Microplate hybridization for Borna disease virus RNA in human peripheral blood mononuclear cells.

We developed a simple and sensitive microplate hybridization procedure with which to identify Borna disease virus cDNA in amplified products from human peripheral blood mononuclear cells. The mean values for the positive PCR products were significant compared with those for any of the negative products, indicating that this method can be applied to rapidly diagnose a large number of clinical specimens.     

Arteries to the great and second toes based on three-dimensional analysis of 100 cadaveric feet

Summary Three-dimensional arteriography was used to analyse the arterial supply of the great and second toes of 100 cadaveric feet down to the microsurgical level. This information will aid in successful composite tissue transfers of these toes to the hand. The arterial blood supply of the great toe came principally from the first dorsal metatarsal a. (78%) and the first plantar metatarsal a. (22%), and secondarily from the medial tarsal aa. and the three terminal branches of the medial plantar a. For the second toe, the first dorsal metatarsal a. (78%) and the first plantar metatarsal a. (22%) supplied blood from the medial side, and the second dorsal metatarsal a. (78%) and the second plantar metatarsal a. (22%) supplied blood from the lateral side. Seven arterial patterns were found in the interdigital web space. The socalled general pattern was seen in the first web space in 65% of the feet examined. In the second web space it was found in 85%. The first intermetatarsal space sometimes contained a large artery arising directly from the dorsalis pedis or first proximal perforating a. as well as the first dorsal and first plantar metatarsal arteries. In this space, arterial patterns were classified into 4 types and 9 subtypes based on the origins and proximal courses of these arteries. The so-called standard pattern was found in only 19% of the feet, while an arterial pattern with a common proximal trunk on the plantar side for the first dorsal and first plantar metatarsal aa. was found most frequently (46%). In the second intermetatarsal space the second dorsal metatarsal a. was usually located on the dorsum.

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Les artères de l'hallux et du deuxième orteil. Analyse tridimensionnelle de cent pieds de cadavres

Résumé L'artériographie en trois dimensions a été utilisée pour analyser la vascularisation artérielle de l'hallux et du deuxième orteil de cent pieds de cadavres jusqu'au niveau microchirurgical. Ces renseignements aideront au succès des transferts de tissus composites de ces orteils vers la main. La vascularisation artérielle de l'hallux vient principalement de la première artère métatarsienne dorsale (78 %) et de la première artère métatarsienne plantaire (22 %) et secondairement des artères tarsiennes médiales et des trois branches terminales de l'artère plantaire médiale. Pour le deuxième orteil, la première artère métatarsienne dorsale (78 %) et la première artère métatarsienne plantaire (22 %) vascularisent le versant médial, la deuxième artère métatarsienne dorsale (78 %) et la deuxième artère métatarsienne plantaire (22 %) vascularisent le versant latéral. Sept dispositions artérielles ont été trouvées dans la commissure interdigitale. Une disposition dénommée commune a été vue dans la premiere commissure sur 65 % des pieds examinés, dans la deuxième commissure dans 85 % des cas. Le premier espace intermétatarsien contient parfois une volumineuse artère, naissant directement de l'artère dorsale du pied ou de la première artère perforante proximale, mais aussi des premières artères métatarsiennes dorsale ou plantaire. Dans cet espace, les dispositions artérielles sont classées en 4 types et 9 sous-types, basés sur l'origine et le trajet proximal de ces artères. La disposition standard a été trouvée seulement sur 19 % des pieds, alors qu'une disposition comprenant un tronc proximal commun situé sur le versant plantaire, fournissant les premières artères métatarsiennes dorsale et plantaire, a été trouvée plus fréquemment (46 %). Dans le deuxième espace intermétatarsien, la deuxième artère métatarsienne dorsale était habituellement située sur le versant dorsal.

     

Monoclonal antibodies to a CD4 peptide derivative which includes the region corresponding to an immunoglobulin CDR3: evidence of the involvement of pre-CDR3-related region in HIV-1 and host cell interaction.

