The ANAE reaction pattern of reticular stromal cells of lymphatic organs from normal and hydrocortisone--treated mice

The pattern of alpha-naphthyl-acetate esterase (ANAE) activity in stromal and lymphatic cells has been examined in lymphatic organs from normal and hydrocortisone (HC)-treated mice. The distribution of ANAE-positive lymphocytes as well as stromal cells in peripheral lymphatic organs was different in T- and B-dependent regions, while in the thymus there was the difference between cortex and medulla. The HC application evoked a marked changes in the pattern of ANAE-positive cells distribution of the thymus, producing only a slight ones in the remaining organs. The influence of HC on the ANAE reaction intensity and distribution in the examined tissue sections is discussed.     

Migration inhibitory factor expression in experimentally induced endotoxemia.

Macrophage migration inhibitory factor (MIF) is an important constituent of the host response to stress and infection and is the first mediator that has been identified to be released from immune cells upon stimulation with glucocorticoids. MIF also has been shown to be secreted from the anterior pituitary gland, monocytes/macrophages, and T cells activated by various proinflammatory stimuli. Once released, MIF acts to counter-regulate the inhibitory effect of glucocorticoids on inflammatory cytokine production. To characterize more precisely the role of MIF in the host response to infection, we undertook a systematic analysis of MIF expression in various organs of the rat after endotoxin (lipopolysaccharide) administration. MIF protein and mRNA were analyzed by immunohistochemistry and in situ hybridization, respectively. MIF was found to be expressed constitutively in organs such as the lung, liver, kidney, spleen, adrenal gland, and skin. Significant quantities of MIF protein were detected preformed in various cell types and appeared to be released as a consequence of endotoxemia. In virtually all tissues examined, the loss of MIF protein 6 hours after lipopolysaccharide administration was accompanied by the induction of MIF mRNA and, at 24 hours, by the restoration of immunoreactive, intracellular MIF. The constitutive production of MIF by several cell and tissue types together with its rapid release from intracellular pools distinguishes MIF from other cytokines or hormonal mediators and significantly expands the physiological role of this unique counter-regulator of glucocorticoid action.     

Macrophage migration inhibitory factor expression in male and female ethanol-fed rats.

Macrophage migration inhibitory factory (MIF) regulates macrophage accumulation at sites of injury and can promote the inflammatory response. We studied MIF expression in the intragastric feeding rat model for alcoholic liver injury. Male and age-matched female rats were fed ethanol or dextrose with fish oil. Two groups of male rats were fed medium-chain triglycerides with ethanol or dextrose. Analysis of liver histopathology, lipid peroxidation, endotoxin, mRNA, and immunohistochemistry for MIF, tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) were carried out. Male and female rats fed fish oil and ethanol showed necroinflammatory liver injury and had the highest expression of MIF, TNF-alpha, and IFN-gamma in the liver. Decreased levels of MIF protein were seen in rats with higher endotoxin levels, suggesting that preformed MIF is released into the circulation. MIF is an important mediator of the inflammatory response in alcoholic liver disease and a potential therapeutic target.     

Acute toxicity of an anti-Fas antibody in mice.

By histopathologic examination of various organs in 3 normal strains, C3H/HeN, ICR, and DBA/1J, of mice treated intravenously once with anti-Fas antibody (Jo2), we failed to determine any target organ, except the liver, responsible for the acute lethality induced by the Fas/anti-Fas antibody interaction. However, we could show the presence of Fas-mediated apoptosis in other organs aside from the liver and normal mouse strain differences in susceptibility to anti-Fas antibody. Among these strains, C3H/HeN was the most susceptible to the antibody, followed by ICR and DBA/1J. We observed Fas-mediated apoptosis in the liver, spleen, thymus, lymph nodes, Peyer's patch, intestine, skin, coagulation glands, ovary, uterus, and vagina in all 3 strains and additionally in the epididymides and seminal vesicles in the DBA/1J strain. We also demonstrated that Fas-mediated apoptosis of small lymphocytes in the mantle zone of splenic lymphatic follicles preceded that of the hepatocytes or thymic cells. Since cellular damage was most severe in the liver among all the apoptotic organs in the 3 mouse strains, liver injury induced by anti-Fas antibody is speculated to play a significant role in the death.     

