Multiple Histiocytic Foam Cell Nodules in the Tongue of Miniature Dachshund Dogs.

Miniature dachshund dogs are a common breed in Japan and are known to be predisposed to granulomatous diseases. Here we report the pathologic features of multiple lingual nodules in 7 miniature dachshunds. Seven dogs had multiple nodules of variable sizes mainly on the ventral and lateral surface of the tongue. In addition, 1 dog also had masses on the left oral mucosa. Three cases had recurrence after surgical resection. Histologically, the lingual nodules were composed of aggregates of foam cells with clear vacuolated cytoplasm that were negative for oil red O, PAS, and alcian blue. They stained positively for CD204 (macrophage scavenger receptor) and MHC class II and negatively for Iba-1, E-cadherin, adipophilin, cytokeratins, S-100, and nestin. These findings indicate that the multiple lingual nodules in miniature dachshunds are an unusual, unique lesion consisting of macrophage-derived foam cells, which does not correspond to canine lingual diseases reported to date.

Hyaluronan metabolism in overloaded temporomandibular joint.

This study aimed to examine hyaluronan (HA) metabolism in relation to the onset and progression of temporomandibular joint osteoarthritis (TMJ-OA) induced by mechanical overloading. Two-month-old and 6-month-old C57BL/6N mice were divided into experimental and untreated control groups (n = 5/group). A sliding plate was attached to the maxillary incisors of the experimental mice for 10 days to overload the condylar cartilage in TMJ. In experimental group, profound cartilage degradation was detected in haematoxylin-eosin, Safranin-O-Fast Green-stained sections. It was also shown that the cartilage degradation was greater in older mice in both the control and the experimental groups. The number of HABP-positive cells was decreased by mechanical overloading and with age. The reduction of HA expression was correlated with the progression of cartilage degradation induced by mechanical overloading. The absolute quantification of the mRNA expression related to HA synthesis and HA degradation was also performed in each group. The mRNA expression levels of HA synthase (HAS) 2 and 3 were lower in the experimental group compared with the control group in the younger mice. In contrast, the mRNA expression levels of the HA degradation gene, HYAL2 and KIAA1199, were higher in the experimental group compared with the control group in the older mice. Thus, mechanical overload differently affected the balance of HA degradation and HA synthesis in the older and younger mice, respectively. In conclusion, mechanical overloading affects HA metabolism and it might initiate or amplify the condylar cartilage degradation.

Macrophage deficiency in osteopetrotic (op/op) mice inhibits activation of satellite cells and prevents hypertrophy in single soleus fibers.

Effects of macrophage on the responses of soleus fiber size to hind limb unloading and reloading were studied in osteopetrotic homozygous (op/op) mice with inactivated mutation of macrophage colony-stimulating factor (M-CSF) gene and in wild-type (+/+) and heterozygous (+/op) mice. The basal levels of mitotically active and quiescent satellite cell (-46 and -39% vs. +/+, and -40 and -30% vs. +/op) and myonuclear number (-29% vs. +/+ and -28% vs. +/op) in fibers of op/op mice were significantly less than controls. Fiber length and sarcomere number in op/op were also less than +/+ (-22%) and +/op (-21%) mice. Similar trend was noted in fiber cross-sectional area (CSA, -15% vs. +/+, P = 0.06, and -14% vs. +/op, P = 0.07). The sizes of myonuclear domain, cytoplasmic volume per myonucleus, were identical in all types of mice. The CSA, length, and the whole number of sarcomeres, myonuclei, and mitotically active and quiescent satellite cells, as well as myonuclear domain, in single muscle fibers were decreased after 10 days of unloading in all types of mice, although all of these parameters in +/+ and +/op mice were increased toward the control values after 10 days of reloading. However, none of these levels in op/op mice were recovered. Data suggest that M-CSF and/or macrophages are important to activate satellite cells, which cause increase of myonuclear number during fiber hypertrophy. However, it is unclear why their responses to general growth and reloading after unloading are different.

