Central Macrophage Research Articles

Origin and fate of the central macrophages of erythroblastic islands in the fetal and neonatal mouse liver.

Erythroblastic islands were examined by ultrastructure and ultrastructural histochemistry in fetal and early neonatal livers of the mouse. The liver primordium of day 11 embryos contained not only immature hemopoietic cells in the hepatic cords but also macrophages in expanded sinusoids. At 12 and 13 days of gestation, macrophages bearing large cytoplasmic inclusions increased in number, and some of them moved from the sinusoids into the hepatic cords. A ring of erythroblasts surrounded the macrophages, and erythroblastic islands could be identified at 14 days of gestation. Fetal livers contained two kinds of macrophages: sinusoidal macrophages and central macrophages of the erythroblastic islands. These macrophages exhibited a similar binding pattern to Griffonia simplicifolia isoagglutinin-IB4 (GS-IB4) and soybean agglutinin (SBA). Fetal hepatocytes, however, did not appear to engage in active phagocytosis, and the binding patterns of GS-IB4 and SBA differed significantly between hepatocytes and the two kinds of macrophages. In the early postnatal mouse, a marked decrease in the number of erythroblastic islands occurred. Erythroblasts left the central macrophages, and the macrophages subsequently underwent degeneration. The erythroblastic islands finally disappeared at the end of liver hemopoiesis, and the degenerated central macrophages were removed by scavenger macrophages in the perisinusoidal space. Our data demonstrate that scavenger macrophages play an essential role in the development of hepatic hemopoiesis, with special reference to the formation and dissolution of erythroblastic islands.

Origin and fate of the central macrophages of erythroblastic islands in the fetal and neonatal mouse liver

Origin and fate of the central macrophages of erythroblastic islands in the fetal and neonatal mouse liver

Hemopoiesis in human fetal and embryonic liver.

In this review, we describe the topographic distribution of hemopoietic cells of the lymphoid, myeloid, and erythroid lineages in the human fetal and embryonic liver. The data are based on studies of frozen tissue, allowing the determination of a broad range of hemopoietic antigens, and studies on paraffin-embedded tissue, allowing the combination of optimal morphology and immunodetection of lineage-specific antigens. The different hemopoietic lineages each show their own immunophenotype and distribution; intercellular and microenvironmental relationships were easily determined. In a few cases, some scarce CD34-positive early progenitor cells were seen. The number of proliferating cells, identified by monoclonal antibody (MAb) Ki-67, varied from 350 to 2500 (median = 1,500) per square millimeter of tissue. Erythroid cells reacted with antisera to glycophorin A, CDw75, and CD43 and partly surround a central macrophage, whereas the myelomonocytic cells reacted with CD45, CD43, CD74, and antilysozyme serum, and with LN3 from 14 weeks onward. Myelopoietic (CD15 positive) cells were localized mainly around portal triad vessels and increased in number with gestational age. The lymphoid cells showed CD45, CD43, CD45RA/MT2, CD45RA/MB1, MB2, and CD74 reactivity. B cells and their precursors were scattered among the hepatocytes without any sign of focal development in the age range studied. We seldom found cells positive for delta-H chain or C3bR (CD35); C3dR (CD21)-positive cells were even more scarce. Cells reactive with MAb WT1 (CD7) were present in a scattered single pattern (< or = 20/mm2) among the parenchymal cells; cells expressing mature T-cell markers (CD2, CD3, CD5) were rare. Large (> 15 mu) CD43-positive hemopoietic cells in the fetal liver were distinguished that exclusively expressed CD43, probably representing early hemopoietic progenitor cells.

Murine thymic nurse cells and rosettes: analysis of adhesion molecule expression using confocal microscopy and a simplified enrichment method.

