Molecular evaluation of a new highly sensitive aryl hydrocarbon receptor in ostriches.

A 1,000-fold difference has been reported in dioxin sensitivity between avian species. This difference is because the 2 amino acids in the type 1 aryl hydrocarbon receptor (AhR1), at positions 325 and 381, correspond to Leu324 and Ser380 in chickens. The chicken had been reported to be the only avian species to possess a sensitive form of AhR1. This is the first study to reveal that the ostrich (Struthio camelus), a nonchicken species, also has a pair of amino acids (Ile-325 and Ser-381) that show high ligand affinity. However, the alignment of the AhR1 cDNA sequence showed that the AhR sequence in the ostrich was different than that of other avian species even though the critical amino acids were observed at positions 325 and 381. Ostrich AhR1 was also evaluated in a heterologous expression study. Ostrich AhR1 showed very high transcriptional activity of the cytochrome P450 1A5 (CYP1A5) gene in African Green Monkey Cercopithecus aethiops kidney cells (COS-7) treated with Sudan III. In primary cultures of ostrich kidney cells, CYP1A5 expression was induced by Sudan III at a lower (or almost identical) concentration to that observed in the chicken. The present study revealed a new AhR ligand sensitive avian species (i.e., the ostrich).

Activation of microglia and p38 mitogen-activated protein kinase in the dorsal column nucleus contributes to tactile allodynia following peripheral nerve injury.

The activation of glial cells in the CNS has been suggested to be involved in abnormal pain sensation after peripheral nerve injury. Previous studies demonstrated phosphorylation of p38 mitogen-activated protein kinase (MAPK) in spinal cord glial cells after peripheral nerve injury, and such phosphorylation has been suggested to be involved in the development of neuropathic pain. The aim of this study was to examine the dorsal column nuclei for phosphorylation of p38 MAPK following peripheral nerve injury and to explore a possibility of its contribution to neuropathic pain. Immunohistochemical labeling for phosphorylated p38 (p-p38) MAPK was performed in histological sections of the rat spinal cord and medulla oblongata after the fifth lumbar (L5) spinal nerve ligation (SNL). The number of p-p38 MAPK-immunoreactive (IR) cells was significantly increased in the L5 dorsal horn and the gracile nucleus ipsilateral to the injury at days 3-21 after SNL. Double immunofluorescence labeling with cell-specific markers revealed that p-p38 MAPK-IR cells co-expressed OX-42, suggesting their microglial identity. Increased immunofluorescence labeling for OX-42 indicated that microglial cells were activated by SNL in the L5 dorsal horn and the gracile nucleus ipsilateral to the injury. Continuous infusion of a p38 MAPK inhibitor into the cisterna magna for 14 days beginning on the day of SNL suppressed the development of tactile allodynia, but not thermal hyperalgesia induced by nerve injury. These results demonstrate that SNL activates p38 MAPK pathway in microglia in the gracile nucleus as well as in the spinal cord dorsal horn. Activation of p38 MAPK in medullary microglia may contribute to the pathogenesis of neuropathic pain.

A synthetic peptide inhibitor for alpha-chemokines inhibits the growth of melanoma cell lines.

Melanoma growth stimulatory activity (MGSA/GROalpha) is a 73 amino acid peptide sharing sequence characteristics with the alpha-chemokine superfamily. MGSA/GROalpha is produced by diverse melanoma cell lines and reported to act as an autocrine growth factor for the cells. We tested the binding of MGSA/GROalpha to melanoma cell lines, Hs 294T and RPMI7951, and found that these cells could bind to MGSA/GROalpha but not to interleukin-8. Recently, we defined a novel hexapeptide, antileukinate, which is a potent inhibitor of binding of alpha-chemokines to their receptors on neutrophils. When antileukinate was added to melanoma cells, it inhibited the binding of MGSA/ GROalpha. The growth of cells from both melanoma cell lines was suppressed completely in the presence of 100 microM peptide. The cell growth inhibition was reversed by the removal of the peptide from the culture media or by the addition of the excess amount of MGSA/GROalpha. The viability of Hs 294T cells in the presence of 100 microM peptide was > 92%. These findings suggest that MGSA/GROalpha is an essential autostimulatory growth factor for melanoma cells and antileukinate inhibits their growth by preventing MGSA/GROalpha from binding to its receptors.

Human standing posture control system depending on adopted strategies.

