular and immunological approaches in aggressive breast cancer, small cell lung cancer, liver carcinoma, and colo-rectal cancer; however, for clinic diagnosis, monoclonal antibodies for REST recognition are unavailable. The goal of this work was to design, produce and characterize monoclonal antibodies against the REST DNA binding damain (DBD) that would be suitable for immunoassays. We searched for conserved domains, and immunogenic and antigenic sites in the REST structure via in silico analysis. For mice immunization, we used a recombinant REST DBD purified by affinity chromatography, and then Hybridomas were generated by mouse spleen fusion with myeloma cells. Finally, for monoclonal antibody characterization, we performed enzyme-linked immunosorbent (ELISA), western blot, dot blot, immunocytochemistry (ICC) and immunoprecipitation assays. Results showed that the DBD is conserved in REST isoforms and contains immunogenic and antigenic sites. We generated three clones producing monoclonal antibodies against REST DBD, one of them specifically recognized native REST and was suitable for ICC in samples from patients. [ABSTRACT FROM AUTHOR]

Summary: Background: Plasmacytoid dendritic cells (pDCs) are involved in the pathogenesis of psoriasis by secreting interferon‐α. Vitamin D3 analogues are widely used to treat psoriasis, and the representative analogue calcipotriol (CAL) uniquely downregulates the cytokine production and chemotactic activity of pDCs. However, the molecular mechanism of action of CAL is not well understood. Aim: To investigate effects of CAL on the Toll‐like receptor 9–myeloid differentiation primary response gene 88 (TLR9‐MyD88) signalling pathway, which induces cytokine production, in murine pDCs. Methods: pDCs were isolated from mouse spleen cells by negative selection or were generated from mouse bone ‐marrow cells, and were stimulated with CpG‐oligodeoxynucleotide (ODN) with or without CAL for 24 h. mRNA expression of TLR9 and MyD88 was assessed by real‐time PCR, and the amount of TLR9 was measured by western blotting. Results: CAL suppressed the CpG‐ODN‐induced increased expression of MyD88 and TLR9 in pDCs. Conclusions: CAL may downregulate pDCs by inhibiting TLR9‐MyD88 signalling. [ABSTRACT FROM AUTHOR]

Graft versus host disease (GVHD) is a major complication of allogeneic hematopoietic stem cell transplantation, leading to significant morbidity and mortality. Host‐derived TNF‐α play a role in the induction of allo‐reactive donor T cell activation and the pathogenesis of GVHD. On the other hand, the precise role of donor‐derived TNF‐α in GVHD remains unclear. To elucidate this issue, we designed an acute GVHD model using (B6×D2) F1 recipient mice transferred with spleen cells derived from either wild‐type or TNF‐α–/– C57BL/6 mice. Surprisingly, we found that spleen cells from TNF‐α–/– mice induce more severe graft versus host reaction (GVHR) than wild‐type spleen cells upon transfer into B6D2F1 mice. Transplantation of TNF‐α–/– mouse spleen cells was associated with enhanced anti‐host CTL generation and augmented deletion of host cells. Moreover, mice receiving TNF‐α–/– cells showed significantly higher levels of serum IFN‐γ, which was mainly produced by donor CD8+ T cells. We also demonstrated that TNF‐α deficiency in donor spleen cells caused a marked elevation of TNF‐α producing capacity by LPS‐stimulated host macrophages. Such enhanced GVHR was completely prevented by using TNF‐α–/–IFN‐γ–/– splenic cells. Our findings demonstrate, for the first time, that donor‐derived TNF‐α suppress GVHR by inhibiting IFN‐γ‐dependent donor type‐1 immunity which is essential for host TNF‐α elevation. [ABSTRACT FROM AUTHOR]