Thymic nurse cells (TNC) and T-cell stromal rosettes (ROS) are two in vivo models for stromal cell-thymocyte interactions. We describe a simplified enrichment method for TNC and ROS that overcomes the necessity for large amounts of tissue. The complexes were further analyzed with confocal microscopy, and three subunits of ROS were defined on the basis of their central cell phenotype, i.e., macrophage, dendritic, or epithelial cell rosettes. Because adhesion molecules are proposed to play a crucial role in T-cell development, we investigated CD44, LFA-1, and ICAM-1 expression in such complexes. The epithelial component of TNC expresses CD44 and ICAM-1, whereas intra-TNC thymocytes are LFA-negative. With regard to ROS, all subsets expressed CD44, and macrophage and dendritic cell ROS were also ICAM-1-positive and LFA-1-positive. The current protocol opens the possibility for further in vivo analysis of stromal cell-thymocyte interactions, e.g., for studies of scarce gene mutant mice.

Kinetics of Central Nervous System Microglial and Macrophage Engraftment: Analysis Using a Transgenic Bone Marrow Transplantation Model

AbstractTo determine the kinetics of tissue macrophage and microglial engraftment after bone marrow (BM) transplantation, we have developed a model using the ROSA 26 mouse. Transplanted ROSA 26 cells can be precisely identified in recipient animals because they constitutively express β-galactosidase (β-gal) and neomycin resistance. B6/129 F2 mice were irradiated and transplanted with BM from ROSA 26 donors and their tissues (spleen, marrow, brain, liver, and lung) examined at various time points to determine the kinetics of engraftment. Frozen sections from transplanted animals were stained histochemically for β-gal to identify donor cells. At 1, 2, 6, and 12 months posttransplantation, 98% to 100% of granulocyte-macrophage colonies were of donor (ROSA 26) origin determined by β-gal staining and by neomycin resistance. Splenic monocytes/macrophages were 89% donor origin by 1 month confirming quick and complete engraftment of hematopoietic tissues. At this time, only rare ROSA 26 tissue macrophages or microglia were observed. Alveolar macrophage engraftment was evident by 2 months and had increased to 61% of total tissue macrophages at 1 year posttransplantation. The kinetics of liver Kupffer cell engraftment were similar to those seen in the lung. However, donor microglial engraftment remained only 23% of total microglia at 6 months and increased to only 30% by 1 year. Also, donor microglia were predominantly seen at perivascular and leptomeningeal, and not parenchymal, sites. The data show that microglia derive from BM precursors but turn over at a significantly slower rate than other tissue macrophages. No clinical or histological graft-versus-host disease was observed in the recipients of ROSA 26 BM. These kinetics may impact strategies for the gene therapy of lysosomal storage diseases. Because individual donor cells can be identified in situ, the ROSA 26 model should have many applications in transplantation biology including studies of homing and differentiation.

Kinetics of Central Nervous System Microglial and Macrophage Engraftment: Analysis Using a Transgenic Bone Marrow Transplantation Model

To determine the kinetics of tissue macrophage and microglial engraftment after bone marrow (BM) transplantation, we have developed a model using the ROSA 26 mouse. Transplanted ROSA 26 cells can be precisely identified in recipient animals because they constitutively express β-galactosidase (β-gal) and neomycin resistance. B6/129 F2 mice were irradiated and transplanted with BM from ROSA 26 donors and their tissues (spleen, marrow, brain, liver, and lung) examined at various time points to determine the kinetics of engraftment. Frozen sections from transplanted animals were stained histochemically for β-gal to identify donor cells. At 1, 2, 6, and 12 months posttransplantation, 98% to 100% of granulocyte-macrophage colonies were of donor (ROSA 26) origin determined by β-gal staining and by neomycin resistance. Splenic monocytes/macrophages were 89% donor origin by 1 month confirming quick and complete engraftment of hematopoietic tissues. At this time, only rare ROSA 26 tissue macrophages or microglia were observed. Alveolar macrophage engraftment was evident by 2 months and had increased to 61% of total tissue macrophages at 1 year posttransplantation. The kinetics of liver Kupffer cell engraftment were similar to those seen in the lung. However, donor microglial engraftment remained only 23% of total microglia at 6 months and increased to only 30% by 1 year. Also, donor microglia were predominantly seen at perivascular and leptomeningeal, and not parenchymal, sites. The data show that microglia derive from BM precursors but turn over at a significantly slower rate than other tissue macrophages. No clinical or histological graft-versus-host disease was observed in the recipients of ROSA 26 BM. These kinetics may impact strategies for the gene therapy of lysosomal storage diseases. Because individual donor cells can be identified in situ, the ROSA 26 model should have many applications in transplantation biology including studies of homing and differentiation.

