Medulla Of Lymph Nodes Research Articles

Neural-Cadherin Influences the Homing of Terminally Differentiated Memory CD8 T Cells to the Lymph Nodes and Bone Marrow.

Memory T (TM) cells play an important role in the long-term defense against pathogen reinvasion. However, it is still unclear how these cells receive the crucial signals necessary for their longevity and homeostatic turnover. To understand how TM cells receive these signals, we infected mice with lymphocytic choriomeningitis virus (LCMV) and examined the expression sites of neural cadherin (N-cadherin) by immunofluorescence microscopy. We found that N-cadherin was expressed in the surroundings of the white pulps of the spleen and medulla of lymph nodes (LNs). Moreover, TM cells expressing high levels of killer cell lectin-like receptor G1 (KLRG1), a ligand of N-cadherin, were co-localized with N-cadherin+ cells in the spleen but not in LNs. We then blocked N-cadherin in vivo to investigate whether it regulates the formation or function of TM cells. The numbers of CD127hiCD62Lhi TM cells in the spleen of memory P14 chimeric mice declined when N-cadherin was blocked during the contraction phase, without functional impairment of these cells. In addition, when CD127loKLRG1hi TM cells were adoptively transferred into anti–N-cadherin–treated mice compared with control mice, the number of these cells was reduced in the bone marrow and LNs, without functional loss. Taken together, our results suggest that N-cadherin participates in the development of CD127hiCD62Lhi TM cells and homing of CD127loKLRG1hi TM cells to lymphoid organs.

Ultrasound Elastography for the Evaluation of Lymph Nodes.

The differential diagnosis of lymphadenopathy is important for predicting prognosis, staging, and monitoring the treatment, especially for cancer patients. Conventional computed tomography and magnetic resonance imaging characterize lymph node (LN) with disappointing sensitivity and specificity. Conventional ultrasound with the advantage of high resolution has been widely used for the LN evaluation. Ultrasound elastography (UE) using color map or shear wave velocity can non-invasively demonstrate the stiffness and homogeneity of both the cortex and medulla of LNs and can detect early circumscribed malignant infiltration. There is a need of a review to comprehensively discuss the current knowledge of the applications of various UE techniques in the evaluation of LNs. In this review, we discussed the principles of strain elastography and shear wave-based elastography, and their advantages and limitations in the evaluation of LNs. In addition, we comprehensively introduced the applications of various UE techniques in the differential diagnosis of reactive LNs, lymphoma, metastatic LNs, and other lymphadenopathy. Moreover, the applications of endoscopic UE and endobronchial UE are also discussed, including their use for improving the positive rate of diagnosis of fine-needle aspiration biopsy.

Application of conventional ultrasonography combined with contrast-enhanced ultrasonography in the axillary lymph nodes and evaluation of the efficacy of neoadjuvant chemotherapy in breast cancer patients.

Objective:Axillary lymph node status assessment has always been an important issue in clinical treatment of breast cancer. However, there has been no effective method to accurately predict the pathological complete response (pCR) of axillary lymph node after neoadjuvant chemotherapy (NAC). The objective of our study was to investigate whether conventional ultrasonography combined with contrast-enhanced ultrasonography (CEUS) can be used to evaluate axillary lymph node status of breast cancer patients after NAC.Methods:A total of 74 patients who underwent NAC were recruited for the present study. Prior to and after NAC, examinations of conventional ultrasonography and CEUS were performed. After evaluating the images of conventional ultrasonography, four characteristics were recorded: lymph node medulla boundary, cortex of lymph node, lymph node hilus, and lymph node aspect ratio. Two additional imaging characteristics of CEUS were analyzed: CEUS way and CEUS pattern. Receiver operating characteristiccurve analysis was applied to evaluate their diagnostic performance.Results:After 6~8 cycles of NAC, 46 (71.9%) patients had negative axillary lymph node, and 18 (28.1%) patients turned out non-pCR. According to statistical analysis, lymph node medulla, lymph node aspect ratio and CEUS way were independently associated with pCR of axillary lymph node after NAC. The area under the curve of the prediction model with three imaging characteristics was 0.882 (95% confidence interval: 0.608–0.958), and the accuracy to predict the patients’ lymph node status was 78.1% (p < 0.01).Conclusions:Conventional ultrasonography combined with CEUS technology can accurately predict axillary lymph nodes status of breast cancer patients after NAC.Advances in knowledge:The usefulness of CEUS technology in predicting pCR after neoadjuvant chemotherapy is highlighted.