Immunophenotypical characterization of macrophages in rat bleomycin-induced scleroderma.

Scleroderma is a skin disorder characterized by persistent fibrosis. Macrophage properties influencing cutaneous fibrogenesis remain to be fully elucidated. In this rat (F344 rats) model of scleroderma, at 1, 2, 3, and 4 weeks after initiation of daily subcutaneous injections of bleomycin (BLM; 100 µl of 1 mg/ml daily), skin samples were collected for histological and immunohistochemical evaluations. Immunohistochemically, the numbers of cells reacting to ED1 (anti-CD68; phagocytic activity) and ED2 (anti-CD163; inflammatory factor production) began to increase at week 1, peaked at week 2, and decreased thereafter. In contrast, the increased number of cells reacting to OX6 (anti-MHC class II molecules) was seen from week 2 and remained elevated until week 4. α-Smooth muscle actin-positive myofibroblasts were increased for 4 weeks. Double labeling revealed that galectin-3, a regulator of fibrogenic factor TGF-ß1, was expressed in CD68+, CD163+, and MHC class II+ macrophages and myofibroblasts. mRNA expression of TGF-ß1, as well as MCP-1 and CSF-1 (both macrophage function modulators), were significantly elevated at weeks 1 to 4. This study shows that the increased number of macrophages with heterogeneous immunophenotypes, which might be induced by MCP-1 and CSF-1, could participate in the sclerotic lesion formation, presumably through increased fibrogenic factors such as galectin-3 and TGF-ß1; the data may provide useful information to understand the pathogenesis of the human scleroderma condition.

Effect of exercise training on the density of endothelial cells in the white adipose tissue of rats.

We examined the effects of a 9-week exercise training (TR) in Wistar male rats, beginning at 4 weeks of age, on the density of endothelial cells (ECs) in epididymal white adipose tissue (WAT) and the mRNA expression of angiogenic factors in adipose tissue stromal vascular fraction (SVF) cells. The number of ECs and mRNA expressions were assessed by lectin staining and real-time reverse transcriptase-polymerase chain reaction, respectively. Compared with control (CR) rats, TR rats gained weight more slowly and had significantly lower final weight of WAT due to the reduction in the size and the number of adipocytes. TR significantly increased the number of ECs per square millimeter and per adipocyte (1.37- and 1.23-fold, respectively) in WAT. This is probably because the number of adipocytes is fewer while the number of ECs is constant in the WAT of TR rats, because the regression line of TR rats for adipocyte number-dependent EC number was shifted toward the left without significant differences in the slopes between groups. TR also induced the upregulation of mRNA expression of vascular endothelial growth factor (Vegf)-A and Vegf-receptor-2 in SVF cells, thereby retaining a constant number of ECs in the WAT.

Cardiac hamartoma in a young squirrel monkey who died suddenly.

A case of cardiac hamartoma in a 2-month-old squirrel monkey is reported. The monkey showed a loss of appetite and died suddenly. Microscopically, an encapsulated nodular lesion was found at the right atrial wall. The lesion consisted of irregularly shaped, slender myocytes intermingled with a few fibroblasts and collagen fibers. Neither nuclear atypia nor inflammatory cell infiltrate was seen. The constituting cells had stratified striations in the cytoplasm and reacted immunohistochemically for desmin, indicating the nature of myocytes. Based on the above findings, a diagnosis of cardiac hamartoma was made. This is the first case of cardiac hamartoma in this species.

Biomechanical and biochemical characteristics of the mandibular condylar cartilage.