Scanning Electron Microscopic Study of Erythroblastic Islands Obtained by “Spleen-Stamp Culture” Method

During hematopoiesis of mammalian animals, an erythroblastic island in the bone marrow has been reported to contain a central macrophage surrounded by many erythroblasts. Fetal mouse livers and spleens, which are treated with collagenase, have been used to get the erythroblastic island in tissue cultures. In this study, we have cultured some stamped cells obtained from spleens of anemic mice and examined their structures by scanning electron microscopy.Hemolytic anemia of female BALB/c mice was induced by acetylphenylhydrazine (4mg/g body weight) injection for 5 days. Spleens were resected on the second day after the last injection and a “spleen-stamp culture“ method was performed. Briefly, they were cut with razor blades (Fig. 1a) and splenic cells on the cut tissue surface were stamped on collagen-coated coverslips (Fig.1b). The stamped splenic cells were covered with dialysis membranes (Fig. 1c). They were cultured with 55% IMDM, 20% L-15 and 25% FBS, containing 2U/ml erythropoietin (EPO) in humidified atmosphere of 5% CO2 at 37°C for 1-3 days.

Importance of cell-cell interactions in regulation of erythropoiesis.

In the bone marrow, erythropoiesis occurs in distinct anatomic units called erythroblastic islands that consist of a central macrophage surrounded by a ring of developing erythroblasts. The formation of erythroblastic islands involves adhesive interactions between the central macrophage and the ring of developing erythroblasts, between adjacent erythroblasts in the ring, and between macrophages/erythroblasts and the components of extracellular matrix. These adhesive interactions are mediated by specific pairs of cell surface receptors and counterreceptors (ligands) including integrins, the immunoglobulin superfamily, and cadherins. Several lines of evidence have suggested that cell-cell interactions among various cell types in the bone marrow play an important role in supporting and regulating hematopoietic differentiation. Cell surface adhesion molecules expressed during erythroid development and the likely counterreceptors for these molecules are discussed in this review.

Hydranencephaly, cerebellar hypoplasia, and myopathy in chick embryos infected with aino virus.

Pathogenesis of Aino virus (AIV), a suspected causative agent of congenital abnormalities of calves, has not yet been established by experimental infection of dams. To investigate the pathogenesis, 10(3) median tissue culture infective doses per 0.2 ml of AIV strain JaNAr 28 was inoculated into the yolk sac of 8-day-old chick embryos. At 4, 7, 10, and 13 days post-inoculation (PI) 20 eggs were opened and macro- and microscopic studies combined with virus recovery and immunohistochemical detection of the virus antigen were performed. At 7 to 13 days PI chick embryos manifested marked hydranencephaly, cerebellar hypoplasia, arthrogryposis, and scoliosis, with the highest incidences of 86.7%, 73.3%, 80.0%, and 20.0%, respectively. At 4 days PI the viral antigen was found in nerve cells, gitter cells in mild necrotic foci of the central nervous system (CNS), degenerative myotubules, and macrophages in the interstitium, which was associated with the early phase of AIV-induced encephalitis and polymyositis, with occasional accompanying hemorrhage and clumping of myotubular fragments. From 7 to 10 days PI, AIV antigen increased markedly in the liquefactive necrosis and in both degenerative and normal-looking myotubules in conjunction with developing hydranencephaly and arthrogryposis. The encephalitis and myositis had a tendency to mitigate by 10 days PI, coincident with a slight decrease in amount of AIV antigen. At 13 days PI there was almost no detectable AIV antigen in CNS and skeletal muscles, probably due to depletion of cells having affinity to AIV.

Theiler's murine encephalomyelitis virus kills restrictive but not permissive cells by apoptosis.

