Cat-scratch disease diagnosed serologically using an enzyme immunoassay in a patient with neuroretinitis.
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Antibodies, Bacterial↗
Biomedical subjects
T J Martin
Publications and source records attributed to T J Martin.
Expression of parathyroid hormone-related protein in cells of osteoblast lineage.
The expression of parathyroid hormone-related protein (PTHrP) was studied in a range of cell cultures representative of the osteoblast lineage and in rat calvarial sections. Primary newborn rat calvarial cells, a rat preosteoblastic cell line (UMR 201), a mouse stromal cell line (ST 2), a mouse calvaria-derived osteoblastic cell line (KS 4), and rat osteosarcoma cell lines (UMR 106-01 and -06), all expressed PTHrP when examined by reverse transcription polymerase chain reaction (RT-PCR). Using a radioimmunoassay we also demonstrated PTHrP in the conditioned medium of the cultured cells, with the exception of UMR 106-01 and -06 cells. Treatment of UMR 201 cells with all-trans-retinoic acid which induces them to acquire a more differentiated phenotype, also led to a time-dependent decrease in PTHrP mRNA levels as determined by RT-PCR, Northern blot analysis, and in situ hybridization. Decreased PTHrP levels in the conditioned medium of the treated cells was also observed. These results suggested that PTHrP production might be greater in less mature osteoblasts. Examination of the populations obtained from newborn rat calvariae by sequential collagenase digestion revealed that the early digests exhibited low ALP activity, low expression of PTH/PTHrP receptor mRNA, and no adenylate cyclase response to PTHrP(1-34). These populations showed the highest level of mRNA and production of PTHrP. Cells from later digests, the "osteoblast-rich" populations, had reduced PTHrP expression. Immunohistochemistry and in situ hybridization in sections of newborn rat calvariae showed PTHrP expression in cuboidal osteoblasts located adjacent to bone and in spindle-shaped cells in the periosteal region. It is concluded that PTHrP is produced by cells of the osteoblast lineage, supporting the hypothesis that PTHrP may function physiologically as a paracrine factor in bone.
Animals↗
Effects of continuous calcitonin treatment on osteoclast-like cell development and calcitonin receptor expression in mouse bone marrow cultures.
Continuous treatment with calcitonin (CT) to inhibit osteoclastic bone resorption results in acquired resistance. The mechanisms of this "escape" phenomenon are not yet established. The aim of this study was to examine the effects of continuous treatment with CT on the generation of osteoclasts and calcitonin receptor (CTR) expression in mouse bone marrow cultures. This was done by daily CT treatment of mouse bone marrow cultures from day 0, when only undifferentiated mononuclear precursors of osteoclast-like cells were present, or commencing from day 6, when differentiated osteoclast-like cells were abundant. The response to CT treatment was determined by quantitation of cells positive for tartrate-resistant acid phosphatase (TRAP) and binding of 125I-salmon CT. Calcitonin receptor and TRAP mRNA levels were determined using semi-quantitative reverse transcription/polymerase chain reaction. When cultures were treated with CT from day 0, TRAP-positive multinucleated cells appeared. These cells expressed only very low levels of CTR or CTR mRNA and were morphologically indistinguishable from osteoclast-like cells formed in control cultures. They also displayed the ability to resorb bone. Continuous CT treatment of cultures from day 6 rapidly reduced the CTR mRNA levels, with a t1/2 of 6 to 12 h, and these levels remained low thereafter. 125I-salmon CT binding capacity, as determined by autoradiography, was lost in parallel. These effects were specific for the CTR since there was no consistent effect on TRAP mRNA levels. Based on these data, we suggest that the "escape" phenomenon may result from a prolonged CT-induced loss of CT responsiveness due, at least in part, both to reduced synthesis of CTR, and to the appearance in bone of CTR-deficient osteoclasts.
Acid Phosphatase↗
Immunochemical detection of parathyroid hormone-related protein in the saccus vasculosus of a teleost fish.