The human masticatory system consists of a mandible which is able to move with respect to the skull at its bilateral temporomandibular joint (TMJ) through contractions of the masticatory muscles. Like other synovial joints, the TMJ is loaded mechanically during function. The articular surface of the mandibular condyle is covered with cartilage that is composed mainly of collagen fibers and proteoglycans. This construction results in a viscoelastic response to loading and enables the cartilage to play an important role as a stress absorber during function. To understand its mechanical functions properly, and to assess its limitations, detailed information about the viscoelastic behavior of the mandibular condylar cartilage is required. The purpose of this paper is to review the fundamental concepts of the biomechanical behavior of the mandibular condylar cartilage. This review consists of four parts. Part 1 is a brief introduction of the structure and function of the mandibular condylar cartilage. In Part 2, the biochemical composition of the mandibular condylar cartilage is summarized. Part 3 explores the biomechanical properties of the mandibular condylar cartilage. Finally, Part 4 relates this behavior to the breakdown mechanism of the mandibular condylar cartilage which is associated with the progression of osteoarthritis in the TMJ.

Malignant seminoma with multiple visceral metastases in a guinea fowl (Numida meleagris) kept in a zoo.

A case of seminoma found in an adult guinea fowl (Numida meleagris) that has been exhibited in a zoo is reported. The right testis was extremely enlarged and replaced by round and polyhedral pleomorphic neoplastic cells showing nest, sheet, and diffuse growth patterns. The neoplastic cells had acidophilic cytoplasm and hyperchromatic and eccentrically placed nuclei. Metastatic lesions composed of diffuse growth of neoplastic cells similar to those of the primary tumor were seen in the liver, lungs, kidneys, and heart, and neoplastic emboli were often detected within blood vessels of these organs, indicating hematogenous metastasis. This is the first report of malignant seminoma with multiple metastases in the visceral organs in the guinea fowl.

A Case of Feline T-cell Lymphoma with Tropism for Striated Muscle and Peripheral Nerve.

An 11-year-old female American shorthair cat was presented with a 3-month history of hindlimb ataxia and knuckling of the left forelimb. Clinical abnormalities included weight loss, hyperaesthesia of the neck and back, cardiac murmur and systemic muscle atrophy. The cat died 10 days after the initial presentation and a necropsy examination was performed. Grossly, extensive pale lesions were seen in the wall of the left ventricle and the septum of the heart. There were no detectable masses in the heart, skeletal muscles or peripheral nerves. Histopathological examination revealed diffuse, extensive infiltration of atypical lymphoid cells in the heart; the cardiac muscles were markedly degenerate and atrophic and were replaced by the neoplastic cells. Neoplastic cells with similar morphology were seen in all specimens of the skeletal muscles and peripheral nerves. Clonality analysis of the paraffin wax-embedded heart tissue revealed a monoclonal rearrangement of the gene encoding the T-cell receptor γ chain. Based on these findings, the case was diagnosed as T-cell lymphoma with tropism for striated muscle and peripheral nerve.

Regional differences in fiber characteristics in the rat temporalis muscle.

The behavioral differences in muscle use are related to the fiber type composition of the muscles among other variables. The aim of this study was to examine the degree of heterogeneity in the fiber type composition in the rat temporalis muscle. The temporalis muscle was taken from 10-week-old Wistar strain male rats (n = 5). Fiber types were classified by immunohistochemical staining according to their myosin heavy chain content. The anterior temporalis revealed an obvious regional difference of the fiber type distribution, whereas the posterior temporalis was homogeneous. The deep anterior temporalis showed a predominant proportion of type IIA fibers and was the only muscle portion displaying slow type fibers (< 10%). The other two muscle portions, the superficial anterior and posterior temporalis, did not differ significantly from each other and contained mainly type IIB fibers. Moreover, the deep anterior temporalis was the only muscle portion showing slow type fibers (< 10%). In the deep portion, type IIX fibers revealed the largest cross-sectional area (1943.1 +/- 613.7 microm(2)), which was significantly (P < 0.01) larger than those of type IIA and I + IIA fibers. The cross-sectional area of type IIB fibers was the largest in the remaining two muscle portions and was significantly (P < 0.01) larger than that of type IIX fibers. In conclusion, temporalis muscle in rats showed an obvious heterogeneity of fiber type composition and fiber cross-sectional area, which suggests multiple functions of this muscle.