Small Molecules for Multi-Wavelength Near-Infrared Fluorescent Mapping of Regional and Sentinel Lymph Nodes in Colorectal Cancer Staging.

Assessing lymph node (LN) status during tumor resection is fundamental for the staging of colorectal cancer. Current guidelines require a minimum of 12 LNs to be harvested during resection and ultra-staging regional lymph nodes by sentinel lymph node (SLN) assessment is being extensively investigated. The current study presents novel near-infrared (NIR) fluorescent dyes for simultaneous pan lymph node (PanLN; regional) and SLN mapping. PanLN-Forte was intravenously injected in mice and assessed for accumulation in regional LNs. SLN800 was injected intradermally in mice, after which the collection and retention of fluorescence in SLNs were measured using indocyanine green (ICG) and its precursor, SLN700, as references. LNs in the cervical, inguinal, jejunal, iliac, and thoracic basins could clearly be distinguished after a low dose intravenous injection of PanLN-Forte. Background fluorescence was significantly lower compared to the parent compound ZW800-3A (p < 0.001). SLN700 and SLN800 specifically targeted SLNs with fluorescence being retained over 40-fold longer than the current clinically used agent ICG. Using SLN700 and SLN800, absolute fluorescence in SLN was at least 10 times higher than ICG in second-tier nodes, even at 1 hour post-injection. Histologically, the fluorescent signal localized in the LN medulla (PanLN-Forte) or sinus entry (SLN700/SLN800). PanLN-Forte and SLN800 appear to be optimal for real-time NIR fluorescence imaging of regional and SLNs, respectively.

ECRG4 expression in normal rat tissues: expression study and literature review.

The Esophageal Cancer Related Gene 4 (ECRG4) is a highly conserved tumour suppressor gene encoding various peptides (augurin, CΔ16 augurin, ecilin, argilin, CΔ16 argilin) which can be processed and secreted. In the present work, we examined ECRG4 expression and location in a wide range of rat organs and reviewed the available literature. ECRG4 mRNA was identified in all examined tissues by quantitative PCR (qPCR). ECRG4 immunoreaction was mainly cytoplasmic, and was detected in heart and skeletal muscles, smooth muscle cells showing only weak reactions. In the digestive system, ECRG4 immunostaining was stronger in the esophageal epithelium, bases of gastric glands, hepatocytes and pancreatic acinar epithelium. In the lymphatic system, immunoreactive cells were detectable in the thymus cortex, lymph node medulla and splenic red pulp. In the central and peripheral nervous systems, different neuronal groups showed different reaction intensities. In the endocrine system, ECRG4 immunoreaction was detected in the hypothalamic paraventricular and supraoptic nuclei, hypophysis, thyroid and parathyroid glands, adrenal zona glomerularis and medulla and Leydig cells, as well as in follicular and luteal cells of the ovary. In the literature, ECRG4 has been reported to inhibit cell proliferation and increase apoptosis in various cell types. It is down-regulated, frequently due to hypermethylation, in esophageal, prostate, breast and colon cancers, together with glioma (oncosuppressor function), although it is up-regulated in papillary thyroid cancer (oncogenic role). ECRG4 expression is also higher in non-proliferating cells of the lymphatic system. In conclusion, our identification of ECRG4 in many structures suggests the involvement of ECRG4 in the tumorigenesis of other organs and also the need for further research. In addition, on the basis of the location of ECRG4 in neurons and endocrine cells and the fact that it can be secreted, its role as a neurotransmitter/neuromodulator and endocrine factor must be examined in depth in the future.

Highlights from the latest research in nanomedicine.

