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Sponge matrix allografts. A model for analysis of killer cells infiltrating mouse allografts.

A method for isolation of allograft-infiltrating cells in a functionally viable state is described in this article. The method is based on the use of a spongious tissue into which cells of strain A (e.g., fibroblasts or tumor cells) are grown. The resulting graft is then transplanted to a strain B animal, and the infiltrating cells are released from it by gentle compression. The graft-infiltrating cells are completely recovered, and they may be processed for further experimentation and analysis by employing exclusively physical methods of cell preparation. As an example of future applications, some preliminary results on density- and charge fractionation of the graft-infiltrating cells are also reported.

Animals

Identification of mononuclear cells infiltrating basal cell carcinomas.

The identification of mononuclear cells extracted from 12 basal cell carcinomas has been carried out by means of various assays such as the E rosette test, FITC anti-Ig conjugates and peroxidase conjugates viewed at an ultrastructural level. The relative quantities of B and T cells have been determined as well as the morphology of the extracted cells. It was found that the T/B ratio is close to that of a delayed-type reaction to tuberculin, suggesting a possible identical immune mechanism, and also that the morphological appearance of some tissue lymphocytes differs from that of circulating lymphocytes due to the presence of both membrane and cytoplasmic Ig.

B-Lymphocytes

Role of mononuclear infiltrating cells in pathogenesis of hepatitis.

Mice that have been injected with Corynebacterium parvum have mononuclear-cell infiltrates in the liver lobules. In such mice a small dose of lipopolysaccharide endotoxin produced a lethal hepatitis, with high serum-transaminase concentrations, glycogen depletion, and hypoglycaemia. It is suggested that lipopolysaccharide triggers the release from the infiltrating mononuclear cells of factors toxic for hepatocytes. Similarly certain parasitic and virus infections and graft-versus-host reactions can sensitise mice to the induction of hepatitis by exposure to small doses of lipopolysaccharide. This model may be applicable to human hepatitis.

Animals

Ultrastructure of cells infiltrating human kidney allografts.

Cell infiltration is commonly observed in human renal allograft biopsies. This infiltration was investigated using electron microscopy for a more precise assessment of the nature of these cells. More than 3000 cells infiltrating twenty-five renal allograft biopsies were studied. Six cellular types were distinguished and a mean percentage of each type was calculated. Only one-half of these cells were normal or transformed lymphocytes (including small lymphocytes: 22-3 +/- 3-8%, 'intermediary' cells: 22 +/- 3-6%, blast-like cells similar to MLC transformed lymphocytes: 8-1 +/- 2-4%. A relatively high number of plasmocytes (12-4 +/- 2-5%) and a still higher percentage of macrophages (28-5 +/- 4-6%) were found. Granulocytes represented only 2 +/- 0-8%of the cell population. Variations of the mean percentage of these cellular types were studied in various clinical situations.

Cell Movement

Tissue T and B cell infiltration of primary and metastatic cancer.

Immunofluorescent techniques were utilized to identify the types of infiltrating lymphocytes adjacent to human malignant tumors arising from a wide range of anatomic sites. 24 of 29 primary tumors and 5 of 8 metastatic lesions showed varying degrees of lymphocytic infiltration. T cells predominated in the infiltrates in primary tumors (mean 80%, range 50-100%) and this pattern was evident regardless of anatomic site or the presence or absence of metastatic spread. By contrast, B cells predominated at the margins of three of five tumor metastases. Mononuclear cells bearing the Fc receptor were not a prominent component of the infiltrates associated with either primary tumors or metastases, but tumor cell binding of fluoresceinated IgG aggregates was observed in 12 of 29 primary tumors. A significant reduction in peripheral blood T cell numbers occurred in a third of the patients studied. This decrease was not clearly related either to the extent of local tumor T cell infiltration or to the presence of disseminated disease. These preliminary findings provide a descriptive analysis of the local and systemic distributions of immunocompetent cells in cancer.