Control of the standing posture of humans involves at least two distinct modes of operation to restore the body balance in the sagittal plane: the ankle strategy and the hip strategy. The objective of the study was to estimate the contribution of vestibular, visual and somatosensory feedbacks to these distinct strategies. The body dynamics was described as the motion of two linked rigid segments that represented the legs and the rest of the body. The posture controller received the inclination angles of the two body segments as inputs and regulated the moments around the ankle and hip joints. The controller had four feedback paths that were characterised by transfer functions connecting the two inputs and the two outputs. To evoke the distinct strategies, the floor conditions were varied by narrowing the support surface under the feet. A continuous pseudo-random external disturbing force was applied to the waist and the thigh independently. The inclination angles of the body segments and the ground reaction force were measured, and the transfer functions of the controller were estimated with the maximum-likelihood system identification procedure. Six healthy male adult subjects participated in the experiment. When the hip strategy became evident under the narrow support surface conditions, the transfer function relating the leg inclination angle and the ankle joint moment decreased its DC gain (16%), whereas the other three transfer functions increased the gains (20-140%) (ANOVA, p < 0.05). Based on a criterion for simplicity in the modification of the posture controller, these changes suggest a new hypothesis that, when posture control becomes difficult, the central nervous system selectively activates the somatosensory feedback paths from the hip joint angle to the moments around the ankle and hip joints.

Constipation is not associated with colonic diverticula: a multicenter study in Japan.

BACKGROUND: The association of diverticula with bowel habits is unclear. We therefore analyzed the association between diverticula and bowel habits in over 1000 Japanese individuals. METHODS: Japanese subjects who underwent total colonoscopies at seven centers in Japan from June to September 2013 were analyzed. Bowel habits were evaluated using the Gastrointestinal Symptom Rating Scale, and stool form was assessed using a part of the Bristol Scale and Rome ΙΙΙ criteria. Diverticula were diagnosed by colonoscopy with a transparent soft-short hood. KEY RESULTS: The study evaluated 1066 subjects, 648 males and 418 females (ratio, 1.55 : 1), of mean age 63.9 ± 13.0 years. After adjusting for age and sex, the presence of constipation was associated with a significantly reduced likelihood of diverticula (odds ratio [OR] = 0.70, 95% confidence interval [CI] 0.52-0.93). When assessed according to the location of diverticula, the presence of constipation was associated with a significantly decreased likelihood of left-sided (OR = 0.39, 95% CI 0.16-0.93), but not right-sided (OR = 1.10, 95% CI 0.48-2.53), diverticula. Furthermore, stool form was unrelated with the presence or absence of diverticula. CONCLUSIONS & INFERENCES: The wide-spread hypothesis that constipation was associated with colonic diverticula was not supported. Rather, we found that the absence of diverticula was associated with constipation, suggesting the need to reassess the etiology of colonic diverticula.

Expression of protein kinase C subspecies in rat retina.

An extract of rat retina was subjected to Mono Q followed by chromatography on hydroxyapatite, and the protein kinase C (PKC) subspecies were identified by immunoblot and biochemical analysis. It was found that, although the relative activities assayed with myelin basic protein as a common phosphate acceptor vary greatly with one another, the alpha-, beta I-, beta II-, gamma-, delta-, epsilon-, zeta-, and another structurally unknown PKC subspecies are expressed in this tissue. Thus, the retina is a unique tissue which expresses most of the PKC subspecies so far identified in mammals.

A novel textured surface for blood-contact.

Blood-contacting surface modifications aimed at reduction of thromboembolic complications have included the texturing of surfaces so as to promote the formation of a stable pseudo-neointima. A technique has been developed whereby a textured surface consisting of regularly spaced micro-fibres was produced on a smooth base plane. Polyurethane vascular patches with and without the textured luminal surface were fabricated and implanted bilaterally in ovine carotid arteries for 1- and 3-week implantation periods (n = 6 per period). One of 6 arteries with textured patches in the 1-week group was occluded. All other arteries were patent. At 1 week, all patent textured patches had adherent thrombus covering the entire patch surface. By 3 weeks, the thrombus had organised to form a stable pseudo-neointima. Non-textured patches at 1 week had only partial surface coverage of thrombus. At 3 weeks, 4 of 6 non-textured patches had significant red thrombus in the lumen. At 3 weeks, there was also evidence of cellular migration from artery onto both textured and non-textured patches. These findings suggest that the major role of the textured surface was as a promoter of a stabilised thrombus base onto which subsequent cellular migration and tissue healing occurred more rapidly than onto a smooth polyurethane surface.

alpha-Chemokine growth factors for adenocarcinomas; a synthetic peptide inhibitor for alpha-chemokines inhibits the growth of adenocarcinoma cell lines.