Theiler's murine encephalomyelitis viruses (TMEV), genus Cardiovirus, family Picorniviridae, are natural enteric pathogens of mice which cause central nervous system demyelination similar to that seen in multiple sclerosis. TMEV can be classified into two groups based on neurovirulence: a highly virulent group, e.g., GDVII virus, and a less virulent group, e.g., BeAn virus. Both viruses, depending on the multiplicity of infection, produced cytopathology in BSC-1 cells similar to that in BHK-21 cells. Since apoptosis has been reported as a mechanism of cell death after infection with many viruses, we examined infected BHK-21 and BSC-1 cells for morphological and biochemical changes consistent with apoptosis. Only the restrictive BSC-1 cells showed evidence of nuclear morphology and internucleosomal DNA degradation indicative of apoptosis. Interestingly, the more virulent GDVII virus was at least 50-fold more efficient in inducing apoptosis than the less virulent BeAn virus. This difference was not due to greater GDVII viral RNA replication or production of infectious virus, since the two viruses were similarly restricted in BSC-1 cells. Apoptosis in BSC-1 cells appears to be triggered by a cytoplasmic event, since inactivation of GDVII viral RNA by UV light abolished the ability of the virus to induce apoptosis. The possible role of apoptosis in the pathogenesis of TMEV infection in mice, especially virus persistence in central nervous system macrophages, is discussed.

Pre- and post-embedding immunoelectron microscopy of Ki-M1P immunoreactive germinal center macrophages using ultra-small gold probes with silver enhancement

The Ki-M1P protein is primarily detected in cells deriving from the monocyte/macrophage cell lineage. The aim of this study was to investigate the occurrence of Ki-M1P immunoreactivity in germinal center macrophages by immunohistochemical and immunocytochemical staining techniques. Ultra-small (0.8 nm) gold probes combined with silver enhancement were used as a detection system for pre- and post-embedding immunostaining both at the light and electron microscopic level. Ki-M1P-positive macrophages were observed at a constant frequency in the germinal centers of the follicles throughout the tonsillar lymphatic tissue. The specific immunostaining was localized in the cytoplasm of these cells. Electron microscopic examination demonstrated the presence of abundant lysosomes in the cytoplasm, and some of the germinal center macrophages contained phagocytosed cells (tingible bodies) showing signs of various degrees of digestion. Ki-M1P immunoreactivity, as revealed by depositions of silver-enhanced ultra-small gold probes, was confined to the periphery of the lysosomes and tingible bodies. The results obtained demonstrate that the use of silver-enhanced ultra-small gold probes is a highly sensitive and specific detection system for pre- and post-embedding immunostaining of the Ki-M1P protein, and provides, in general, a flexible system for combined light and electron microscopic examination of tissue antigens. Furthermore, in the cytoplasm of the germinal center macrophages a spatial association between the Ki-M1P protein and lysosomes and tingible bodies was observed. These findings may indicate that the Ki-M1P protein is connected with phagocytosis and/or processes related to intracellular digestion in these cells.

Normal adult ramified microglia separated from other central nervous system macrophages by flow cytometric sorting. Phenotypic differences defined and direct ex vivo antigen presentation to myelin basic protein-reactive CD4+ T cells compared.

Ramified microglia in the adult central nervous system (CNS) are the principal glial element up-regulating MHC class I and II expression in response to inflammatory events or neuronal damage. A proportion of these cells also express MHC class II constitutively in the normal CNS. The role of microglia as APCs for CD4+ T cells extravasating into the CNS remains undefined. In this study, using irradiation bone marrow chimeras in CD45-congenic rats, the phenotype CD45lowCD11b/c+ is shown to identify microglial cells specifically within the CNS. Highly purified populations of microglia and nonmicroglial but CNS-associated macrophages (CD45highCD11b/c+) have been obtained directly from the adult CNS, by using flow cytometric sorting. Morphologically, freshly isolated microglia vs other CNS macrophages are quite distinct. Of the two populations recovered from the normal CNS, it is the minority CD45highCD11b/c+ transitional macrophage population, and not microglia, that is the effective APC for experimental autoimmune encephalomyelitis-inducing CD4+ myelin basic protein (MBP)-reactive T cells. CD45highCD11b/c+ CNS macrophages also stimulate MBP-reactive T cells without addition of MBP to culture, suggesting presentation of endogenous Ag. This is the first study in which microglia vs other CNS macrophages have been analyzed for APC ability directly from the CNS, with substantial cross-contamination between the two populations eliminated. The heterogeneity of these populations in terms of APC function is clearly demonstrated. Evidence is still lacking that adult CNS microglia have the capacity to interact with and stimulate CD4+ T cells to proliferate or secrete IL-2.