2015 A nanoparticulate hydrogel anticancer vaccine Evaluation of: Muraoka D, Harada N, Hayashi T et al. Nanogel-based immunologically stealth vaccine targets macrophages in the medulla of lymph node and induces potent antitumor immunity. ACS Nano 8(9), 9209–9218 (2014). Therapeutic cancer vaccines are designed to stimulate highly specific immune responses against cancers by targeting tumor-associated antigens. Various vaccines have utilized a wide range of immunogenic agents, including peptides, recombinant viruses, tumor cells and primed antigen-presenting dendritic cells, in combination with immunogenicityboosting adjuvants, immune suppression inhibitors or traditional chemotherapeutic agents [1]. However, cancer vaccines have been of varying effectiveness in clinical trials due to tumor characteristics such as antigenic heterogeneity and immunosuppressive mechanisms. Antigenic heterogeneity may allow the tumors to develop resistance against single-target immunogenic agents, while immunosuppressive mechanisms reduce the effectiveness of the immune response and alter the self-recognition mechanisms, causing cancerous cells to be ignored. Furthermore, tumor-associated antigen-targeting agents show varying immunogenic properties, with tradeoffs between specificity and effectiveness [1–3]. Nanoparticles have been of interest as efficacy-enhancing vaccine delivery agents due to their size and customizability. They are small enough to move into lymphatic systems and enter antigen-presenting cells through engulfment, and can be designed for specific parameters in terms of shape, surface properties and composition in order to enable antigen delivery specificity and presentation longevity [4,5]. Muraoka et al. developed hydrogel nanoparticles (nanogels) of approximately 50 nm in diameter using cholesteryl pullulan (CHP) polysaccharides, which self-assemble in water. These nanogels stably complex with polypeptides through hydrophobic interactions. Stability was shown for over 40 h in serum using nanogels complexed with a long-peptide antigen (LPA,) which either included the murine tumor-specific antigen mERK2 or the human tumor antigen MAGE-A4. Due to their small particle size, lack of charge and potential ligand sites, the CHP–LPA nanogel complexes travel efficiently in the lymphatic system to the draining lymph node, escaping uptake by lymph node dendritic cells until they reach the central medulla portion of the node. There, they are largely engulfed by medullary macrophages. Cellular uptake was evaluated through flow cytometry assessments of rhodamine-labeled CHP nanogels upon immune cell populations isolated from the draining lymph node. LPA that was complexed with other formulations did not show similar uptake patterns. Medullary macrophage antigen cross-presentation was assessed with CHP–mERK2 LPA, a Toll-like receptor 9 agonist and a CpG oligodeoxynucleotide adjuvant. The results showed successful T-cell recognition of mERK2-derived tumor epitopes mainly with the macrophage cells purified from the lymph nodes, with weak T-cell responses from purified dendritic cell populations. These outcomes were confirmed by impaired Highlights from the latest research in nanomedicine

Nanogel-based immunologically stealth vaccine targets macrophages in the medulla of lymph node and induces potent antitumor immunity.

Because existing therapeutic cancer vaccines provide only a limited clinical benefit, a different vaccination strategy is necessary to improve vaccine efficacy. We developed a nanoparticulate cancer vaccine by encapsulating a synthetic long peptide antigen within an immunologically inert nanoparticulate hydrogel (nanogel) of cholesteryl pullulan (CHP). After subcutaneous injection to mice, the nanogel-based vaccine was efficiently transported to the draining lymph node, and was preferentially engulfed by medullary macrophages but was not sensed by other macrophages and dendritic cells (so-called "immunologically stealth mode"). Although the function of medullary macrophages in T cell immunity has been unexplored so far, these macrophages effectively cross-primed the vaccine-specific CD8(+) T cells in the presence of a Toll-like receptor (TLR) agonist as an adjuvant. The nanogel-based vaccine significantly inhibited in vivo tumor growth in the prophylactic and therapeutic settings, compared to another vaccine formulation using a conventional delivery system, incomplete Freund's adjuvant. We also revealed that lymph node macrophages were highly responsive to TLR stimulation, which may underlie the potency of the macrophage-oriented, nanogel-based vaccine. These results indicate that targeting medullary macrophages using the immunologically stealth nanoparticulate delivery system is an effective vaccine strategy.