Aims To elucidate the biological characteristics and stability of a newly identified staphylococcal enterotoxin Q ( SEQ) against heating and digestive enzymes and to evaluate the risk of seq-harbouring Staphylococcus aureus in food poisoning. Methods and Results Purified SEQ was treated with heating, pepsin and trypsin which are related to food cooking, stomach and intestine conditions, respectively. Superantigenic activity of SEQ was assessed by determining the ability of IL-2 induction in mouse spleen cells. The emetic activity of SEQ was assessed using house musk shrew, a small emetic animal model. The results revealed that SEQ exhibits a remarkable resistance to heat treatment and pepsin digestion and has significant superantigenic and emetic activities. Furthermore, a sandwich ELISA for detection of SEQ production was developed, and the results showed that seq-harboring S. aureus isolates produce a large amount of SEQ. Conclusions The newly identified SEQ had remarkable stability to heat treatment and digestive enzyme degradation and exhibited significant superantigenic and emetic activities. In addition, seq-harbouring S. aureus isolated from food poisoning outbreaks produced a large amount of SEQ, suggesting that seq-harbouring S. aureus could potentially be a hazard for food safety. Significance and Impact of the Study This study found, for the first time, that SEQ, a nonclassical SE, had remarkable stability to heat treatment and enzyme degradation and exhibited significant emetic activity, indicating that SEQ is a high-risk toxin in food poisoning. [ABSTRACT FROM AUTHOR]

Receptors for bioactive glycosylation-inhibiting factor (GIF) were demonstrated using a bioactive mutant of recombinant human (rh) GIF, which is comparable to the suppressor T (Ts) cell-derived bioactive GIF in its affinity for the receptors on helper T (Th) hybridoma cells. Both naive T and B cells in normal mouse spleen lacked GIF receptors. However, presentation of specific antigen to naive T cells resulted in the expression of the receptors on activated T cells. Furthermore, activation of small resting B cells with F(ab′)2 fragments of anti-mouse IgM plus IL-4, lipopolysaccharide (LPS) plus IL-4 or LPS plus dextran sulfate induced the expression of the receptors within 48 h of B cell stimulation. It was also found that NK T cells freshly isolated from mouse spleen, but not conventional NK cells, expressed receptors for GIF. CD4+ and CD4– subpopulations of NK T cells showed a similar binding capability. Mature dendritic cells derived from bone marrow did not bear the receptors. The dissociation constant (Kd) of the interaction between the bioactive rhGIF mutant and the high-affinity receptors was 10–100 pM, whereas inactive wild-type rhGIF failed to bind to the receptors. A bioactive derivative of rhGIF suppressed both IgG1 and IgE synthesis by purified B cells activated by LPS and IL-4, indicating that the binding of bioactive GIF to its receptors on activated B cells results in suppression of their differentiation. [ABSTRACT FROM PUBLISHER]

Activity of Adenosine deaminase (ADA), an enzyme known to be deficient in some patients with servere combined immunodeficiency, increased three-fold within a 24-hr exposure of human peripheral blood lymphocytes to phytohaemagglutinin (PHA) in culture. This increase took place before the onset of DNA synthesis. Increased levels of ADA activity were also observed in lymphocytes incubated with pokeweed mitogen (PWM) for 60 hr. DNA synthesis induced by PHA, PWM or mixed lymphocyte cultures (MLC) was strongly inhibited by adenosine at concentrations of 10-4M or higher when human peripheral blood lymphocytes were cultured in a medium supplemented with horse serum, which lacks ADA. 10-6-10-8M coformycin, a potent inhibitor of ADA, inhibited PHA-, PWM- and MLC-induced DNA synthesis to a variable extent, whereas thymidine incorporation induced by Salmonella lipopolysaccharide (LPS) in mouse spleen cell cultures was strongly inhibited (by 75% or more) by 10-6M coformycin. Combination of 10-7-10-8M coformycin and 10-4-10-5M adenosine synergistically inhibited mitogen- or MLC-induced DNA synthesis in human and mouse lymphocyte cultures. These results, together with observations on children with ADA deficiency, provide evidence that adenosine deaminase is highly important for lymphocyte proliferation. Human peripheral blood lymphocytes incubated with PHA, 10-5M adenosine and 10-7M coformycin showed some cytotoxicity whereas the rate of 51Cr release from normal lymphocytes was not modified by the drugs. These findings suggest that in vivo clones of lymphocytes responding to specific antigens might be eliminated by coformycin, which may prove to be useful as a specific immunosuppressive agent. [ABSTRACT FROM AUTHOR]