Ethanol-induced depletion of lymphocytes from the mesenteric lymph nodes of C57B1/6 mice is associated with RNA but not DNA degradation.

The effects of ethanol (EtOH) consumption by adult female C57B1/6 mice on lymphocyte populations of the mesenteric lymph nodes (MLNs) were determined by feeding mice with the Lieber-DeCarli liquid diet by a pair-feeding paradigm. Histological analysis of the MLNs of EtOH-fed mice showed a progressive loss of lymphocytes from the medullary regions at 3, 5, and 7 days after initiation of the EtOH diet. The stromal cells in the medullary region also demonstrated a progressive alteration in stellate morphological features at times corresponding to those of loss of lymphocytes from this region. Microscopic evaluation of the follicle regions of MLNs obtained from mice fed an EtOH-containing diet showed no appreciable alterations in morphological characteristics. The number of tingible body macrophages in the germinal centers of the follicles, however, was increased after 3 days of EtOH diet feeding and declined progressively after this time. Flow cytometric analysis of isolated lymphocytes showed a depletion of both T and B cell populations from the MLNs. In contrast to B cells, however, T cells were depleted through 7 days of EtOH diet feeding. Total RNA isolated from the MLNs of mice consuming the EtOH-containing diet was progressively degraded. No degradation of DNA was observed. These study results establish that continuous consumption of dietary EtOH adversely affects the cellularity of MLN, resulting in a progressive loss of lymphocytes that is associated with degradation of total RNA.

Very late activation antigen 4-vascular cell adhesion molecule 1 interaction is involved in the formation of erythroblastic islands.

Erythroblastic islands are anatomical units consisting of a central macrophage surrounded by erythroblasts. We studied the adhesion molecules involved in the formation of these structures. Central macrophages of erythroblastic islands isolated from the spleens of phlebotomized mice were clearly stained for vascular cell adhesion molecule 1 (VCAM-1). The surrounding erythroblasts of the erythroblastic islands strongly expressed the alpha 4 integrin of very late activation antigen 4 (VLA-4: alpha 4 beta 1 integrin), the counter receptor of VCAM-1, whereas most reticulocytes and erythrocytes did not. Both monoclonal antibodies (mAbs) against alpha 4 integrin and VCAM-1 disrupted the erythroblastic islands cultured in the presence of erythropoietin. Moreover, adhesion of splenic erythroblasts to tumor necrosis factor alpha-stimulated mouse splenic endothelial cells, which showed high expression of VCAM-1 but not intercellular adhesion molecule 1, was inhibited by the anti-VCAM-1 and anti-alpha 4 mAbs. These findings suggest that VLA-4-VCAM-1 interaction plays a crucial role in the formation of erythroblastic islands.

The association of erythroblasts with macrophages promotes erythroid proliferation and maturation: a 30-kD heparin-binding protein is involved in this contact