Does senescence affect lymph node number and morphology? A systematic review

Immunosenescence may contribute to an observed increase in infections and specific cancers in the elderly. Lymph nodes play a key role in the body's immune system. A systematic review was undertaken to investigate the effects of senescence on lymph node number and morphology. Electronic databases Ovid MEDLINE, Embase and Google Scholar were searched for relevant articles examining normal lymph node number and morphology with senescence. Data on lymph node number, gross anatomy and histo-architecture were collated and analysed. A total of 20 articles (15 human and 5 animal studies) were eligible for inclusion; many were limited by poorly standardized methods and relatively small sample sizes. However, there is evidence to suggest both a decrease in lymph node number and histological lymph node degeneration with senescence, at least in some lymph node basins. Degenerative changes include loss of lymphoid tissue from both the cortex and the medulla of lymph nodes, a reduction in the number and size of germinal centres, and changes such as hyalinization, fibrosis, fat deposition, a decrease in high endothelial venules and 'transparency'. In this first systematic review to examine changes in lymph nodes with senescence, evidence was accrued to suggest a decline in lymph node number and morphological degeneration in older age groups. These changes might adversely affect immune function and the prognosis of infections and selected cancers in the elderly. Further research is required to confirm these morphological changes and to explore their potential immunological and functional effects.

Identification of IL-7–Producing Cells in Primary and Secondary Lymphoid Organs Using IL-7–GFP Knock-In Mice

IL-7 is a cytokine crucial for development and maintenance of lymphocytes and other hematopoietic cells. However, how IL-7-expressing cells are distributed in lymphoid organs is not well known. To address this question, we established and analyzed IL-7-GFP knock-in mice. Thymic epithelial cells (TECs) expressed high GFP levels in the cortex and medulla, as detected with an anti-GFP Ab. Thymic mesenchymal cells also expressed GFP. Flow cytometry analysis suggested that cortical TECs expressed higher GFP levels than did medullary TECs. In bone marrow, immunohistochemistry indicated high levels of GFP in many VCAM-1(+) mesenchymal stromal cells and in some VCAM-1(-) cells. Additionally, half of the VCAM-1(+)CD31(-) stromal cells and some platelet-derived growth factor receptor α(+) stromal cells were GFP(+), as detected by flow cytometry. Moreover, we detected GFP expression in fibroblastic reticular cells in the T cell zone and cortical ridge of lymph nodes. Remarkably, lymphatic endothelial cells (LECs) expressed GFP at high levels within the lymph node medulla, skin epidermis, and intestinal tissues. Additionally, we detected abundant IL-7 transcripts in isolated LECs, suggesting that LECs produce IL-7, a heretofore unknown finding. Furthermore, GFP is expressed in a subpopulation of intestinal epithelial cells, and that expression was markedly upregulated in a dextran sulfate sodium-induced acute colitis model. Overall, IL-7-GFP knock-in mice serve as a unique and powerful tool to examine the identity and distribution of IL-7-expressing cells in vivo.

Plasma Cell Precursors: Long-Distance Travelers Looking for a Home

Little is known about the migration of plasma cell precursors to the lymph node medulla. In this issue of Immunity, Fooksman et al. (2010) propose that this migration is largely independent of chemotactic cues but follows a long linear walk of random orientation.

Capture of influenza by medullary dendritic cells via SIGN-R1 is essential for humoral immunity in draining lymph nodes

A major pathway for B cell acquisition of lymph-borne particulate antigens relies on antigen capture by subcapsular sinus macrophages of the lymph node. Here we tested whether this mechanism is also important for humoral immunity to inactivated influenza virus. By multiple approaches, including multiphoton intravital imaging, we found that antigen capture by sinus-lining macrophages was important for limiting the systemic spread of virus but not for the generation of influenza-specific humoral immunity. Instead, we found that dendritic cells residing in the lymph node medulla use the lectin receptor SIGN-R1 to capture lymph-borne influenza virus and promote humoral immunity. Thus, our results have important implications for the generation of durable humoral immunity to viral pathogens through vaccination.