Adult

[Study of macrophages infiltrating rat cardiac allografts. II.--Cytoenzymatic analysis of cells infiltrating the grafts and of peritoneal macrophages in isograft and allograft recipients (author's transl)].

Cytoenzymatic analysis of cells infiltrating heart transplants in the rat confirms the high participation of monocytes and macrophages. However, when comparing iso and allografts, only slight differences are observed in the lysosomal enzyme specific colorations, whereas a striking difference in these colorations is found between heart infiltrates and peritoneal exsudates from these recipients. This could favour the hypothesis of a local macrophage activating process within the graft but seems more probably due to differences in cell maturation stages.

Acid Phosphatase

Cellular components of allograft rejection: identity, specificity, and cytotoxic function of cells infiltrating acutely rejecting allografts.

Functioning mononuclear cells have been harvested from heterotopic rat cardiac allografts during maximal transplant cellular infiltration. T cells, identified by a T cell-specific absorbed rabbit anti-rat brain serum, constituted two-thirds of the total cells recovered. Approximately 20% of the infiltrating cells bear and synthesize surface immunoglobulin. Macrophages, identified by latex ingestion and morphologic and cytochemical techniques, comprise 9% of the graft infiltrate. Donor-specific cytotoxic T lymphocytes are concentrated within the graft. A separate population of Fc receptor-positive recovered cells mediate antibody-dependent LMC (Ab-LMC). Neither effector cell was adherent or phagocytic. These studies have conclusively established that cytotoxic T lymphocytes accumulate within rejecting allografts; however, the enriched presence of cytotoxic T cells within the grafts is not fully dependent upon antigen recognition per se, since Lew animals grafted with both BN and BUF hearts have Lew anti-BN and Lew anti-BUF killer cells in each graft.

Animals

Rejected human renal allografts: recovery and characteristics of infiltrating cells and antibody.

Viable infiltrating host leukocytes have been isolated from 10 rejected human renal allografts, removed 1 to 67 months after transplantation. The cell populations have been identified by surface characteristics and their cytotoxic capacities were assessed. A heterogenous population of cells of host origin accumulated in the grafts, including T and B lymphocytes, Fc+ cells, and macrophages. Using a 51Cr release assay, specific cytotoxicity against donor alloantigens was determined. Cytotoxicity of the infiltrating cells was almost invariably greater than cytotoxicity mounted by recipient peripheral blood lymphocytes. Deletion studies confirmed previous work and suggested that T cells were primarily responsible for cytolysis in early acute rejection; non-T cells more often in late chronic rejection. Antibodies eluted from the grafts demonstrated both specific antidonor and nonspecific activity as well as cross-reacting anti-HLA activity. Allograft morphology was examined and cellular and humoral host responses were assessed. These studies emphasize the complexities of immune responses produced by the host against transplanted tissues.

Antibody Specificity

A comparison of the hematogenous cell infiltrate evoked by lymphokine injection with that of delayed hypersensitivity reactions.

Lymphokine preparations of high potency obtained by in vitro assay were employed in vivo to determine whether they could produce inflammatory responses showing a cellular infiltrate that qualitatively or quantitatively resembled responses of delayed hypersensitivity. Hematogenous cell infiltrates in guinea pig skin were characterized in terms of the number and types of participating cells following intradermal injection of either lymphokines or an antigen (PPD) to which the animals exhibited delayed hypersensitivity. The dose of lymphokine or PPD per skin test site was selected on the basis of comparable ability to enhance vascular permeability. Delayed hypersensitivity responses showed, as expected, a persistent mononuclear cell exudate both in dermis and subcutis, but most notable in the dermis, during the 24 hours following antigen injection. In contrast, the response to lymphokine over the same period was characteristically neutrophilic and principally in the subcutis. There was no pronounced mononuclear cell infiltrate at any time throughout the reaction to lymphokine. It is concluded that preformed lymphokine produces a pattern of increased vascular permeability appropriate to a mediator of delayed hypersensitivity reactions, provided there is sustained secretion of this material. The ability of lymphokine to cause carbon labeling of dermal capillaries is also pertinent to a mediator of delayed hypersensitivity. The absence of significant mononuclear cell accumulation suggests that the in vitro chemotactic activity of lymphokine toward mononuclear cells may be more important for retention of mononuclear cells in the extravascular connective tissue space than for their selective accumulation.