Although the association of erythroblasts with macrophages has been well documented in the human bone marrow, the function and identification of the intimate contacts occurring between the membranes of these two cell types in the physiology of erythropoiesis is not known. Using in vitro cultures of human peripheral blood derived erythroid progenitors, we have shown the presence of erythroblastic islands consisting of a central macrophage surrounded by a ring of erythroblasts that undergo terminal maturation leading to enucleation. However, when cultures were carried in the absence of intact macrophages, erythroid cells matured to the late erythroblast stage but failed to enucleate. Furthermore, the number of erythroid cells was markedly reduced in macrophage-depleted cultures, suggesting that the erythroblast-macrophage contact promotes proliferation and terminal maturation of erythroid cells leading to their enucleation. To examine the molecule(s) involved in the interaction between erythroblasts and macrophages, we have used a cell attachment assay involving incubation of solubilized surface-labeled erythroblasts with macrophage membrane proteins separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to a nitrocellulose membrane. Erythroblast surface proteins specifically attached to a 30-kD protein from macrophage membranes, whereas no adhesion was seen to the protein standards. An apparently similar protein of 30 kD was also detected on erythroblasts and was shown to mediate erythroblast-erythroblast contact in addition to the erythroblast-macrophage contact. The extraction of plasma membranes with Triton X-100 showed that the 30-kD protein is linked to the membrane skeleton via an integral membrane protein both in erythroblasts and macrophages. Furthermore, our results show that the cell:cell interactions mediated by the 30-kD protein are calcium-independent and could be specifically inhibited by heparin. We conclude that the association of erythroblasts with macrophages promotes erythroid proliferation and maturation leading to erythroblast enucleation and that a 30-kD heparin-binding protein present on the surface of macrophages and erythroblasts is involved in this contact. This protein is capable of binding homotypic and heterotypic cells.

The Association of Erythroblasts With Macrophages Promotes Erythroid Proliferation and Maturation: A 30-kD Heparin-Binding Protein Is Involved in This Contact

Although the association of erythroblasts with macrophages has been well documented in the human bone marrow, the function and identification of the intimate contacts occurring between the membranes of these two cell types in the physiology of erythropoiesis is not known. Using in vitro cultures of human peripheral blood derived erythroid progenitors, we have shown the presence of erythroblastic islands consisting of a central macrophage surrounded by a ring of erythroblasts that undergo terminal maturation leading to enucleation. However, when cultures were carried in the absence of intact macrophages, erythroid cells matured to the late erythroblast stage but failed to enucleate. Furthermore, the number of erythroid cells was markedly reduced in macrophage-depleted cultures, suggesting that the erythroblast-macrophage contact promotes proliferation and terminal maturation of erythroid cells leading to their enucleation. To examine the molecule(s) involved in the interaction between erythroblasts and macrophages, we have used a cell attachment assay involving incubation of solubilized surface-labeled erythroblasts with macrophage membrane proteins separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to a nitrocellulose membrane. Erythroblast surface proteins specifically attached to a 30-kD protein from macrophage membranes, whereas no adhesion was seen to the protein standards. An apparently similar protein of 30 kD was also detected on erythroblasts and was shown to mediate erythroblast-erythroblast contact in addition to the erythroblast-macrophage contact. The extraction of plasma membranes with Triton X-100 showed that the 30-kD protein is linked to the membrane skeleton via an integral membrane protein both in erythroblasts and macrophages. Furthermore, our results show that the cell:cell interactions mediated by the 30-kD protein are calcium-independent and could be specifically inhibited by heparin. We conclude that the association of erythroblasts with macrophages promotes erythroid proliferation and maturation leading to erythroblast enucleation and that a 30-kD heparin-binding protein present on the surface of macrophages and erythroblasts is involved in this contact. This protein is capable of binding homotypic and heterotypic cells.

Physiological and clinical significance of nitric oxide.

Nitric oxide, or NO., is a gas under atmospheric conditions. It is noxious because of its free-radical structure. It is biosynthesized from the amino acid L-arginine. The responsible enzymes are widely distributed in the human body. It has been shown that this simple molecule, NO., plays important roles in mammalian physiology. It is one of the factors regulating vascular tone and blood pressure, inhibition of platelet aggregation, neurotransmission in the peripheral and central nervous systems and macrophage function. Evidence for the pathophysiological significance of NO. is now accumulating.

Scavenger macrophages and central macrophages of erythroblastic islands in liver hemopoiesis of the fetal and early postnatal mouse: a semithin light- and electron-microscopic study.