Subcapsular sinus macrophages prevent lethal vesicular stomatitis virus (VSV) infection (43.11)

Abstract When a pathogen escapes innate host responses at the site of entry, it has the potential to spread systemically by accessing tissue-draining lymph vessels that channel interstitial fluid to the venous circulation. Before reaching the blood, the tissue-derived lymph must first pass through draining lymph nodes (LNs). LN act as filter stations preventing pathogens from reaching the systemic circulation and they are the staging grounds for adaptive immune responses, providing a specialized environment for the presentation of antigens to B and T cells. It is unclear how viruses are cleared from afferent lymph and how antiviral immune responses are generated. We have previously identified a population of CD11b+CD169+ macrophages on the floor of the subcapsular sinus (SCS) and in the medulla of LNs that capture viral particles within minutes after subcutaneous (sc) injection. Using VSV, an acutely cytopathic RNA virus closely related to rabies virus, we show here that these macrophages are critical to prevent viral access to nerve termini located in the SCS. Indeed, mice depleted of SCS macrophages develop an ascending CNS paralysis that leads to death in 50% of the animals 6-10 days after sc infection. Macrophage-depleted mice die despite mounting an increased neutralizing antibody (nAb) response. The notion that nAb are dispensable for protection against a sc infection (the natural route of transmission of this virus) is further demonstrated by the fact that antibody-deficient mice, which die upon intravenous infection, survive a sc VSV infection as long as SCS macrophages are present. While nAb are unnecessary for survival, B cells are required, and their protective role presumably relies on maintenance of LN architecture and normal macrophage phenotype.

Subcapsular sinus macrophages in lymph nodes clear lymph-borne viruses and present them to antiviral B cells

Lymph nodes prevent the systemic dissemination of pathogens such as viruses that infect peripheral tissues after penetrating the body's surface barriers. They are also the staging ground of adaptive immune responses to pathogen-derived antigens. It is unclear how virus particles are cleared from afferent lymph and presented to cognate B cells to induce antibody responses. Here we identify a population of CD11b+CD169+MHCII+ macrophages on the floor of the subcapsular sinus (SCS) and in the medulla of lymph nodes that capture viral particles within minutes after subcutaneous injection. Macrophages in the SCS translocated surface-bound viral particles across the SCS floor and presented them to migrating B cells in the underlying follicles. Selective depletion of these macrophages compromised local viral retention, exacerbated viraemia of the host, and impaired local B-cell activation. These findings indicate that CD169+ macrophages have a dual physiological function. They act as innate 'flypaper' by preventing the systemic spread of lymph-borne pathogens and as critical gatekeepers at the lymph-tissue interface that facilitate the recognition of particulate antigens by B cells and initiate humoral immune responses.

Association of SIGNR1 with TLR4–MD-2 enhances signal transduction by recognition of LPS in gram-negative bacteria

SIGNR1, a member of a new family of mouse C-type lectins, is expressed at high levels in macrophages (Mphi) within the splenic marginal zone, lymph node medulla, and in some strains, in peritoneal cavity. We previously reported that SIGNR1 captures gram-negative bacteria, such as Escherichia coli and Salmonella typhimurium, as well as Candida albicans. We have now investigated the precise ligands and innate responses that involve SIGNR1. The interaction of SIGNR1 with FITC-dextran and E. coli was completely inhibited by LPS from E. coli and Salmonella minnesota. Using LPS from various types of rough mutants of Salmonella, we found that SIGNR1 primarily recognizes oligosaccharides in the non-reductive end of the LPS core region. In transfectants, expression of SIGNR1 enhanced the oligomerization of Toll-like receptor (TLR) 4 molecules as well as the degradation of IkappaB-alpha after stimulation with E. coli under low-serum conditions. The enhanced TLR4 oligomerization was inhibited by pre-treatment of the cells with anti-SIGNR1 mAb or with mannan. A physical association between SIGNR1 and the TLR4-MD-2 complex was also observed by immunoprecipitation. Finally, we found that transfection of SIGNR1 into the macrophage-like RAW264.7 cells resulted in significant augmentation of cytokine production. These results suggest that SIGNR1 associates with TLR4 to capture gram-negative bacteria and facilitate signal transduction to activate innate M responses.

A novel endothelial L-selectin ligand activity in lymph node medulla that is regulated by alpha(1,3)-fucosyltransferase-IV.