Participation of Tumor-Associated Myeloid Cells in Progression of Amelanotic Melanoma (RMM Tumor Line) in F344 Rats, with Particular Reference to MHC Class II- and CD163-Expressing Cells.

Tumor progression is often influenced by infiltration of myeloid cells; depending on the M1- or M2-like activation status, these cells may have either inhibitory or promoting effects on tumor growth. We investigated the properties of tumor-associated myeloid cells in a previously established homotransplantable amelanotic melanoma (RMM tumor line) in F344 rats. RMM tumor nodules were allowed to reach the sizes of 0.5, 1, 2 and 3 cm, respectively. Immunohistochemistry and flow cytometry was performed for macrophage markers CD68 and CD163, and for the antigen-presenting cell marker, MHC class II. Although no significant change was observed in the number of CD68+ and CD163+ macrophages during RMM progression, the number of MHC class II+ antigen-presenting cells was reduced in 3 cm nodules. Real-time RT-PCR of laser microdissection samples obtained from RMM regions rich in MHC class II+ cells demonstrated high expressions of M1-like factors: IFN-γ, GM-CSF and IL-12a. Furthermore, fluorescence-activated cell sorting, followed by real-time RT-PCR for CD11b+ MHC class II+ (myeloid antigen-presenting cells), CD11b+ CD163+ (M2 type myeloid cells), CD11b+ CD80+ (M1 type myeloid cells) and CD11b+ CD11c+ (dendritic cells) cells was performed. Based on the levels of inflammation- and tumor progression-related factors, MHC class II+ antigen-presenting cells showed polarization towards M1, while CD163+ macrophages, towards M2. CD80+ and CD11c+ myeloid cells did not show clear functional polarization. Our results provide novel information on tumor-associated myeloid cells in amelanotic melanoma, and may become useful in further research on melanoma immunity.

Cutaneous Histiocytic Sarcoma with Regional Lymph Node Metastasis in a Netherland Dwarf Rabbit (Oryctolagus cuniculus).

A 10-year-old male Netherland dwarf rabbit (Oryctolagus cuniculus) was presented with a red nodular mass (1 cm in diameter) with ulceration and hair loss in the skin of the left upper lip. Cytological examination revealed atypical round cells. The mass was excised surgically. Histologically, the mass was composed of large round to polyhedral neoplastic cells with marked cytological atypia. The neoplastic cells were often binucleated or multinucleated. Immunohistochemically, the neoplastic cells were intensely positive for Iba1 and vimentin, but fewer neoplastic cells expressed E-cadherin. Nuclear immunoreactivity for Ki67 was detected in approximately 41% of the neoplastic cells. Metastasis to the left cervical lymph nodes was detected 6 months after the surgical excision. Based on clinical, histopathological and immunohistochemical findings, the present case was diagnosed as cutaneous histiocytic sarcoma. To the authors' knowledge cutaneous histiocytic disease has not been reported previously in lagomorphs.

Metastatic Liposarcoma in a South African Fur Seal (Arctocephalus pusillus).

A 14-year-old female South African fur seal (Arctocephalus pusillus) was presented with a large skin mass on the right shoulder. At necropsy examination, multiple white nodules were found in the lungs, liver, spleen and right axillary lymph nodes. Histologically, the skin mass was composed of round to polygonal neoplastic cells with round to oval nuclei and variably sized cytoplasmic vacuoles. Cellular and nuclear atypia were prominent. Immunohistochemically, the neoplastic cells expressed vimentin, but not cytokeratins, S100 protein, adipophilin or desmin. The cytoplasmic lipid droplets stained positively with oil red O. Metastasis was seen in the lungs, liver, spleen and right axillary lymph nodes, with similar morphological features to the skin mass. Based on these findings, a diagnosis of a pleomorphic liposarcoma with systemic metastasis was made. No previous reports of metastatic liposarcomas have been published in marine mammals.

Plant bZIP protein DNA binding specificity.