The livers of 12-day mouse embryos contain many hemopoietic cells in the hepatic cords and macrophages which are laden with large heterophagosomes in the sinusoids. Macrophages, as scavengers, engulfed circulating primitive erythroblasts from the yolk sac, as well as nuclei expelled from erythroblasts. At the 14th day of gestation, scavenger macrophages in mitosis were seen, extending long and thin cytoplasmic projections from their cell surface. At the 13-14 day of gestation, macrophages migrated from sinusoids into hepatic cords, and erythroblasts gathered around them, thus forming cell clusters designated as erythroblastic islets. Central macrophages formed a cell-socket-like structure on their cell surface for surrounding erythroblasts, reflecting the close association between macrophages and erythroblasts. In the erythroblastic islands, contained in late fetal and neonatal livers, erythroblasts have dissociated from the central macrophages having a starfish-shaped cell profile. The macrophage-erythroblastic association in the islands became less marked rapidly after birth.

Macrophage markers in diagnostic neuropathology.

It can be difficult to differentiate the cells of infiltrating gliomas from macrophages associated with demyelinating lesions or cerebral infarcts, especially in tissue with freezing artifact or tissue that is technically suboptimal. We therefore sought to identify a reliable immunohistochemical marker for central nervous system macrophages that could be used on routinely processed tissue. The reactions of commercially available markers [Ham-56, Mac 387, CD68, CR3/43, DR alpha, lysozyme, CD16, and Ricinus communis agglutinin-1 (RCA-1)] were studied in normal cerebellum, subacute infarcts, viral encephalitis with microglial nodules, radiation necrosis, Alzheimer's disease, medulloblastoma, T-cell lymphoma, oligodendroglioma, gliomatosis cerebri, and microgliomatosis cerebri. We also performed a more extensive investigation of Ham-56 with largely surgical material. We concluded that (a) RCA-1 is a sensitive marker for microglia, either resting or responding to injury, but staining must be interpreted carefully because endothelial cells and reactive astrocytes frequently stain as well. This marker was useful in highlighting focal inflammation in HIV encephalitis. (b) CD68 is a specific marker for resting microglia. A case of microgliomatosis cerebri was intensely stained. (c) Ham-56 is a sensitive, specific (with the exception of endothelial cells), and reliable marker of central nervous system macrophages in routinely processed autopsy and surgical material. It is especially useful in distinguishing the macrophages in demyelinating disease and cerebral infarcts from neoplastic glial cells.

Lymphoid pneumonitis in 50 adult patients infected with the human immunodeficiency virus: Lymphocytic interstitial pneumonitis versus nonspecific interstitial pneumonitis

Lymphocytic interstitial pneumonitis (LIP) and nonspecific interstitial pneumonitis (NIP) are pulmonary complications of human immunodeficiency virus (HIV) infection that occur in the absence of a detectable opportunistic infection or neoplasm. We reviewed lung biopsy specimens from 50 adult HIV-infected patients, of whom four had LIP and 46 had NIP. The majority (47 of 50) of specimens from patients with NIP showed mild chronic interstitial pneumonitis ( CIP NIP ), with three showing features of diffuse alveolar damage, organizing phase. In contrast to CIP NIP , the five specimens from four patients with LIP demonstrated more extensive lymphocytic interstitial infiltrates that extended into the alveolar septal interstitium. The majority of the interstitial lymphocytes in both NIP and LIP were of T-cell origin and stained for UCHL-1. The etiologies of NIP and LIP remain unknown. Since the common opportunistic infections were excluded by routine methods, we sought, with special techniques, to investigate whether HIV, Epstein-Barr virus (EBV), or cytomegalovirus (CMV) could be identified in lung biopsy specimens from these patients. By in situ hybridization, we found one LIP specimen with expression of large amounts of HIV RNA primarily within macrophages in germinal centers; in the remaining specimens, occasional cells expressing HIV RNA were found (two LIP and four NIP). Neither CMV nor EBV was found by in situ hybridization in seven specimens; in these same specimens EBV was detected using the polymerase chain reaction in only one case of NIP, similar to results in control specimens. These results, together with the knowledge that lymphocytic pulmonary lesions may be caused by lentiviruses in humans and animals, suggest that HIV plays a significant role in the pathogenesis of both NIP and LIP in adult HIV-infected patients; in contrast, our data do not demonstrate a direct role for either EBV or CMV.