Lymphocytes home to peripheral lymph nodes (PLNs) via high endothelial venules (HEVs) in the subcortex and incrementally larger collecting venules in the medulla. HEVs express ligands for L-selectin, which mediates lymphocyte rolling. L-selectin counterreceptors in HEVs are recognized by mAb MECA-79, a surrogate marker for molecularly heterogeneous glycans termed peripheral node addressin. By contrast, we find that medullary venules express L-selectin ligands not recognized by MECA-79. Both L-selectin ligands must be fucosylated by α(1,3)-fucosyltransferase (FucT)-IV or FucT-VII as rolling is absent in FucT-IV+VII−/− mice. Intravital microscopy experiments revealed that MECA-79–reactive ligands depend primarily on FucT-VII, whereas MECA-79–independent medullary L-selectin ligands are regulated by FucT-IV. Expression levels of both enzymes paralleled these anatomical distinctions. The relative mRNA level of FucT-IV was higher in medullary venules than in HEVs, whereas FucT-VII was most prominent in HEVs and weak in medullary venules. Thus, two distinct L-selectin ligands are segmentally confined to contiguous microvascular domains in PLNs. Although MECA-79–reactive species predominate in HEVs, medullary venules express another ligand that is spatially, antigenically, and biosynthetically unique. Physiologic relevance for this novel activity in medullary microvessels is suggested by the finding that L-selectin–dependent T cell homing to PLNs was partly insensitive to MECA-79 inhibition.

SIGN-R1, a novel C-type lectin expressed by marginal zone macrophages in spleen, mediates uptake of the polysaccharide dextran.

The marginal zone macrophages of the spleen are implicated in the clearance of polysaccharides, but underlying mechanisms need to be pinpointed. SIGN-R1 is one of five recently identified mouse genes that are homologous to human DC-SIGN and encode a single, external, C-terminal C-type lectin domain. We find that a polyclonal antibody to a specific SIGN-R1 peptide reacts primarily and strongly with a subset of macrophages in the marginal zone of spleen and lymph node medulla. In both sites, SIGN-R1 exists primarily in an aggregated form, resistant to dissociation into monomers upon boiling in SDS under reducing conditions. Upon transfection into three different cell lines, high-mol.-wt forms bearing SIGN-R1 are expressed, as well as reactivity with ER-TR9, a mAb previously described to react selectively with marginal zone macrophages. SIGN-R1-expressing macrophages preferentially sequester dextrans following i.v. injection. Likewise, when phagocytic cells are enriched from spleen and tested in culture, dextran is selectively endocytosed by a subset of very large SIGN-R1(+) cells representing approximately 5% of total released macrophages. Uptake of FITC-dextran by these macrophages in vivo and in vitro is blocked by ER-TR9 and polyclonal anti-SIGN-R1 antibodies. Following transfection with SIGN-R1, cell lines become competent to endocytose dextrans. The dextran localizes primarily to compartments lacking transferrin receptor and the LAMP-1 CD107a panlysosomal antigen. Therefore, SIGN-R1 mediates the uptake of dextran polysaccharides, and it is predominantly expressed in the macrophages of the splenic marginal zone and lymph node medulla.

Development of immune hyperinnervation in NGF-transgenic mice.

Sympathetic innervation of lymphoid tissues is localized to specific tissue compartments, but little is known of the "factors" that are important in establishing this pattern during development. Numerous studies have shown interactions of nerve growth factor (NGF) with the immune system, which may include modulation of immune innervation. We previously have shown that NGF transgenic mice, which overexpress NGF in skin and not immune tissues, have a dramatic hyperinnervation of splenic marginal zone and peripheral lymph node medulla and capsule. The purpose of the current studies was to determine if the presence of elevated NGF would alter immune system development and the process of sympathetic ingrowth. The results show that the splenic innervation in NGF transgenics gradually diverged from controls during the first two postnatal weeks, with the greatest change occurring between postnatal days 13 and 16 when the splenic organization was reaching the adult pattern. In contrast, the peripheral lymph nodes were hyperinnervated at an earlier age. mesenteric lymph nodes never diverged from the normal pattern. NGF levels in transgenic spleen were much higher than controls at postnatal days 1 and 2, when little innervation was present, and declined as the tissue matured, possibly because of NGF uptake by the ingrowing sympathetic fibers. This suggests that immune tissues are capable of concentrating NGF, which in turn may modulate the level of innervation by the sympathetic nervous system.

Immunohistochemical identification of caprine arthritis-encephalitis virus in paraffin-embedded specimens from naturally infected goats.