PURPOSE: The experiments aimed to determine if alpha-chemokine inhibitors are effective suppressors of the growth of adenocarcinomas, a neoplasm with a high mortality rate. METHODS: Expression of growth-related oncogene (GROalpha) and interleukin-8 (IL-8) was determined by enzyme-linked immunosorbent assay. Inhibition of alpha-chemokine binding to tumor cells was assessed in the presence and absence of the hexapeptide, antileukinate. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays were performed to determine the effect of alpha-chemokines, monoclonal antibodies (mAb), and antileukinate on cell proliferation. Finally, antileukinate inhibition of human, lung adenocarcinoma tumor growth, was determined in BALB/c nude mice. RESULTS: All of the adenocarcinomas tested produced either GROalpha or IL-8 or both. Proliferation of lung, stomach and colon adenocarcinoma cells was inhibited by anti-GROalpha mAb and/or anti-IL-8 mAb while recombinant human GROalpha stimulated the proliferation of lung and stomach adenocarcinomas. Antileukinate inhibited GROalpha binding to specific receptors on adenocarcinoma cells and inhibited the proliferation of all adenocarcinomas tested. Colon-derived adenocarcinomas specifically bound IL-8 and this binding was also inhibited by antileukinate. Administration of antileukinate in vivo inhibited the tumor growth of adenocarcinoma A549. CONCLUSIONS: GROalpha and IL-8 are necessary for the growth of lung, stomach and colon adenocarcinomas, and can be inhibited by the hexapeptide, antileukinate. The findings suggest the possibility of using alpha-chemokine receptor inhibitors in the treatment of adenocarcinoma.

Catalytic properties of yeast protein kinase C: difference between the yeast and mammalian enzymes.

With bovine myelin basic protein as a model common substrate, protein kinases C (PKC) purified from yeast (Saccharomyces cerevisiae) and mammalian tissue (rat brain) were shown to exhibit clearly different catalytic properties. The major sites of phosphorylation in bovine myelin basic protein by the yeast PKC were identified: Thr-19, Thr-34, and Thr-65. These sites are distinctly different from those for the mammalian PKC: Ser-8, Ser-46, Ser-55, Ser-110, Ser-132, Ser-151, and Ser-161, which were previously identified [Kishimoto, A., Nishiyama, K., Nakanishi, H., Uratsuji, Y., Nomura, H., Takeyama, Y., & Nishizuka, Y. (1985) J. Biol. Chem. 160, 12492-12499]. The results suggest that the yeast and mammalian enzymes may play distinct roles in cellular regulation. No evidence is available, however, that a yeast-type PKC exists in mammalian tissues. An oligopeptide containing the sequence around Thr-19 of bovine myelin basic protein, Lys-Tyr-Leu-Ala-Ser-Ala-Ser-Thr(19)-Met-Asp-His-Ala, can be used as a substrate for selective assaying of the yeast PKC.

Synthetic utility of stannyl enolates as radical alkylating agents.

[reaction: see text] The radical-initiated beta-ketoalkylation of haloalkanes with tributylstannyl enolates is described. Stannyl enolates derived from aromatic ketones are reactive toward the homolytic beta-ketoalkylation of simple haloalkanes as well as those activated by an electron-withdrawing group. The reactivity of stannyl enolates as radical alkylating agents can be utilized for an efficient three-component coupling reaction among stannyl enolates, haloalkanes, and electron-deficient alkenes.

Homolytic carbostannylation of alkenes and alkynes with tributylstannyl enolates.

In the presence of AIBN, tributylstannyl enolates derived from aromatic ketones reacted with electron-deficient alkenes and a variety of alkynes to give the corresponding carbostannylated adducts. The reactions with methyl acrylate gave alpha-tributylstannylmethyl-gamma-ketoesters, unlike the known Michael-type reaction of stannyl enolates forming delta-ketoesters. The carbostannylation of alkynes proceeded in an anti addition mode to afford beta,gamma-unsaturated ketones. The reactivity of stannyl enolates as radical transfer agents could be utilized for radical cyclization of 1,6-enynes.

Cellular and intracellular localization of epsilon-subspecies of protein kinase C in the rat brain; presynaptic localization of the epsilon-subspecies.

