Immunochemical Localisation of Cathepsin S, Cathepsin L and MHC Class II-Associated p41 Isoform of Invariant Chain in Human Lymph Node Tissue

Zavanik-Bergant, V.; Sekirnik, Andreja; Golouh, Rastko; Türk, Vito; Kos, Janko

doi:10.1515/bc.2001.096

Cited by 13 publications

(13 citation statements)

References 24 publications

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“…Tissue sections (5 m) from formalin-fixed, paraffin-embedded human lymph nodes were prepared [22] and labeled for CyC, MHC II (HLA-DR), CD68, and FDC. Labeling of lymph node tissue and confocal immunofluorescence microscopy was performed as described previously [21,23].…”

Section: Immunohistochemical Analysis Of Lymph Nodesmentioning

confidence: 88%

“…Cells were fixed with 4% paraformaldehyde in PBS (pH 7.2) for 1 h and permeabilized with 0.1% Triton X-100 for an additional 5 min. Cells were labeled with antibody and visualized by immunofluorescence microscopy using a confocal laser-scanning microscope, Carl Zeiss LSM 510, as described [21], unless stated otherwise. Anti-CyC 1A2 mAb was labeled with Alexa Fluor TM 488 fluorophore, using Zenon TM One Mouse IgG 1 labeling kit, as recommended by the producer (Molecular Probes) prior to colocalization study of CyC with golgin-97 [11] in immature and mature DC.…”

Section: Confocal Immunofluorescence Microscopymentioning

confidence: 99%

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Differentiation- and maturation-dependent content, localization, and secretion of cystatin C in human dendritic cells

Zavašnik–Bergant

Repnik

Schweiger

et al. 2005

Journal of Leukocyte Biology

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Antigen-presenting cells (APC) play a pivotal role in the initiation of the T cell-mediated and antigen-specific immune response. The suggested role of endogenous inhibitor cystatin C (CyC) is to modulate cysteine proteases (cathepsins) present in human APC. To test this hypothesis, dendritic cells (DC) were generated in vitro from isolated monocytes, and changes in content, localization, and secretion of CyC and cathepsins S, L, and H (CatS, -L, and -H, repsectively) were followed in response to interleukin-4, enabling monocyte differentiation, and to tumor necrosis factor alpha (TNF-alpha), enabling DC maturation. A large increase in intracellular CyC accompanied the differentiation of monocytes to immature DC, also shown by strong immunolabeling of Golgi in immature DC. On DC maturation, intracellular CyC levels decreased, and CyC was mostly absent from the Golgi. On prolonged incubation of mature DC with TNF-alpha, CyC was found located in the proximity of the plasma membrane, indicating that the transport of CyC from Golgi was not blocked as the result of the arrested exocytosis in mature DC. The secretion of CyC ceased, consistent with the peak of the surface expression of phenotypic markers (CD40, CD54, CD80, CD83, CD86, and major histocompatibility complex class II), characteristic for the mature DC stage, whereas the secretion of cathepsins did not correlate with the maturation stage. The difference in localization of CyC and of CatS, -L, and -H in immature and mature DC shows that the regulatory potential of CyC toward CatS, -L, and -H inside DC is limited. However, these interactions may occur extracellularly in lymph, as suggested by the large excess of CyC over secreted CatS, -L, and -H, and they may facilitate DC migration to lymph nodes.

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Section: Immunohistochemical Analysis Of Lymph Nodesmentioning

confidence: 88%

Section: Confocal Immunofluorescence Microscopymentioning

confidence: 99%

Differentiation- and maturation-dependent content, localization, and secretion of cystatin C in human dendritic cells

Zavašnik–Bergant

Repnik

Schweiger

et al. 2005

Journal of Leukocyte Biology

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“…Monoclonal antibodies (MAbs) (Krka, d.d., Ljubljana, Slovenia) used in ELISA, IHA and Western blots were prepared from mouse hybridoma cell lines (Schweiger et al, 1997;Zavašnik-Bergant et al, 2001). The cell culture supernatants were purified by affinity chromatography on protein A-Sepharose as recommended by the producer (Pharmacia, Uppsala, Sweden).…”

mentioning

confidence: 99%

Cathepsin S in tumours, regional lymph nodes and sera of patients with lung cancer: relation to prognosis