Plant bZIP proteins exhibit a relaxed DNA-binding specificity for DNA sequence motifs containing an ACGT core. Gel mobility shift experiments employing ten different recombinant plant bZIP proteins demonstrated that nucleotides flanking the ACGT core affected binding specificity and identified three different types of ACGT elements: G-box, CACGTG; C-box, GACGTC; and A-box, TACGTA, motifs. These ten different bZIP proteins could be categorized into three groups according to their qualitative and quantitative specificity for G-box and C-box elements. Dissociation constant values (Kd values) of these bZIP proteins for high affinity G-box and C-box elements and reciprocal competition gel mobility shift assays confirmed our classification scheme. Group 1 proteins exhibit a stronger binding affinity for G-box elements, group 2 proteins bind both G-box and C-box motifs with comparable binding affinity, whereas the group 3 proteins display a stronger binding affinity for C-box oligonucleotides. Studies using a panel of G-box and C-box oligonucleotides differing in their flanking sequences identified high affinity binding sites. All ten plant bZIP proteins examined, except TGA1a, exhibited type A G-box binding activity preferring class I G-box elements. In contrast to the situation observed for G-box elements, C-box motifs displayed a very much more stringent flanking nucleotide requirement for binding activity. Protein/DNA binding experiments using scanning mutants of a high affinity G-box element and G-box/C-box hybrid elements demonstrated that bZIP protein binding activity depends upon the affinity of protein dimer subunits for ACGT half-sites. Information provided by our systematic analysis of plant bZIP DNA binding specificity can be used to identify high affinity binding sites for the plant bZIP proteins studied here. Assuming that only high affinity bZIP binding sites are likely to function in vivo, identification of these sites will allow us to predict which genes are activated by a particular bZIP protein.

Glucocorticoid-mediated generation of suppressor macrophages with high density Fc gamma RII during acute cold stress.

Acute cold stress induces adherent cells with suppressor function, resulting in immunosuppression. Glucocorticoids (GC) are known as an inhibitor of the immune system and are increased by stress through stimulation of the hypothalamic-pituitary-adrenal axis. To elucidate the mechanisms underlying acute cold stress-induced immunosuppression, functions and surface phenotypes of murine peritoneal cells of the monocyte/macrophage lineage from acute cold-stressed mice (5 C for 3 or 24 h) in addition to the role of GC in the immunomodulation were investigated. Flow cytometric analysis revealed that the proportion of MAC-1+ cells with a high density of Fc gamma RII (Fc gamma RIIbright cells) was markedly increased in the peritoneal exudate cells from acute cold-stressed mice. These Fc gamma RIIbright cells were also stained with F4/80, a monoclonal antibody directed specifically against the mouse mature macrophages. The prominent suppressor activity for Concanavalin A (Con A) responses of control spleen cells was found in Fc gamma RIIbright cells, whereas MAC-1+ cells, with a low density of Fc gamma RII (Fc gamma RIIdull cells), from the stressed mice did not suppress the Con A responses. Fc gamma RIIbright cells from control mice also suppressed the Con A responses; the inhibitory effect was considerably less than that of cells from acute cold-stressed mice. As was anticipated, serum corticosterone levels were markedly increased in acute cold-stressed mice. In addition, expression of GC receptor messenger RNA was observed in Fc gamma RIIbright cells from these mice. The increase in Fc gamma RIIbright cells in peritoneal exudate cells caused by acute cold stress was inhibited by adrenalectomy or administration of a saturating amount of the GC antagonist RU 38486 (mifepristone). On the contrary, administration of the GC agonist, dexamethasone, markedly increased the proportion of Fc gamma RIIbright cells in peritoneal exudate cells of control mice. These results suggest that the generation of Fc gamma RIIbright suppressor cells of monocyte/macrophage lineage by acute cold stress was mediated to a greater or lesser degree by the action of GC through the GC receptor.

Application of a live varicella vaccine in children with acute leukemia or other malignant diseases.