Levamisole has been reported to act as an immunological adjuvant. Experiments reported here on the effect of this agent on a variety of murine lymphoid culture systems were designed to gain an insight into its mechanism of action. We have found levamisole to be a weak mitogen for mouse spleen cells producing a dose related response which peaks at 48 hr in culture. The drug acted to augment the response of spleen cells to sub-optimal concentrations of concanavalin A, but had no unusual effect on the lipopolysaccharide stimulation of B-cell DNA synthesis in vitro. Levamisote was directly stimulatory on enriched T-cell populations and was found to have two actions: (1) to stimulate a subpopulation of T cells and (2) to augment the response of suboptimal mitogen concentrations of concanavalin A. In addition, we have found that murine thymocytes stimulated by concanavalin A were greatly potentiated in the presence of levamisole, but this population of cells could not be stimulated directly by the drug. [ABSTRACT FROM AUTHOR]

Mouse spleen and lymph node lymphocytes with receptors for complement (CRL) were prepared by a standard rosetting method and the identity of the central lymphocyte studied by indirect immunofluorescence with heterologous antisera specific for immunoglobulin and for T cells. About 30%, of CRL were shown to be T cells and the remainder immunoglobulin-bearing (B) cells. In thymus-less 'nude' mice a population of immunoglobulin-negative, non-T cell CRL was found. [ABSTRACT FROM AUTHOR]

Novel trypsin-like serine proteases (mouse trypsin-type serine proteases 1 and 2 [MTSP-1 and -2]) were purified to homogeneity from mouse spleen. Each protease consisted of a single polypeptide with a molecular mass of about 29 kDa, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis under reducing conditions. Both were totally inhibited by diisopropylfluorophosphate, soybean trypsin inhibitor, aprotinin, antipain, and leupeptin and partially inhibited by chymostatin and dithiothreitol, suggesting that they are trypsin-like serine proteases. They hydrolyzed synthetic substrates for trypsin-like proteases but not those for chymotrypsin-like proteases, elastase and kalli-krein. MTSP-1 hydrolyzed terf-butyloxycarbonyl (Boc)-Asp(OBzl)Pro-Arg-amino-4-methyl-coumaryl-7-amide (MCA) and Boc-De-Glu-Gly-Arg-MCA faster than Boc-Phe-Ser-Arg-MCA. On the other hand, MTSP-2 hydrolyzed Boc-Phe-Ser-Arg-MCA most rapidly, with a specific activity 15 times higher than that of MTSP-1. The N-terminal amino acid sequence of MTSP-1 was Ile-Val-Gly-Gly-Tyr-Thr-His-Leu-Asp-Asn-Gln-Val-Pro-Tyr. This sequence was 71% homologous with the N-terminal of bovine trypsin. The Boc-Phe-Ser-Arg-MCA hydrolyzing activity of mouse spleen significantly (p < 0.01) increased to about 1.5-fold the basal activity 2 weeks after an injection of Freund's complete adjuvant, suggesting that these proteases are involved in the immune response. [ABSTRACT FROM AUTHOR]

Tumor-promoting phorbol esters activated a calcium-activated, phospholipid-dependent protein kinase by direct complexation with this protein kinase and phospholipid in the presence of divalent cations and this complexation was identical to association of kinase/receptor activity with cell membranes. Treatment of isolated mouse spleen lymphocytes with phorbol ester tumor promoters resulted in a rapid shift in the subcellular localization of both the phorbol ester receptor and a calcium-activated, phospholipid-dependent protein kinase activity. Both activities shifted from almost entirely soluble to largely membrane-associated, which is consistent with a single protein possessing both activities. Activation of partially purified kinase/receptor activity by phorbol ester or calcium alone or in combination occurred in parallel to the formation of a complex between the kinase/receptor and phospholipid. Magnesium also was important both for complex formation and for activation of the protein kinase. Although phorbol ester did not appear to affect the affinity of the kinase/receptor for phospholipid, it did increase the extent of formation of a stable complex between the receptor and the phospholipid. These observations support the hypothesis that the cell membrane is the locus of action of both the phorbol esters and the calcium-activated, phospholipid-dependent protein kinase activity. [ABSTRACT FROM PUBLISHER]