Animals

Characterization of mononuclear cell infiltrates in psoriatic lesions.

The dermal mononuclear cell infiltrates of psoriatic lesions were characterized by receptors for sheep erythrocytes (T-lymphocytes), C3b receptors (macrophages and B-lymphocytes) and C3d receptors (B-lymphocytes), using hemadsorption to cryostat sections in a closed chamber. T-lymphocytes and macrophages were the predominant inflammatory cells. Very few B-lymphocytes were detected. These findings are discussed in relation to the possible pathogenetic signficance of cellular immunity in psoriasis.

B-Lymphocytes

Elevated Triggering Receptor Expressed on Myeloid Cells 2 Expression in Tumor-Associated Macrophages Suppresses Cytotoxic T Cell Infiltration and Facilitates Immune Escape in Colorectal Cancer.

BACKGROUND & AIMS: Emerging evidence supports a crucial role for tumor-associated macrophages in shaping the immunosuppressive tumor microenvironment. Furthermore, research has identified that the triggering receptor expressed on myeloid cells 2 has immunomodulatory functions. The present investigated the potential effect of triggering receptor expressed on myeloid cells 2 expression in tumor-associated macrophages on facilitating immune evasion in colorectal cancer. METHODS: Immunohistochemical analysis of clinical specimens, complemented by extensive data mining from The Cancer Genome Atlas, revealed a significant upregulation of triggering receptor expressed on myeloid cells 2 in colorectal cancer-associated tumor-associated macrophages, with this upregulation exhibiting a correlation with poor patient prognosis. RESULTS: Mechanistically, triggering receptor expressed on myeloid cells 2+ tumor-associated macrophages were found to drive fibroblast activation through transforming growth factor-β signaling, inducing fibroblast-activated protein-positive cancer-associated fibroblasts that secrete collagen I/III to establish dense peritumoral barriers. Spatial profiling revealed that these fibrous structures physically impede CD8+ T-cell infiltration, restricting cytotoxic lymphocytes to stromal compartments. Intriguingly, triggering receptor expressed on myeloid cells 2 deficiency enhanced the secretion of matrix metalloproteinase 13 by macrophages, thereby promoting extracellular matrix degradation and improving T-cell penetration. In vivo, Trem2-knockout mice showed a reduction in tumor growth with enhanced intratumoral CD8+ T-cell infiltration compared with wild-type controls. CONCLUSIONS: Our findings establish triggering receptor expressed on myeloid cells 2+ tumor-associated macrophages as central regulators of stromal remodeling and suggest that therapeutic targeting of the triggering receptor expressed on myeloid cells 2/transforming growth factor-β/fibroblast-activated protein pathway may overcome immune resistance in patients with colorectal cancer.

Colorectal Neoplasms

Effect of X-irradiation on host-cell infiltration and growth of a murine fibrosarcoma.

Whole body X-irradiation (400 rad) of C57BL mice, either before or after i.m. implantation of the syngeneic fibrosarcoma, FS6, influenced both the growth of the tumours and their cellular composition, particularly their macrophage content. Pre-irradiation resulted in slower initial growth of tumours, and a concomitant lack of host-cell infiltration, but when tumours began to grow at a rate parallel to controls infiltration by host cells was demonstrable. Similarly, irradiation of the tumour-bearing host resulted in a temporary cessation of growth, and a decrease in the macrophage content, which did not return to control levels for 2-3 weeks after irradiation. The significance of these results is discussed in relation to the possibility that infiltrating host cells, particularly macrophages, may stimulate the growth of this tumour.