A CD4 peptide of amino acid residues 68-130 [CD4(68-130)], which had the capacities to inhibit HIV-1 replication and HIV-1-induced syncytium formation, was used as an immunogen for the preparation of mAb. The mAbs prepared were classified into at least five types (I-V) in terms of their recognition sites by ELISA using various kinds of smaller CD4 peptides. Among them, the type I mAb no. 35 recognizing amino acid residues 72-84, which lies just before the region corresponding to an immunoglobulin third complementarity-determining region (CDR3), showed the strongest effects in reducing both HIV-1 infection and HIV-1-induced syncytium formation, although a large amount of no. 35 mAb was necessary to reduce such HIV-1 activities compared with those of anti-Leu-3a and OKT4A mAbs which recognize CD4 epitopes near a portion corresponding to an immunoglobulin CDR2. Western blot analysis showed that the reactivities of CD4 molecule in CD4-positive cells or sCD4 molecule with types I-V mAbs were stronger than that with anti-Leu-3a mAb. Flow cytometry showed that no. 35 mAb was faintly reactive with native CD4 molecule on cell surface at the concn showing the inhibitory effects on HIV-1 infection and syncytium formation. In addition, a smaller peptide CD4(66-92), one of the good epitope peptides for no. 35 mAb, also showed strong inhibitory effect on HIV-1 infection as well as a weaker inhibitory effect on syncytium formation. These results suggest that, in addition to the CD4 CDR2-related region, the pre-CDR3-related region is also involved in the early events of the interactions between the host cell and HIV-1.     

Virus-like particles in cultured C3H/St mouse cells treated with a carcinogenic polycyclic hydrocarbon

Virus-like particles with cylindrical form were found in cultured alveolar macrophages and lung fibroblasts of C3H/St mice, after treating these cells with a carcinogenic polycyclic hydrocarbon, 1, 2, 5, 6,-Dibenzanthracene (DBA). Their morphology looked identical to those observed in vivo in the reactive cells which engulfed DBA crystals implanted into the brain and muscle of the same mouse strain. These particles were not found in either the untreated cells of C3 H/St mice or the treated cells of BALB/c mice. In the alveolar macrophages, these particles appeared first at 3 days after DBA treatment and reached the maximum number around the 30th day. They still kept their morphology in the degenerating cells which had lost the cytoplasmic organelles. These findings suggest the possibility that DBA induced the expression of the viral genome endogeneous to C3H/St cell.     

Cleavage of integrin by mu-calpain during hypoxia in human endometrial cells

PROBLEM: The distribution and activation of mu-calpain and possible cleavage of integrin in human endometrial cells under hypoxic condition were investigated. METHOD OF STUDY: Human endometrial epithelial and stromal cells were subjected to hypoxia, and subsequently used for immunostaining and western blot analysis. RESULTS: The proform of mu-calpain was detected in the cytoplasm of normal cells, and displayed a substantial decrease after hypoxia. Conversely, the active form of mu-calpain was not detected in normal cells, but was abundant after hypoxia. The cytoplasmic domain of integrin beta3 was also detected in the cytoplasm of endometrial cells. Western blot analysis confirmed that both the proform of mu-calpain and the integrin beta3 cytoplasmic domain decreased during hypoxia. CONCLUSIONS: Mu-calpain is activated in human endometrial cells during hypoxia and that subsequent cleavage of the integrin beta3 cytoplasmic domain may give some adverse effects to the function of human endometrium.     

Demonstration of Borna disease virus (BDV) in specific regions of the brain from horses positive for serum antibodies to BDV but negative for BDV RNA in the blood and internal organs

Sero- and molecular-epidemiological studies on Borna disease virus (BDV) infection show that BDV RNA is not always detected in the peripheral blood mononuclear cells (PBMCs) from serum anti-BDV antibody-positive individuals such as horses, sheep, cattle, cats, and humans. In this study we demonstrated BDV RNA signals by polymerase chain reaction only in restricted regions of the brain from horses with locomotor disease. Four of six horses examined showed apparently positive reactions for anti-BDV antibodies. Specific regions of the brain of these four horses were positive for BDV RNA but the internal organs, lymph nodes, and PBMCs were negative. Histological studies of their brains revealed no apparent histological abnormalities such as inflammatory reactions. These results suggest that BDV chronically infects certain restricted regions of brain in seropositive horses. Received: 6 January 1997     