The cellular and intracellular localization of the epsilon-subspecies of protein kinase C (PKC) in the rat brain was demonstrated by immunocytochemistry using specific antibodies against epsilon-PKC. The epsilon-PKC-specific immunoreactivity was most abundant in the hippocampal formation, olfactory tubercle and Calleja's islands, was moderate in the cerebral cortex, anterior olfactory nuclei, accumbens nucleus, lateral septal nuclei and caudate-putamen and low in the thalamus and medulla. The epsilon-PKC-immunoreactivity was scanty in the perikarya, except for the pyramidal cells of CA3 region of the hippocampus and the immunoreactivity was mainly present in neuropils and nerve fibers. The distribution of epsilon-PKC immunoreactive neurons was consistent with that obtained by in situ hybridization histochemistry. Electron microscopic observations of the hippocampus revealed that the epsilon-PKC is predominantly present in the cytoplasm of axon and nerve terminals and that this enzyme is associated with mitochondrial membrane and vesicles. These results suggested that epsilon-PKC is probably involved in presynaptic functions in CNS, perhaps even neurotransmitter release.

Castleman disease presenting with ophthalmic signs and symptoms.

PURPOSE: To describe a patient with multicentric Castleman disease who was initially examined with ophthalmic signs and symptoms. METHODS: Case report. A 71-year-old man was initially examined with swelling of both upper eyelids and diplopia of 2 months' duration. RESULTS: Medical evaluation and right axillary lymph node biopsy disclosed Castleman disease. Systemic corticosteroid treatment temporarily resolved signs and symptoms, but the patient died of recurrence with cytomegalovirus and Aspergillus infection 10 months after initial examination. CONCLUSIONS: Multicentric Castleman disease is a rare but distinct disorder that may present initially with ocular signs and symptoms. This disease must be included in the differential diagnosis of orbital pseudotumor and lymphoma.

A synthetic peptide inhibitor for alpha-chemokines inhibits the tumour growth and pulmonary metastasis of human melanoma cells in nude mice.

Growth-related oncogene-alpha (GROalpha) was first described as an autocrine mitogen and growth factor for melanoma cells. More recent studies show that GROalpha, interleukin-8 (IL-8) and other members of the alpha-chemokine superfamily are also angiogenic. Therefore, we sought to determine if inhibitors of the alpha-chemokine receptor would be effective in inhibiting the tumour growth and pulmonary metastasis of human melanoma cells. We determined that melanocytes and 12 human melanoma cell lines produce both GROalpha and IL-8. The proliferation of A375SM, a highly metastatic cell line, and C8161-C were significantly increased by human recombinant GROalpha and inhibited by anti-human GROalpha monoclonal antibody. Antileukinate, a potent inhibitor of alpha-chemokine receptor binding, inhibited the binding of GROalpha to its receptors in melanocytes and all 12 melanoma cell lines tested. Antileukinate also suppressed proliferation of A375SM and C8161-C cells in a dose-dependent manner, and the suppression was not due to cytotoxic effects. Furthermore, continuous administration of antileukinate inhibited the tumour growth and pulmonary metastasis of A375SM cells in athymic BALB/c nude mice. These findings suggest that antileukinate inhibits the growth of melanoma cells by preventing GROalpha from binding to its receptors. This suggests a possible use of alpha-chemokine receptor inhibitors such as antileukinate in the treatment of malignant melanoma.

Necrotizing fasciitis caused by Vibrio vulnificus differs from that caused by streptococcal infection.

We reviewed the clinical record of all patients admitted to Saga Medical School Hospital during the most recent 10 years and found that 17 (0.03%) were diagnosed as having necrotizing fasciitis. Bacteriological examination demonstrated that Vibrio vulnificus was the pathogen responsible in five patients (29%). The disease caused by V. vulnificus occurred in the warmer half of the year. All of the patients had underlying chronic liver dysfunction, and three of them had previously consumed raw seafood. In these patients, the predominant skin lesions were oedema and subcutaneous bleeding, such as ecchymosis and purpura, while superficial necrosis was not recognized. Three patients died of systemic complications. By contrast, all of the five patients with necrotizing fasciitis caused by Streptococcus pyogenes had the disorder in winter, and only one of them had chronic liver dysfunction. In skin lesions, subcutaneous bleeding was rare but necrosis was seen often. Despite the high incidence of systemic complications, no patients with streptococcal necrotizing fasciitis died. These findings suggest that the clinical features of necrotizing fasciitis caused by V. vulnificus are different from those of necrotizing fasciitis caused by classical pathogens, and that the two should be differentiated as early as possible to improve the prognosis.

Basal ganglia mass lesions in juvenile rheumatoid arthritis.