Stress-induced galectin-1 influences immune tolerance in the spleen and thymus by modulating CD45 immunoreactive lymphocytes

Galectin-1 (Gal-1) is differentially expressed in normal and pathological tissues and regulates immune cell homeostasis. Restraint stress increases serum Gal-1 in rats. However, the function of stress-induced Gal-1 in serum is unknown. We determined if stress-induced Gal-1 in serum accumulates in immunocompetent organs as protection from physiological and/or psychological stress. Western blotting showed that the intensity of Gal-1 bands in stressed groups was significantly higher than that in controls. RT–PCR analysis indicated that the Gal-1 mRNA level did not increase after restraint stress. The numbers of Gal-1 immunoreactive cells in the splenic periarterial lymphatic sheath (PLS) and the thymus medulla of the stressed group were increased compared with those in controls. Furthermore, stress-induced Gal-1 immunoreactive cells corresponded to CD45 immunoreactive lymphocytes (CD45⁺) in the PLS of the spleen and the medulla of the thymus. Thus, stress-induced Gal-1 immediately accumulates in the spleen and thymus, and may modulate the immune response through apoptosis by binding to CD45⁺ lymphocytes in immune organs following physiological and/or psychological stress.     

Early changes of hematopoietic tissues in mice treated with intravenous pulse-doses of 7,12-dimethylbenz(a)anthracene

Repeated intravenous injections of 7,12-dimethylbenz(a)anthracene (abbr. DMBA), four times weekly, induced several kinds of tumors in 26 mice (83.9%) out of 31 ddO mice, including 13 leukemias and 6 ovarian tumors. All leukemias were of lymphatic type, and 8 cases out of 13 leukemias were of thymus type. The thymus was involved by leukemic cells in 12 cases. All ovarian tumors were diagnosed as granulosa cell tumors histologically. During the treatment of DMBA, two types of changes in organs and blood were observed: one was a striking decrease of weight or organs or number of cells followed by prompt recovery as found in the spleen, red blood cells, and granulocytes in the peripheral blood; the other was moderate but long-standing decrease with little recovery as found in the thymus, ovaries, uterus, and lymphocytes in the peripheral blood. Unlike DMBA, the weight of the thymus was strikingly decreased by predonisolone, one of corticosteroids, whereas reactions of the spleen and the lymph nodes were not remarkable. Autoradiographical studies revealed a severe reduction followed by a marked rebound of labeling index in the red pulp of the spleen, but not in the white pulps of the spleen and cortex of the thymus. From these observations, it is concluded that a mild but long standing injury was noticed in the tissues which were the main sites of the tumorigenesis in mice by DMBA, and a severe but transient injury may not be related to induction of tumors in mice by DMBA.     

正常大鼠不同脏器微血管通透性的定量研究

大鼠颈动脉注射１％ＦｌＮａ荧光显微镜下活体观察肠系膜微血管血流状态及ＦｌＮａ的渗出情况。同时，在不同时点经股动脉采血，测定血浆内ＦｌＮａ浓度随时间的变化，用组织匀浆测定不同脏器中ＦｌＮａ浓度随时间的变化规律，并辅以冰冻切片进行观察。活体观察发现。ＦｌＮａ注入体内后，经微血管通融向周围组织渗出，最后汇集了淋巴管。血浆及组织匀浆ＦｌＮａ浓度的测定表明，不同脏器组织ＦｌＮａ浓度随时间的衰减过程各不相同，     

Lymphatic system of mice born from mothers treated with ampicillin or cloxacillin during gestation

Ampicillin, an antibiotic, widely used for combating bacterial infections exerts great influence on cells of the immune system of mouse and man. We have studied the effect of ampicillin and cloxacillin treatment of mice in the final week of pregnancy on the development of the lymphatic system of their offsprings. The mice born from antibiotic or saline treated mothers were examined 30 days after delivery. The examination of relative organ weight, cellularity and histopathological picture of lymphatic system (thymus, spleen and lymph nodes) and some other organs was performed. In the group of mice from ampicillin treated mothers we have found decreased relative weight of thymus and spleen and increased weight of lymph nodes with increased cellularity in thymus and lymph nodes. In the group of mice from cloxacillin treated mothers increased cellularity of thymus and lymph nodes was found. The histopathological study of lymphatic organs structure did not reveal any specific changes but the symptoms of focal degeneration and fat necrosis were found in livers of mice born from ampicillin treated mothers. Moreover, in mice born from antibiotic treated mothers the significant lymphocytosis in the peripheral blood was assessed. It was accompanied by an increase of granulocyte number in offsprings from ampicillin treated mothers and increase of monocyte number in those of cloxacillin treated. In conclusion, it could be suggested that ampicillin treatment during pregnancy would exert some effect on the development of lymphopoietic system of fetuses.     