The expression of caprine arthritis-encephalitis virus capsid protein was studied in seropositive naturally infected asymptomatic goals (10< seropositive naturally infected encephalitic kids (12) and goats (4), and noninfected control goats (3). Rabbit antiserum to recombinant viral capsid and matrix proteins were used in a biotin-streptavidin-alkaline phosphatase complex immunohistochemical method on sections of formalin- and ethanol-fixed tissue specimens. Macrophages in inflamed areas of the lung (8/12), in the brain (5/16), and in the spinal cord (4/16) from encephalitic animals harbored viral antigens, as revealed by immunohistochemistry and use of a capsid protein-specific antiserum. Altogether 12/16 encephalitic animals tested positive for viral antigen. Viral antigens were found in 5/10 seropositive asymptomatic goals in macrophages located in the lung (3), the udder (1), and the medulla of lymph nodes (4). None of the control animals tested positive for viral antigen. Ethanol fixation showed highest sensitivity, and the lowest antigen concentration that revealed a positive signal discernible from background was twofold higher in ethanol-fixed specimens than in formalin-fixed specimens. The evaluation was performed on artificial antigen substrates embedded with defined concentrations of recombinant viral capsid protein. Immunohistochemistry is a valuable supplement to the methods presently available for diagnosis in cases suspicious of caprine arthritis-encephalitis.

Morphological changes in lymph nodes and expression of VCAM1 and cytokines at the late stages of SIV-induced disease in rhesus monkeys

Four patterns of structural alterations were found in lymph nodes (LNs) from rhesus monkeys 17 to 34 months after infection with simian immunodeficiency virus (SIV-mac251). SIV p27gag antigen and viral particles were localized either between the processes of follicular dendritic cells (FDCs) or in the cytoplasm of macrophages. In hyperplastic follicles, enlarged germinal centres contained numerous Ki67+ proliferating centroblasts which were rather rare in light zones occupied by the CD23+ FDC network. Involuted follicles contained a small number of Ki67+ centroblasts and the CD23 labelling was limited to a very small apical zone. A correlation was found between the morphological characteristics of the follicles (hyperplasia-involution) and the level of expression of the vascular cell adhesion molecule 1 (VCAM1) on FDCs. A gradient in VCAM1 intensity with no expression in the subcapsular-intermediary sinuses, low membrane labelling in the mantle and strong expression in the FDC network was observed. IL1 alpha+ and IL6+ (interleukin) cells (lymphocytes and macrophages) were detected in the mantle, the interfollicular area and the medulla of LNs. Expression of the tumour necrosis factor alpha and ultrastructural markers of interferon alpha production were found in a few FDC and macrophages. Our findings indicate a close relationship between the morphofunctional properties of FDC and the LN structure in SIV infection.

Identification of macrophages and dendritic cells in the osteopetrotic (op/op) mouse.

We used a panel of monoclonal antibodies and immunocytochemistry to identify macrophages and dendritic cells in mice that are deficient in macrophage colony stimulating factor (M-CSF or CSF-1) because of the recessive osteopetrotic (op/op) mutation. Prior work had shown that osteopetrosis is associated with a lack of osteoclasts, phagocytic cells required for remodelling in bone. Additional macrophage populations proved to be very M-CSF dependent. op/op mice had few and sometimes no peritoneal cavity phagocytes, splenic marginal zone metallophils, and lymph node subcapsular sinus macrophages. Other populations, however, reached substantial levels in the absence of M-CSF, including phagocytes in the thymic cortex, splenic red pulp, lymph node medulla, intestinal lamina propria, liver (Kupffer cells), lung (alveolar macrophages) and brain (microglia). Dendritic cells, which are specialized accessory cells for T-dependent immune responses and tolerance, were readily identified in skin and in the T-dependent regions of spleen, lymph node and Peyer's patch. The identification of dendritic cells utilized antibodies to MHC class II products and four different antigens that are primarily expressed by these accessory cells. Our findings indicate that only a few macrophage populations are critically dependent upon M-CSF in vivo. With respect to dendritic cells, the data are consistent with prior in vitro work where it was noted that GM-CSF but not M-CSF supported dendritic cell viability, function and growth.