A 2-year-old boy suffered aphasia, hypotonia, dystonia, and loss of activity and spontaneous speech during an active stage of juvenile rheumatoid arthritis with pericarditis, fever, anemia, and a high antinuclear antibody titer. These neurologic signs slowly improved with corticosteroid treatment but fluctuated over 1 year. The neuroimaging studies revealed irregular mass lesions in the basal ganglia bilaterally mainly involving the globus pallidus. They gradually decreased in size and almost disappeared after 1 year. A stereotactic brain biopsy revealed a slight proliferation of astrocytes. Chorioretinitis was also observed during the clinical course. A chronic inflammatory process involving cerebral vessels was suspected, although angiography did not demonstrate cerebral vasculitis. The possibility of central nervous system lymphoma could not be eliminated. The type of aphasia and the relation to the lesion sites are discussed.

[Experimental pulmonary metastasis of Chinese hamster mesenchymal chondrosarcoma cell lines].

Pulmonary metastases of chinese hamster mesenchymal chondrosarcoma cell lines, MCS-1 (undifferentiated type) and MCS-8 (differentiated type), were examined by intravenous transplantation into athymic nude mice. The incidence of pulmonary metastasis of MCS-1 was 100% and that of MCS-8 was 33% at the 23rd day after transfer. Mean number of metastatic nodules in the lung was 41 in the former and only 3 in the latter. Mean survival time of mice with MCS-1 injection (5 x 10(4) cells) was 27 days and that with MCS-8 (5 x 10(4) cells) was 48 days after transfer. At the 42nd day after transfer of MCS-8, however, the incidence of pulmonary metastasis was 100%. These data suggest that the tumor growth rate in the metastatic lesion, as well as the affinity to the target organ, is very important for evaluation of experimental metastasis.

[Hospital infection with methicillin-resistant Staphylococcus aureus (MRSA) in Saga Medical School Hospital, a rapid increase in coagulase type-VIII strains].

Hospital infection with MRSA has increased in Saga Medical School Hospital. The causative MRSA consisted predominantly of coagulase type-II strain before 1989, but after 1990, coagulase type-VII MRSA increased rapidly. This type-VII strain has marked multiple drug-resistance, and the pattern of drug sensitivity of MRSA in this hospital was different from that of MRSA detected in other facilities, which are clinically serious problems, therefore, we conducted an etiological study of the background of the increase in MRSA infection in our hospital. The results of the study are summarized as follows: 1) The proportions of MRSA (on strain from one patient) to all types of S. aureus detected in the hospital were 26% for 1986, 23% for 1988, 37% for 1989, 30% for 1990 and 60% for 1991. The proportion increased greatly in 1991. 2) Coagulase type VII-MRSA was first detected only in 5 patients in 1989, then it tended to spread, and this type (probably derived from the same strain) accounted for 47% of MRSA infection in patients examined in 1991. 3) The study of the drug sensitivity pattern and etiological survey of the infection showed that coagulase type VII-MRSA prevalent in the hospital consisted of two types: CLDM, and EM-sensitive, IPM/CS, and MINO-resistant and TSST-1 non-producing and enterotoxin non-producing type, and CLDM, and EM-sensitive, IPM/CS, and MINO-resistant and TSST-1 non-producing type with enterotoxin serotype A. 4) Coagulase type VII-MRSA (Probably derived from the same strain) was detected in physicians and nurses working in affected wards and also in the patients's room.(ABSTRACT TRUNCATED AT 250 WORDS)

[Isolation of penicillin-resistant Streptococcus pneumoniae in Saga Medical School Hospital].

A recent nationwide increase in beta-lactams-resistant Streptococcus pneumoniae has attracted a great deal of attention. We studied the drug sensitivity of S. pneumoniae isolated from various clinical specimens in Saga Medical School Hospital between April 1988 and December 1991. To determine the drug sensitivity of the strains, we used a micro-dilution method and determined the MIC. Drug resistance was evaluated using MIC of ampicillin (ABPC) as a reference MIC, and the results were roughly classified into the following three groups: sensitive (< or = 0.1 microgram/ml), moderately resistant (0.2-3.13 micrograms/ml) and highly resistant (> or = 6.25 micrograms/ml). The isolation frequency was calculated on the basis of one strain from one patient. No strain of S. pneumoniae with high resistance against ABPC was found in 1988 (94 strains of S. pneumoniae were isolated) and 1990 (115 strains isolated), but one such strain (0.8%) was found among 129 strains isolated in 1989, and 2 such strains (2.4%) among 84 strains isolated in 1991. Moderately resistant strains were isolated at the frequencies of 12.8%, 15.5%, 22.6%, and 21.4% respectively, in 1988, 1989, 1990, and 1991. A sum of the frequencies of "moderately resistant" and "highly resistant" (2.4%) strains was 23.8% in 1991. The frequency of resistant strains is increasing and the intensity of resistance is also being elevated.