The CD8-expressing dendritic cells (DC) present in mouse spleen have been shown to have a regulatory effect on the CD4 and CD8 T cells they activate, restricting subsequent T cell proliferation by either inducing apoptotic T cell death (CD4 T cells) or by limiting endogenous cytokine production (CD8 T cells). To determine the role of the CD8 molecule itself in these regulatory phenomena, the DC from CD8 null mice were studied. The DC marker DEC-205 (NLDC 145) was used as a surrogate marker for CD8, since the expression of these two molecules on splenic DC was closely correlated. DC levels were normal, and the incidence of DEC-205+ and DEC-205- DC was normal in CD8 null mice, indicating that the absence of CD8 did not affect DC development. The proliferative response of T cells to allogeneic DEC-205+ DC from either CD8–/– or CD8+/+ mice was similar and was much less than the response to DEC-205- DC from these mice. This applied to both the CD4 and the CD8 T cell responses. Thus the lack of the CD8 molecule did not affect the stimulatory or regulatory properties of the DC. The regulatory CD8+ DEC-205+ DC therefore differ in that respect from antigen-presenting ‘veto’ cells, where CD8 itself is involved in transmitting negative signals to the T cells. DEC-205 may prove to be a more pertinent marker of the regulatory DC population. [ABSTRACT FROM AUTHOR]

Dengue type 2 virus (DV) infection induces production of a cytokine, the cytotoxic factor (CF) in the spleen of mice. The present study was undertaken to investigate the production of nitrite (NO2-) by the spleen cells of mice in vitro and in vivo following inoculation of DV or CF. Maximum NO2- production occurred at 45 min after inoculation of 5 µg CF, both in vitro and in vivo. The NO2- was produced by macrophages and T cells and not by B cells. Pretreatment of CF with anti- CF antisera inhibited production of NO2-. DV-stimulated spleen cell culture supernatants showed peak production of CF and NO2- at 72 h. In DV-infected mouse spleen, maximum NO2- production occurred at 8-11 days post-infection, which correlated with peak cytotoxic activity in the spleen. Pretreatment of spleen cells with NG-monomethyl L-arginine (NMMA) inhibited NO2- production. NO2- production was abrogated in a dose-dependent manner by treatment of spleen cells with Ca2+ channel blocking drug. Nifedipine. The findings demonstrate that DV induced CF induces production of NO2- in spleen cells, probably in a Ca2+-dependent manner, and may be a mechanism of target cell killing. [ABSTRACT FROM AUTHOR]

A mouse spleen cell/plasmacytoma fusion designed to generate hybridomas making monoclonal antibodies to human lymphocyte 5'-nucleotidase yielded three hybridomas secreting material which inhibited about 50% of human and mouse lymphocyte ecto 5'-nucIeotidase activity. The inhibition proved not to be due to antibody but to material of molecular weight 44.000 ± 7,000 which was not part of an immunoglobulin molecule although it may be related lo a B cell Fc γ receptor. In a haemolytic plaque assay this material inhibited the production of IgG but not IgM antibody by spleen cells of mice immunized with dinitrophenylated keyhole limpit haemocyanin. By contrast, IgG production by pokeweed mitogen-stimulated human tonsillar lymphocytes (assessed by reverse haemolytic plaque assay) was partially inhibited only by ascitic fluid of one of the hybridomas. The factor was called BAN (B cell anti 5'-nucleotidase). The suppressive action of BAN on IgG production was blocked by antibodies against 5-nucleotidase suggesting that the lymphocyte enzyme may be acting as a BAN receptor. [ABSTRACT FROM AUTHOR]