Role of macrophage migration inhibitory factor in hepatitis B virus-specific cytotoxic-T-lymphocyte-induced liver injury

Macrophage migration inhibitory factor (MIF) plays a pivotal role in the development of various inflammatory diseases. Here, we found that anti-mouse MIF antibody treatment reduced liver injury and inflammatory cell infiltration into the liver after injection of antigen-specific cytotoxic T lymphocytes into hepatitis B virus transgenic mice.     

Viral protein sythesis by tissues from avian leukosis virus-infected chickens. II. Effect of passive neutralizing antibody in normal and agammaglobulinaemic chickens.

Synthesis in vitro of avian leukosis virus (ALV) group (gs proteins, p27 and p12, by various tissues from chickens infected within a few days after hatching was studied by means of autoradiography of immunoelectrophoretic patterns. Viral protein was synthesized in all tissues of chicks examined between days 18 and 50 of age after which time liver, kidney, bursa, thymus, and spleen became negative. The lung and genital organs of the chicks, however, continued to synthesize viral protein up to 100 days of age, when the experiment was ended. Repeated injections of neutralizing chicken antibody to ALV (Ab) starting on day 26 or 37 caused gs protein to decrease in spleen, liver, and thymus within 5 days but not in bursa, lung, and genital organs. Agammaglobulinaemic (Agamma) chickens showed prolonged persistence of gs protein synthesis in the spleen and liver; this synthesis was abrogated by passive Ab. Liver from Agamma chickens, however, also became negative without Ab treatment. The relative roles of antibody and cellular immunity in influencing ALV replication during the initial phase of infection before lymphoma development are discussed.     

EARLY CHANGES OF HEMATOPOIETIC TISSUES IN MICE TREATED WITH INTRAVENOUS PULSE-DOSES OF 7,12-DIMETHYLBENZ(a)ANTHRACENE

Repeated intravenous injections of 7,12-dimethylbenz(a)anthracene (abbr. DMBA), four times weekly, induced several kinds of tumors in 26 mice (83.9%) out of 31 ddO mice, including 13 leukemias and 6 ovarian tumors. All leukemias were of lymphatic type, and 9 cases out of 13 leukemias were of thymus type. The thymus was involved by leukemic cells in 12 cases. All ovarian tumors were diagnosed as granulosa cell tumors histologically. During the treatment of DMBA, two types of changes in organs and blood were observed: one was a striking decrease of weight of organs or number of cells followed by prompt recovery as found in the spleen, red blood cells, and granulocytes in the peripheral blood; the other was moderate but long-standing decrease with little recovery as found in the thymus, ovaries, uterus, and lymphocytes in the peripheral blood. Unlike DMBA, the weight of the thymus was strikingly decreased by predonisolone, one of corticosteroids, whereas reactions of the spleen and the lymph nodes were not remarkable. Autoradiographical studies revealed a severe reduction followed by a marked rebound of labeling index in the red pulp of the spleen, but not in the white pulps of the spleen and cortex of the thymus. From these observations, it is concluded that a mild but long standing injury was noticed in the tissues which were the main sites of the tumorigenesis in mice by DMBA, and a severe but transient injury may not be related to induction of tumors in mice by DMBA.     

The role of macrophage migration inhibitory factor in the cascade of events leading to reperfusion-induced inflammatory injury and lethality

Ischemia and reperfusion (I/R) injury is associated with a systemic inflammatory response, characterized by intense tumor necrosis factor (TNF)-alpha production and TNF-alpha-dependent tissue injury. Macrophage migration inhibitory factor (MIF) is a potent proinflammatory cytokine that may induce TNF-alpha release and play an important role in innate immune and inflammatory responses. The aim of this work was to assess whether MIF was involved the inflammatory cascade and injury that follows intestinal I/R. To this end, wild-type (WT) and MIF-deficient (MIF(-/-)) mice underwent 60 minutes of ischemia followed by 60 minutes of reperfusion, after which they were culled for the assessment of inflammatory parameters. I/R was accompanied by an increase in circulating levels of MIF and an increase of vascular permeability, hemorrhage, and production of TNF-alpha in the intestine and lungs. The latter parameters were markedly suppressed in reperfused MIF(-/-) mice, and this was associated with decreased lethality (80% in WT versus 20% in MIF(-/-) mice). Interestingly, the reperfusion-associated neutrophil accumulation in the intestine and lungs was similar in WT and MIF(-/-) mice. Leukocytes isolated from lungs of MIF(-/-) mice were less activated, as assessed by their response to zymosan in a luminol-enhanced chemiluminescence assay. In conclusion, our results suggest that MIF plays an important role in the cascade of events leading to TNF-alpha production and reperfusion-induced tissue injury and lethality in mice.     

