Estrogen stimulates neuronal plasticity in the deafferented hypothalamic arcuate nucleus in aged female rats

The hypothalamic arcuate nucleus (ARCN) was examined ultrastructurally 3 weeks after the complete deafferentation of the medial basal hypothalamus (MBH) with the island isolation technique in ovariectomized aged female rats (720-930 days of age). The mean numbers of axodendritic and axosomatic synapses in the ARCN decreased to about one-third of those in the intact controls. However, the treatment with estradiol benzoate (2 micrograms/day) during the 3 weeks following the day of brain surgery brought about a marked increase in the numbers of these synapses. The data suggest that the ARCN neurons of aged female rats still retain plasticity to react to deafferentation under influences of estrogen.     

Three-dimensional two-layer collagen matrix gel culture model for evaluating complex biological functions of monocyte-derived dendritic cells

Dendritic cell-like cells (Mo-DCs) generated from peripheral blood monocytes with interleukin-4 (IL-4) and granulocyte-macrophage colony-stimulating factor (GM-CSF) have been used as tools to treat cancer patients (DC-vaccines). Because Mo-DCs have multiple antigen presentation-related functions, including phagocytosis, migration, cytokine production, and T cell stimulation, establishment of a method for simultaneously evaluating the various functions of Mo-DCs is important. We developed a new in vitro three-dimensional two-layer collagen matrix culture model that consists of a collagen gel containing Mo-DCs as the lower layer and a collagen gel containing necrotic GCTM-1 tumor cells and/or T cells as the upper layer. We used this system to observe simultaneously multiple functions of Mo-DCs by phase-contrast or fluorescence microscopy and to assess IL-12 secretion during more than 2 weeks of culture. We also observed interactions between Mo-DCs and necrotic GCTM-1 or T cells on an individual cell basis by time-lapse videomicroscopy. In addition, we collected Mo-DCs from the collagen gels by collagenase treatment and analyzed the expression of antigen presentation-related molecules such as HLA-DR, CD80, CD83, and CD86 on Mo-DCs. This model may be a useful tool for evaluation of the various functions of Mo-DCs used as DC vaccines and for studies of the complex behaviors of Mo-DCs in vivo.     

Cytomegalovirus infections following umbilical cord blood transplantation using reduced intensity conditioning regimens for adult patients.

Cytomegalovirus (CMV) infection is a major complication after allogeneic hematopoietic stem cell transplantation (Allo-HSCT); however, we have little information on the clinical features of CMV reactivation after cord blood transplantation using reduced-intensity regimens (RI-CBT) for adults. We reviewed medical records of 140 patients who underwent RI-CBT at Toranomon Hospital between January 2002 and March 2005. All the patients were monitored for CMV-antigenemia weekly, and, if turned positive, received preemptive foscarnet or ganciclovir. Seventy-seven patients developed positive antigenemia at a median onset of day 35 (range, 4-92) after transplant. Median of the maximal number of CMV pp65-positive cells per 50,000 cells was 22 (range, 1-1806). CMV disease developed in 22 patients on a median of day 35 (range, 15-106); 21 had enterocolitis and 1 had adrenalitis. CMV antigenemia had not been detected in 2 patients, when CMV disease was diagnosed. CMV disease was successfully treated using ganciclovir or foscarnet in 14 patients. The other 8 patients died without improvement of CMV disease. In multivariate analysis, grade II-IV acute graft-versus-host disease was a risk factor of CMV disease (relative risk 3.48, 95% confidential interval 1.47-8.23). CMV reactivation and disease develop early after RI-CBT. CMV enterocolitis may be a common complication after RI-CBT.     

An improved method for the detection of HIV antigen in the blood of carriers

The measurement of HIV antigen levels in sera or plasma of HIV-infected individuals is critical for determining the existence of antigen or infectious virus before seroconversion and for prognosis. Pretreatment of sera or plasma of HIV carriers by heating at 70 degrees C for 10 min at an acidic pH enabled us to estimate antigens efficiently in immune complexes. This procedure will also be useful in determining antigen levels in HIV carriers more precisely.     

Protective effect of human granulocyte colony-stimulating factor on microbial infection in neutropenic mice.

A purified human granulocyte colony-stimulating factor (hG-CSF) was studied for its protective effect on the induction of neutropenia and enhanced susceptibility to microbial infections in mice receiving cyclophosphamide (CPA). A severe reduction in peripheral blood neutrophils was induced 4 days after injection with 200 mg of CPA per kg although the level normalized rapidly thereafter. When mice were injected subcutaneously once a day with 2.5 micrograms of hG-CSF beginning on the day after CPA injection, the reduction was prevented markedly, even 4 days later. On the other hand, in mice receiving CPA 4 days prior to infection, a weakened resistance to intraperitoneal challenge with a strain of Pseudomonas aeruginosa was induced. This weakened resistance was dose-dependently restored to normal by four daily injections with hG-CSF. A daily dose of 1.0 microgram was required for complete restoration, although hG-CSF did not directly inhibit bacterial growth in vitro. In hG-CSF-treated mice, morphologically mature neutrophils migrated rapidly into the peritoneal cavities where bacteria were inoculated, followed by a rapid elimination of bacteria from the locality as compared with controls. In addition, the same treatment with hG-CSF was able to protect significantly against systemic infections caused by Serratia marcescens, Escherichia coli, Staphylococcus aureus, and Candida albicans. These data show the possibility that prophylactic therapy with hG-CSF may augment the resistance of immunocompromised patients to infections.     