Animals

[Study of macrophages infiltrating rat cardiac allografts. 1.--Description of the technique of heterotopic cardiac transplantation in the rat and morphological analysis of the cells infiltrating the graft (author's transl)].

The technique of heterotopic cardiac transplantation in the rat is described in detail. A simple, nontraumatic method of obtaining cellular prints from the heart is proposed for the study of cells infiltrating the allograft. Analysis of the morphological aspect of the cellular infiltrate present at the time of rejection in a series of allografts revealed a very high proportion of cells belonging to the monocyte-macrophage series, thus suggesting an important role for these cells in the rejection process.

Animals

Effect of Tertiary Lymphoid Structures on Immune Cell Infiltration in the Tumor Microenvironment and Prognosis in Lung Adenocarcinoma.

Tertiary lymphoid structures (TLSs) modulate immune responses in various solid tumors, but their comprehensive role in lung adenocarcinoma (LUAD) remains unclear. In this study, we analyzed RNA-seq data from 539 LUAD patients in The Cancer Genome Atlas (TCGA) and microarray data from 223 samples from the Gene Expression Omnibus (GEO, GSE13213, and GSE37745). TLS signatures were evaluated via unsupervised consensus clustering based on 12 chemokine transcriptome signatures. The relationships between TLS and clinical characteristics, tumor microenvironment (TME) cell infiltration, and prognosis were assessed using ESTIMATE and CIBERSORT. A prognostic model was established using LASSO regression and validated with external datasets. Additionally, H&E and IHC analyses were performed to explore associations between intratumoral TLS density, immune-related molecular expression, and patient prognosis in LUAD. Consensus clustering of the TCGA cohort revealed two distinct LUAD patient clusters according to TLS abundance. Cluster 1 exhibited greater immune cell infiltration, more favorable prognosis, and increased expression of immune checkpoint molecules. We developed a prognostic model comprising eight survival-associated genes that act as independent prognostic factors for patient survival. H&E/IHC analyses revealed that TLS density-regardless of pathological stage-was associated with better prognosis; higher intratumoral TLS density/proportion was also related to more favorable outcomes. IHC confirmed that survival-associated genes (CD5, HLA-DMB, and P2RY13) are independent prognostic indicators in LUAD. Our study demonstrated the close relationship between TLS signatures and an active immune microenvironment, highlighting their potential as independent prognostic indicators in LUAD.

Humans

Partial characterization of cytotoxic cells infiltrating sponge matrix allografts.

Adapting the Roberts and Hayry sponge allograft model, we have demonstrated the presence of an enriched, specifically cytotoxic population of cells which infiltrate rejecting sponge allografts. The number of cells infiltrating a rejecting sponge allograft peaks on day 14 after transplantation. Utilizing a short-term 51chromium cytotoxicity assay, peak antiallogeneic killing was demonstrable on day 14 also. Only T cell killing was apparent for the first 15 days after transplantation. After day 20, specific cytolysis was present which was not sensitive to anti-theta serum and complement. The infiltrating cytotoxic cells are large, specifically cytotoxic, do not proliferate in culture, do not respond to mitogen, and do not respond in mixed lymphocyte culture even to the same alloantigen to which the animal had been sensitized. In contrast, spleens from sponge-bearing animals kill poorly, respond to mitogen, and respond vigorously in mixed lymphocyte culture to specific and nonspecific alloantigens. The following hypotheses are set forth with regard to the cytotoxic lymphocytes (1) Such cells may be end stage and cannot proliferate. (2) The cytotoxic cells may kill the stimulator cells more rapidly than they can be stimulated to proliferate. (3) The sponge cell population may be enriched for nonspecific supressor cells. (4) The sponge cells may be devoid of helper T cells.