Phospholipids and cholesterol in the liver mitochondrial fraction in mice with transplantable leukemia P 388

BDF1 hybrid mice, were given 10(4) lymphatic leukemia cells P 388 intraperitoneally. They were killed on 5th day and 11th day of the experiment. In the plasma the level of cholesterol was determined. Inner organs, i.e. liver, spleen, lymph nodes and thymus were verified histopathologically. Mitochondrial fraction lipids were extracted from liver and separated by thin layer chromatography. It was proved that the amount of mitochondrial fraction lipids of leukemic mice was considerably higher than in healthy mice and it increased together with the process of the neoplasma. In the group of leukemic animals the level of phosphatidylcholine, phosphatidylethanolamine, cholesterol and sphingomyelin increased in comparison with healthy mice. The results concerning phospholipids and cholesterol level in leukemic animals were interpreted in the aspect of the interaction of these compounds with vitamin E, polyunsaturated fatty acids and free radicals.     

Intrasplenic hepatocyte transplantation in rats with experimental liver injury: morphological and morphometric studies

Isolated rat hepatocytes were transplanted into the splenic parenchyma of syngeneic animals. The effects on the degree of colonization by the transplanted cells of three forms of experimental liver injury in recipient animals were studied. Significant colonization was observed in animals with carbon tetrachloride (CCl4)-induced injury combined with portacaval shunt (PCS) and in animals with common bile duct (CBD) ligation but not in control animals or in animals with CCl4-induced injury alone. Transplanted cells in the CCl4/PCS group resembled normal hepatocytes. In contrast, in the CBD group, the intrasplenic hepatocytes exhibited a pattern of 'ductular metaplasia' similar to that observed in the obstructed liver of the recipients. Transplanted syngeneic hepatocytes can thus proliferate in the spleen in response to liver injury in the recipient. The morphological appearances of the transplanted cells can be modified depending on the nature of the liver injury.     

Immune interferon. II. Different cellular site for the production of murine macrophage migration inhibitory factor and interferon.

The production of macrophage migration inhibitory factor (MIF) and immune interferon (IF) by concanavalin A (Con A)-stimulated cultures of thymus, lymph node and spleen cells was investigated. It was found that all cultures produced MIF activity, whereas only spleen cells produced marked IF activity. The capacity to produce IF was found to be correlated with the macrophage content of a cell preparation as evidenced by staining for esterase-positive cells. Furthermore, column-purified spleen T cells produced MIF but no IF. Migration inhibition caused by residual mitogen could be ruled out. On the other hand, when macrophages grown from bone marrow cells were pre-exposed to supernatants of mitogen-stimulated lymphocytes, IF activity was released into freshly added medium while no significant MIF activity was found. IF was also found in supernatants of macrophage cultures after exposure to conventional inducers in vitro (polyinosinic-polycytidylic acid, Corynebacterium parvum) or in vivo (C. parvum), whereas no MIF was detected. An anti-Type I IF serum neutralized IF in supernatants from Con A-stimulated spleen cells but did not affect MIF in the same supernatants. This indicates that IF and MIF activity are associated with different molecules. It is, therefore, concluded that under the described conditions, IF and MIF are produced by different cells. T cells are the prime producers of MIF while IF is released by macrophages following induction by lymphokines.     

Virus-specific transcription and translation in organs of BALB / Mo mice: Comparative study using quantitative hybridization, in Situ hybridization,and immunocytochemistry

Expression of the endogenous Moloney leukemia virus (=M-MuLV) was studied in a variety of tissues of BALB/Mo mice. Previous experiments using quantitative hybridization techniques have shown that M-MuLV expression occurred predominantly in the lymphatic organs spleen and thymus. Here we have employed in situ hybridization and immunocytochemical techniques to detect virus RNA and protein expression in single cells of a large variety of different organs. The two techniques were employed in parallel and compared with quantitative hybridization experiments using total nucleic acids extracted from the different organs. This allowed estimation of the fraction of positive cells in a given organ and to relate it to the total viral-specific RNA present, and thus to assess the value and the limitations of each method for detection of virus expression. The results confirm that most or all cells of the lymphatic system express regularly high concentrations of viral RNA and proteins. We show furthermore that cells of many nonlymphatic organs can be productively infected with virus expressing viral functions. These include cells of the reproductive system, epithelial cells of intestine, brain, and liver, and hormone-producing cells of the pancreas. With the exception of the reproductive system, the fraction of virus-producing cells in all nonlymphatic organs, however, was much lower than in spleen and thymus. Our results indicated that productive infection with M-MuLV is not restricted to cells of specific tissues, although the frequency of infection may vary considerably. They furthermore demonstrate that regardless of cell type viral proteins are always synthesized once viral RNA has been transcribed. Our results suggest that active cell proliferation and hormone production may be among the cellular parameters that influence susceptibility to productive virus infection.     