et al. 2001

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Cysteine proteinase cathepsin S (Cat S) is expressed mainly in lymphatic tissues and has been characterised as a key enzyme in major histocompatibility complex class II (MHC-II) mediated antigen presentation. Cat S has been measured in tissue cytosols of lung parenchyma, lung tumours and lymph nodes and in sera of patients with lung tumours and of healthy controls, by specific enzyme-linked immunosorbent assay (ELISA). A difference in Cat S level was found between tumour and adjacent control tissue cytosols of 60 lung cancer patients (median 4.3 vs. 2.8 ng mg−1protein). In lymph nodes obtained from 24 patients of the same group, the level of Cat S was significantly higher than in tumours or lung parenchyma (P< 0.001). Additionally, significantly higher levels were found in non-infiltrated than in infiltrated lymph nodes (median 16.6 vs 7.5 ng mg−1protein). Patients with low levels of Cat S in tumours and lung parenchyma exhibited a significantly higher risk of death than those with high levels of Cat S (P= 0.025 – tumours;P= 0.02 – parenchyma). Immunohistochemical analysis (IHA) of lung parenchyma revealed a staining reaction in alveolar type II cells, macrophages and bronchial epithelial cells. In regional lymph node tissue, strong staining of Cat S was found in lymphocytes and histiocytes. Nevertheless, Cat S was detected also in tumour cells, independently of their origin. Our results provide evidence that Cat S may be involved in malignant progression. Its role, however, differs from that of the related Cats B and L and could be associated with the immune response rather than with remodelling of extracellular matrix. © 2001 Cancer Research Campaign http://www.bjcancer.com

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“…We have previously reported strong colocalization of p41 Ii with cathepsin S in CD68 + sinus-lining macrophages. 19 This is to be expected, since cathepsin S is a key enzyme capable to degrade Ii, releasing MHC II binding site. 15 Therefore, in sinus-lining macrophages p41 Ii represents cathepsin's S substrate.…”

Section: Discussionmentioning

confidence: 96%

“…Monoclonality of prepared mouse anti-Ii antibodies (2C12 mAb and 5F6 mAb) was confirmed by isoelectric focusing as described. 19 Specific antibodies against human cathepsins L, H and S, used in this study, were also prepared and characterized in our laboratory as previously reported: mouse anti-cathepsin S 1E3 mAb 20 mouse anti-cathepsin H 1D10 mAb 18 and sheep anti-cathepsin L polyclonal antibody (pAb). 21 Characterization of anti-invariant chain monoclonal antibodies by ELISA The specificity and cross-reactivity of anti-Ii (2C12 mAb and 5F6 mAb) were compared by antigen-immobilized ELISA using standard protocol at our department.…”

Section: Production and Selection Of Anti-invariant Chain Monoclonal mentioning

confidence: 99%

Inhibitory p41 isoform of invariant chain and its potential target enzymes cathepsins L and H in distinct populations of macrophages in human lymph nodes

et al. 2004

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SUMMARYActivation of the CD4 + T-cell mediated immune response relies on the proteolytic capacity of enzymes involved in modulating major histocompatibility complex (MHC) II-associated antigen presentation in antigen-presenting cells (APC). The MHC II-associated chaperone molecule p41 isoform of invariant chain (inhibitory p41 Ii) has been suggested to regulate stability and activity of cathepsin L in these APC. In the present study the human lymph node distribution of noninhibitory p31 Ii and inhibitory p41 Ii have been compared by differential labelling, using two specific monoclonal antibodies. The distribution of p41 Ii, but not p31 Ii, matched the distribution of cathepsins L and H in subcapsular and cortical sinuses and germinal centres. Co-localization of p41 Ii with cathepsin H was confirmed in strongly CD68 + sinus-lining macrophages, acting as APC. Furthermore, p41 Ii was determined together with cathepsins L and H in tingible body macrophages, highly phagocytic, but not antigen-presenting cells inside germinal centres. With respect to the physiological function that these two populations of macrophages have in human lymph nodes, our results support a regulatory function of p41 Ii towards cathepsins L and H in human macrophages, associated with the processes of phagocytosis rather than antigen presentation.

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Immunochemical Localisation of Cathepsin S, Cathepsin L and MHC Class II-Associated p41 Isoform of Invariant Chain in Human Lymph Node Tissue

Cited by 13 publications

References 24 publications

Differentiation- and maturation-dependent content, localization, and secretion of cystatin C in human dendritic cells

Differentiation- and maturation-dependent content, localization, and secretion of cystatin C in human dendritic cells

Cathepsin S in tumours, regional lymph nodes and sera of patients with lung cancer: relation to prognosis

Inhibitory p41 isoform of invariant chain and its potential target enzymes cathepsins L and H in distinct populations of macrophages in human lymph nodes

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