Role of apoptosis controlled by cytochrome c released from mitochondria for luteal function in human granulosa cells

PROBLEM: The aim of this study was to investigate the relationship between apoptosis by the mitochondrial pathway and luteal function in human granulosa cells. METHOD OF STUDY: Granulosa cells were obtained by ultrasound-guided follicular aspiration from patients undergoing in vitro fertilization and embryo transfer. After the addition of RU486, cells were stained with a mitochondria-specific fluorescent dye, MitoTracker Red CM x Ros. Using flow cytometry and National Institute of Health image, the mitochondrial fluorescent area was measured. After staining with Hoechst 33258 dye, the number of apoptotic bodies per 1000 cells were counted at random on photomicrographs. Homogenates were used for sodium dodecyl sulphate-polyacrylamide gel electrophoresis and Western blot analysis using antibodies against cytochrome c or caspase-3. RESULTS: The incidence of apoptotic bodies increased and the mitochondrial membrane potential decreased time dependently. The opposite effect was observed dose dependently with RU486 treatment. Western blot analysis showed increased cytochrome c expression, after treatment with 1-2 microg/mL of RU486 which then decreased with 5-10 microg/mL of RU486. Caspase-3 expression increased dose dependently with RU486. CONCLUSIONS: These results suggest that the activation of caspase-3 caused by cytochrome c released from mitochondria plays an important role in apoptosis-related luteal function in human granulosa cells.     

Experimental study on secondary amyloidosis associated with chronic arthritis

Various organs from rabbits immunized with heat-killed E. coli 0:14 to induce chronic polyarthritis resembling rheumatoid arthritis (RA) were examined for amyloid deposition since amyloidosis is a frequent feature of long-standing RA. Amyloids were confirmed mainly by polarizing as well as nonpolarizing light microscopy after Congo red staining and partly by electron microscopy and immunoperoxidase technique. Amyloid deposits after an additional 2 months from the first appearance of arthritis were most frequently found in the spleen and kidneys suggesting secondary amyloidosis. Amyloidosis was observed in 36% of 75 arthritic rabbits. The incidence of amyloidosis was significantly higher in the arthritic rabbits than in the non-arthritic rabbits (p less than 0.05). This suggests that prolonged sensitization with heat-killed E. coli 0:14 containing endotoxic substance participates in the formation of both arthritis and amyloidosis.     

Differential susceptibility of resting CD4(+) T lymphocytes to a T-tropic and a macrophage (M)-tropic human immunodeficiency virus type 1 is associated with their surface expression of CD38 molecules

Recent evidence has accumulated which definitively shows that chemokine receptors CCR5 and CXCR4 play an essential role as coreceptors for human immunodeficiency virus type 1 (HIV-1) infection. Flow cytometric analysis permitted us to detect CD38, a surface marker of early differentiation, as well as activation of T cells, on about half of healthy donor-derived CD4(+) T cells. In this study, we focused on the susceptibility of CD38(+) and CD38(-) subsets of CD4(+) T cells to HIV-1 infection with different coreceptor tropisms. About 20% of peripheral blood mononuclear cell-derived resting CD4(+) T cells were recovered into the CD38(+) subset fraction by panning with a monoclonal antibody to CD38. Most of the cells in this CD38(high) fraction also expressed CD45RA and CD62L at higher intensities compared with those of CD38(low) fraction. CCR5(+) T cells predominated in the CD38(-) subset, although cell surface expression of CD4 and CXCR4 was almost similar between both subsets. This difference was consistent with a significantly higher susceptibility of the CD38(-) subset to a macrophage (M)-tropic HIV-1 strain. In contrast, it was shown that a T-tropic strain of HIV-1 could replicate more efficiently in the CD38(+) subset, although viral adsorption rates were similar between both subsets. Thus, the differential susceptibility of CD4(+) T cells to M(-) and T-tropic HIV-1 was associated with their surface expression of CD38.