THYMIC LYMPH FOLLICLES A HISTOPATHOLOGICAL STUDY OF 1,356 AUTOPSY CASES*

1) Eighteen cases with definite thymic lymph follicles were found among 1,356 autopsies. Eight of those were cases of Basedow's disease and the remaining 10 consisted of 2 cases of asthma syndrome one of which was associated with a type of allergic granulomatous angiitis, 1 case each of Wegener-like granulomatosis, periarteritis nodosa, rheumatoid arthritis, aplstic anemia, chronic glomerulonephritis, acute yellow liver atrophy, Ekiri syndrome and purulent necrotizing tracheo-bronchitis. 2) No morphological differences were found between the thymic lymph follicles and lymph follicles in other organs. The folliculated thymus did not indicate any specific morphological features. 3) Basically, in cases with thymic lymph follicles, the lymphatic tissue of other organs showed lymphatic follicular hypoplasia. 4) Speculative explanation of the histogenesis and significance of thymic lymph follicle formation was attempted. ACTA PATH. JAP. 16:109-130, 1966     

The relevance of the migration inhibitory factor (MIF) for peripheral tissue response in murine sublethal systemic Aspergillus fumigatus infection

We recently demonstrated that macrophage migration inhibitory factor deficient (MIF (- / -)) mice exhibited a higher susceptibility to lethal systemic Aspergillus fumigatus infections than genetically matched, wild-type (WT) C57BL/6 mice, and displayed altered cytokine profiles in the spleen when challenged by sublethal infections. In this report we focused on the potential involvement of MIF in the response of mice to sublethal systemic A. fumigatus infections in tissues other than spleen. Impaired fungal clearance from lungs, kidneys, liver and brain in MIF (- / -) mice was noted and was associated with histologically-evident differences in signs of inflammation in these organs. Higher values of some indicators of pathologic changes in urine parameters (increases in bilirubin, glucose and ketones), as well as a greater degree of brain tissue damage, pointed to multiple organs being affected in MIF (- / -) mice. Analysis of the lung response revealed differences in the composition of infiltrated cells between MIF-sufficient and MIF-deficient mice. MPO activity and reactive oxygen species production were impaired, as well as production of IL-17 and IFN-γ in MIF (- / -) mice as compared to WT counterparts. Lower systemic IL-1β and IL-6 levels in infected MIF (- / -) mice coincided with reduced blood neutrophil counts and organ infiltration. Collectively, this study identifies MIF as a resistance factor that orchestrates events in several non-lymphoid areas which provide a milieu that accomplishes anti-fungal A. fumigatus defense.     

Moloney leukemia virus gene expression and gene amplification in preleukemic and leukemic BALB/Mo mice

Mice carrying the exogenous Moloney leukemia virus (M-MuLV) as an endogenous virus have been derived previously. This mouse strain (BALB/Mo) transmits the virus as a single Mendelian gene (Mov-1 locus) from one generation to the next. Molecular hybridization experiments were performed to identify the organs in which the M-MuLV gene is activated during postnatal life of BALB/Mo mice and to determine the age-dependent onset of M-MuLV gene expression. Spleen and thymus cells of BALB/Mo mice synthesize M-MuLV-specific RNA soon after birth and virus gene expression reaches high levels at 3–4 weeks of age. A substantial further increase in virus gene expression is not observed in leukemic tissues of older animals. Other organs, such as liver, brain, and kidneys, do not express M-MuLV-specific RNA throughout the animals' life. These observations define lymphatic tissues (spleen and thymus) as the target organs of M-MuLV expression in BALB/Mo mice. Virus gene expression was correlated with somatic amplification of M-MuLV-specific DNA sequences during the preleukemic and leukemic phase of the animals' life. DNA amplification occurs in two steps in target tissues of BALB/Mo mice. A first step to approximately two copies per haploid genome equivalent is observed in preleukemic mice and a second step to three to four copies is observed in leukemic tissues. Nontarget tissues carry one copy of M-MuLV-specific DNA sequences regardless of the age of the animal. These results suggest that M-MuLV-specific gene expression is not sufficient for leukemic transformation and is related to virus-specific DNA amplification in preleukemic animals. A second amplification of M-MuLV DNA sequences appears to be related to leukemic transformation.