The limited infiltration and persistence of chimeric antigen receptor (CAR)-T cells is primarily responsible for their treatment deficits in solid tumors. Here, we present a three-dimensional scaffold, inspired by the physiological process of T-cell proliferation in lymph nodes. This scaffold gathers the function of loading, delivery, activation and expansion for CAR-T cells to enhance their therapeutic effects on solid tumors. This porous device is made from poly(lactic-co-glycolic acid) by a microfluidic technique with the modification of T-cell stimulatory signals, including anti-CD3, anti-CD28 antibodies, as well as cytokines. This scaffold fosters a 50-fold CAR-T cell expansion in vitro and a 15-fold cell expansion in vivo. Particularly, it maintains long-lasting expansion of CAR-T cells for up to 30 days in a cervical tumor model and significantly inhibits the tumor growth. This biomimetic delivery strategy provides a versatile platform of cell delivery and activation for CAR-T cells in treating solid tumors. [ABSTRACT FROM AUTHOR]

Motivation The rapid development of spatial transcriptome technologies has enabled researchers to acquire single-cell-level spatial data at an affordable price. However, computational analysis tools, such as annotation tools, tailored for these data are still lacking. Recently, many computational frameworks have emerged to integrate single-cell RNA sequencing (scRNA-seq) and spatial transcriptomics datasets. While some frameworks can utilize well-annotated scRNA-seq data to annotate spatial expression patterns, they overlook critical aspects. First, existing tools do not explicitly consider cell type mapping when aligning the two modalities. Second, current frameworks lack the capability to detect novel cells, which remains a key interest for biologists. Results To address these problems, we propose an annotation method for spatial transcriptome data called SPANN. The main tasks of SPANN are to transfer cell-type labels from well-annotated scRNA-seq data to newly generated single-cell resolution spatial transcriptome data and discover novel cells from spatial data. The major innovations of SPANN come from two aspects: SPANN automatically detects novel cells from unseen cell types while maintaining high annotation accuracy over known cell types. SPANN finds a mapping between spatial transcriptome samples and RNA data prototypes and thus conducts cell-type-level alignment. Comprehensive experiments using datasets from various spatial platforms demonstrate SPANN's capabilities in annotating known cell types and discovering novel cell states within complex tissue contexts. Availability The source code of SPANN can be accessed at https://github.com/ddb-qiwang/SPANN-torch. Contact dengmh@math.pku.edu.cn. [ABSTRACT FROM AUTHOR]

The advent of single-cell sequencing technologies has revolutionized cell biology studies. However, integrative analyses of diverse single-cell data face serious challenges, including technological noise, sample heterogeneity, and different modalities and species. To address these problems, we propose scCorrector, a variational autoencoder-based model that can integrate single-cell data from different studies and map them into a common space. Specifically, we designed a Study Specific Adaptive Normalization for each study in decoder to implement these features. scCorrector substantially achieves competitive and robust performance compared with state-of-the-art methods and brings novel insights under various circumstances (e.g. various batches, multi-omics, cross-species, and development stages). In addition, the integration of single-cell data and spatial data makes it possible to transfer information between different studies, which greatly expand the narrow range of genes covered by MERFISH technology. In summary, scCorrector can efficiently integrate multi-study single-cell datasets, thereby providing broad opportunities to tackle challenges emerging from noisy resources. [ABSTRACT FROM AUTHOR]

The influence of ageing on the in vitro responses of mouse spleen cells to phytohaemagglutinin and to allogeneic cells was studied in NZB, (NZB × A)F1, A and CBA mice. These responses decline with ageing in those strains which are genetically susceptible to autoimmune disease (NZB, (NZB × A)F1 and A) but not in a non-susceptible strain (CBA). These studies support the view that cellular immunity declines with advancing age in autoimmune-susceptible mice, and suggests that the decline is due to a progressive loss of the thymus-dependent cell pool. [ABSTRACT FROM AUTHOR]