Nucleotide mutations associated with hepatitis B e antigen negativity

One hundred and forty four patients with chronic hepatitis B were tested to identify new mutations associated with hepatitis B e antigen (HBeAg) negativity, using a full genome sequence analysis. All the patients were Chinese and had hepatitis B virus infection of genotype C. Patients with none of the pre-core or core promoter mutations were significantly (P < 0.001) less common in the group with anti-HBe (13%) than in the group with HBeAg (56%). The complete nucleotide sequence was determined in four anti-HBe-positive patients who had neither pre-core nor core promoter mutations and in five HBeAg-positive patients who also had neither of these mutations (the groups were matched for age and sex). Six mutations were found to be significantly more common in the former group than in the latter: G529A (3/4 vs. 0/5), C934A (4/4 vs. 1/5), A1053G (4/4 vs. 1/5), G1915T/A (4/4 vs. 0/5), T2005C/A (4/4 vs. 0/5), and C3026T (3/4 vs. 0/5). Three of the six mutations were significantly more common in the four anti-HBe-positive patients who had neither pre-core nor core promoter mutations, compared to 11 HBeAg-positive patients who had pre-core and core promoter mutations, and also compared to 15 anti-HBe-positive patients who had pre-core and core promoter mutations, suggesting further the specificity of these mutations. Of the six mutations, two resulted in amino acid substitution in the polymerase protein, and one is located near the enhancer I region. The results suggest that the six newly discovered mutations are associated with HBeAg negativity.     

Experimental induction of uterine cancer in rats by N-ethyl-N'-nitro-N-nitrosoguanidine dissolved in polyethylene glycol

In the present experiment we attempted to experimentally induce uterine cancer in rats by injecting into the uterine cavity N-ethyl-N'-nitro-N-nitrosoguanidine (ENNG) dissolved in polyethylene glycol (PEG). Fifty-nine female F-344 rats, 7-8 weeks old, were divided into three groups and each received in the left uterine cavity with laparotomy a single dose of ENNG dissolved in PEG according to the following schedule: Group 1 received 75 mg ENNG/kg body wt.; Group 2 had 20 mg ENNG/kg body wt.: and Group 3 was given only PEG. In Group 1 it was observed that adenocarcinoma and sarcoma were present in the uterine corpus while squamous cell carcinoma occurred in the uterine cervix. In Group 2, although tumors such as adenocarcinoma, adenoma and sarcoma were observed in the uterine corpus, no tumor was present in the uterine cervix. No tumor growth whatsoever was observed in Group 3. From the above results it is apparent that the present method is an efficient means for experimentally inducing uterine cancer and that the site of tumor generation varies according to the concentration of ENNG administered.     

Three Kinds of Foamy Cells in the Spleen: Comparative Histochemical and Ultrastructural Studies

By light and electron microscopy, we observed foamy cells in the spleens from a patient with hemolytic anemia due to red cell adenosine deaminase (ADA) overproduction, a patient with rheumatoid arthritis (RA) treated with gold, and patients with idiopathic thrombocytopenic purpura (ITP)

The foamy cells associated with red cell ADA overproduction were essentially similar to Gaucher-like cells described in patients with thalassemia, and it was suggested that the accelerated destruction of red cells was one of the factors responsible for the development of foamy cells. Foamy cells in ITP and RA were closely associated with an increased destruction of platelets in the spleen. Morphologic transitions between phagocytosed platelets and myelinlike materials were traced in these disorders. In RA, however, foamy cells were heterogeneous from an ultrastructural standpoint, with different cytoplasmic inclusions. In addition to myelinlike materials, dense bodies, vacuoles with flocculent materials, and gold were noted in most of foamy cells. As gold compounds are known to inhibit lysosomal enzymes, we surmise that an acquired disturbance in lysosomal digestion is partially responsible for the accumulation of intermediate metabolites.

In the pathogenesis of foamy cells associated with blood cell dyscrasia, the accelerated destruction of blood cells and/or acquired disorders in catabolic pathways within the macrophages are suggested to be the underlying mechanism of an intralysosomal accumulation of incompletely degraded cellular debris.     

A low molecular weight inhibitor of the alternative complement pathway. I. Its isolation from human urine and the reaction mechanism.

A low molecular inhibitor (LMW-INH) of the alternative pathway activation was isolated form healthy human urine. Its molecular weight was slightly higher than 1000. LMW-INH inhibited C3 convertase formation in fluid phase, on sheep erythrocytes and on zymosan particles. In contrast with beta 1H globulin LMW-INH showed no effect on the C3b binding site for B, and it inhibited the activation of CVF.B complex by D only when LMW-INH was simultaneously present with D. These results indicate that the reaction mechanism of LMW-INH is different from that of beta 1H globulin.