A live varicella vaccine was used in 11 susceptible children in remission from acute leukemia, ten of whom had been in remission for six months or less, and in 6 children with neuroblastoma and retinoblastoma. In the immunological checkup before vaccination, most of them showed a positive reaction in the skin tests with dinitrochlorobenzene, phytohemagglutinin, purified protein derivative, and viral antigens. Leukopenia (three cases, less than 3,000/cu mm) and decreased IgG level (two cases, 380 mg/dl and 445 mg/dl) were observed in the children with leukemia. Anticancer medication was suspended from one week before vaccination to one week after vaccination. The only clinical reaction was a minute rash that appeared three weeks after vaccination in two children with leukemia and that disappeared within three days. Serological responses by complement fixing and neutralizing (NT) tests were detected in all the vaccinated children four weeks after vaccination, and NT antibody was still detected 28 months after vaccination in the two patients tested. Three of the vaccines were exposed to natural varicella at home and in the classroom 2 to 18 months after vaccination, but they were free from any varicella symptoms.

The plasmin heavy chain-urokinase conjugate: a specific thrombolytic agent.

Low molecular weight urokinase (LMW-UK) was coupled to the heavy chain of plasmin to make it able to bind to fibrin. The purified conjugate (PHC-UK conjugate), which consisted of equimolar concentrations of each starting material had a molecular weight of 93,600, bound tightly to fibrin-monomer-Sepharose and was not washed off with 1 M NaCl, but was eluted specifically with epsilon-amino caproic acid. The conjugate showed higher fibrinolytic activity than HMW-UK. A control conjugate prepared by coupling human serum albumin to LMW-UK (HSA-UK conjugate) showed the same fibrinolytic activity as HMW-UK. The half-lives of these two conjugates in rabbits were about 3 times that of HMW-UK. In an experimental pulmonary embolism model in rabbits, the PHC-UK conjugate showed about 10 times higher thrombolytic activity than HMW-UK, while the HSA-UK conjugate showed similar thrombolytic activity as HMW-UK, and moreover caused severe systemic fibrinogen breakdown. Thus the significant increase in thrombolytic activity after injection of PHC-UK conjugate into rabbits may be due to its newly acquired fibrin binding activity, and not to increase in its half-life. It is concluded that the PHC-UK conjugate may be useful in treatment of thrombosis.

Differential vasodilator properties of KRN2391, cromakalim, nitroglycerin and nifedipine in rabbit isolated femoral artery and vein.

1. The selectivity for artery and vein of KRN2391, cromakalim, nitroglycerin and nifedipine was examined in isolated femoral artery and vein preparations of the rabbit. 2. All drugs produced a concentration-dependent relaxation in both femoral artery and vein. 3. Nitroglycerin was more potent in femoral vein than in femoral artery at all concentrations. The EC50 value obtained in the vein was about 14 times smaller than that obtained in artery. 4. Cromakalim and nifedipine were almost equipotent on both vascular preparations. Cromakalim at the highest concentration (10(-5) M) produced 88 and 78% relaxation in femoral artery and vein, respectively. The maximum relaxation induced by nifedipine (10(-6) M) was less than 50% in both preparations. 5. KRN2391 was active at a lower concentration in the vein than in the artery and its maximum relaxation at 10(-5) M was about 90% in both preparations. 6. Glibenclamide (10(-6) M) inhibited the vasorelaxation caused by KRN2391 in both artery and vein. Methylene blue (10(-5) M) also inhibited the relaxant action of KRN2391 but this action was slight in the artery. 7. These results suggest that KRN2391 and nitroglycerin are more potent in the vein than in the artery and cromakalim and nifedipine are equipotent in both. It is considered that the relaxation induced by low concentrations of KRN2391 reflects predominantly its action as a nitrate and that at high concentrations it acts as a K+ channel opener in addition to its nitrate action. The different vascular selectivities of these drugs are thought to relate to the differences in their mechanisms of action in vascular smooth muscle.