Animals

[Enzyme cytochemical and immunocytological differentiation of infiltrative cells in the skin in mycosis fungoides].

In 4 patients with mycosis fungoides in the praemycotic, infiltrative and tumour stage of the disease, a differentiation and functional characterization of the infiltrate cells, obtained from cell suspensions and cutaneous smears, was carried out by means of enzyme cytochemical and immunological methods. The lymphocyte-associated acid esterase is considered to be a marker for mature T-cell populations. Apart from monocytes and histiocytes, acid esterase-positive small lymphocytoid cells with T-cell properties are found in the praenycotic stage. In the tumour stage, large lymphocytoid cells become increasingly prevalent, they show no acid esterase activity, but an intracytoplasmatic-localized reaction in the acid phosphatase activity. On the basis of the cytochemical pattern, it is assumed that these cells represent proliferating lymphoblasts.

Acid Phosphatase

Inflammatory seborrheic keratoses with mononuclear cell infiltration.

Inflammation in inflammatory seborrheic keratoses was found to be composed of mononuclear cells, particularly lymphocytes. Accumulations of mononuclear cells in the papillary and subpapillary dermis frequently showed no tendency to infiltrate the epithelium. However, dramatic interaction of the seborrheic keratosis and the inflammatory process also was common. The lymphocytic exocytosis into the epithelium followed two basic patterns: eczematous, characterized by intercellular and intracellular edema, microvesicle formation, and epithelial necrosis; and lichenoid, in which a bandlike infiltrate attacked the dermoepidermal junction and hyaline bodies occurred in the infiltrated epithelium and in the dermis. Epithelial changes consisted of necrosis of epithelial cells and related parakeratosis; squamous eddies were occasionally found. Inflammatory cell infiltration in seborrheic keratoses possibly represents a process other than irritation, and we propose that mononuclear cell infiltration in seborrheic keratoses may be related to an involutionary process.

Dermatitis, Seborrheic

Identification of mitophagy-related biomarkers with immune cell infiltration in psoriasis.

BACKGROUND: Psoriasis is an inflammatory disorder characterized by scaly erythematous plaques and significant comorbidities. Recent studies have suggested that impaired mitophagy, the cellular mechanism for removing dysfunctional mitochondria, may contribute to the pathogenesis of psoriasis. METHODS: In this study, we analyzed bulk RNA sequencing data from 167 healthy individuals and 177 patients with psoriasis obtained from the Gene Expression Omnibus database (GSE30999 and GSE54456). Mitophagy-related genes were isolated using weighted gene co-expression network analysis. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed and protein-protein interaction networks were constructed for the functional enrichment of genes associated with mitophagy. The correlations between genes associated with mitophagy, signaling pathways, and immune cell infiltration were analyzed. The potential diagnostic value of genes associated with mitophagy was evaluated using receiver operating characteristic (ROC) curves, which were validated in imiquimod-induced psoriatic skin lesions in mice. RESULTS: We identified 3,839 differentially expressed genes between healthy individuals and patients with psoriasis, and 23 genes were selected as hub genes showing a high correlation with mitophagy in psoriasis. GO and KEGG analyses revealed that hub and associated genes were significantly correlated with skin functions, such as epidermal development and keratinocyte differentiation. In addition, mitophagy-related genes were negatively associated with pro-inflammatory and pro-proliferation pathways in psoriasis. Among the immune cells, CD4+ T cells were most significantly affected by mitophagy-related genes. ROC analysis demonstrated that mitophagy-related genes, especially ACER1, C1ORF68, CST6, FLG2, GJB3, GJB5, GPRIN2, KRT2, and SPRR4 were potential biomarkers of psoriasis for use in diagnosis or treatment. CONCLUSIONS: Mitophagy-related genes play crucial roles in psoriasis and have potential use as biomarkers, providing insights into disease mechanisms and therapeutic targets. Further research may lead to the development of new strategies for psoriasis management.

Psoriasis