Using antisera to regions of human parathyroid hormone-related protein (PTHrP) the saccus vasculosus (SV) of the sea bream (Sparus aurata) has been shown to contain immunoreactive PTHrP. By immunohistochemistry (IHC) the epithelial coronet cells in fixed and wax-embedded SV tissue reacted with antisera to the prepro region of human PTHrP (-13 to +2), the N-terminus PTHrP (1-16), and the midmolecule PTHrP (50-69). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of saccus extracts and incubation media contained two major proteins of 14.3 and 15 kDa. By Western blotting these two proteins both reacted with the three antisera used for IHC, suggesting that they are immunochemically similar to human PTHrP (1-84). Ultrastructurally the coronet cells of Sparus saccus vasculosus resembled coronet cells described for other teleosts, with an abundant smooth endoplasmic reticulum (SER) which was more highly organized in the coronets. IHC at EM level showed reaction mainly with the membranes of the SER. These results suggest that S. aurata saccus vasculosus may produce a PTHrP-like molecule similar to human PTHrP.
Animals↗
Ceramide-1-phosphate sugars: new types of glycophospholipids.
Ceramide-1-phosphate sugars were synthesized by direct glycosyl phosphite/phosphate and O-glycosyl trichloroacetimidate/phosphate exchange reactions, respectively. Thus, ceramide-1-O-phosphoric acid 5 gave with sialyl phosphite 1 as sialyl donor directly beta-linked sialyl phosphate 6; deprotection afforded the corresponding glycophospholipid ceramide-1-phosphate N-acetylneuraminate 7. Similarly, from O-glucosyl- and O-galactosyltrichloroacetimidate 10 and 13 with phosphoric acid derivative 5 glycosyl ceramide-1-phosphate sugars 12 and 15, respectively, were obtained.
Carbohydrates↗
Within-session determination of dose-response curves for heroin self-administration in rats: Comparison with between-session determination and effects of naltrexone.
A procedure was employed in the present study to obtain dose-response curves for heroin self-administration within each experimental session. The data generated using this procedure were compared to dose-response data obtained using between-session dose manipulations. The dose of heroin (18, 30, 60 or 100 micrograms/kg/inf) was varied across 4-hourly segments separated by a 20-min time-out period during which heroin was not available. The within-session dose-response procedure yielded data similar to those obtained using between-session dose manipulations when the order of dose presentation was increasing or random. However, the dose-response curve for total drug-intake was flat when the doses were presented in decreasing order. Further analysis of the dose-response curves in the within-session procedure demonstrated that the rate of heroin intake increased in the third and fourth hourly components compared to the first component, suggesting acute tolerance to the reinforcing and/or rate-suppressive effects of heroin. Furthermore, using a random order of dose presentation, administration of 3.0 mg/kg of naltrexone prior to the session shifted the dose-response curve for heroin self-administration 5-fold to the right in the within-session procedure. The data indicate that the within-session dose-response procedure can be used to investigate the pharmacology of heroin self-administration in rodents.
Animals↗
c-fos antisense DNA inhibits proliferation of osteoclast progenitors in osteoclast development but not macrophage differentiation in vitro.
We previously reported that osteoclast formation in vitro, by coculture of mouse bone marrow and primary osteoblastic cells, occurs in two phases: proliferation of osteoclast progenitors followed by terminal differentiation into mature osteoclasts. Using this coculture system, we examined the effects of c-fos antisense and sense phosphorothioate oligonucleotides on osteoclast development and macrophage differentiation. Treatment with c-fos antisense for the first 4 days of coculture inhibited osteoclast formation in a dose-dependent fashion. However, when c-fos antisense was added during the second phase of coculture (4-6 days), osteoclast formation was unaffected. In contrast, c-fos antisense treatment had no effect on the appearance of F4/80 antigen-positive cells of the macrophage lineage in these cultures or on the induction by colony stimulating factor-1 of macrophage colony formation in cultures of mouse bone marrow cells in agar. Neither osteoclast differentiation nor macrophage appearance was inhibited by adding control c-fos sense in the cocultures. When c-fos antisense was added into an assay of bone resorption by mature osteoclasts, pit formation on dentine slices was unaffected. These results indicate that c-fos plays an important role in the proliferative phase of osteoclast progenitors in osteoclast development, but not in the terminal differentiation phase or in the bone resorbing activity of mature osteoclasts. c-fos antisense specifically inhibited osteoclast formation but had no effect on macrophage development.
Analysis of Variance↗
Parathyroid hormone-related protein in hypercalcaemia associated with haematological malignancy.
The incidence of hypercalcaemia and its association with humoral mechanisms involving parathyroid hormone-related protein (PTHrP), parathyroid hormone (PTH), or 1.25(OH)2 vitamin D were assessed in a prospective study of patients admitted to a clinical haematology unit. Hypercalcaemia was detected in 18/165 patients, and was due to primary hyperparathyroidism in 3/17 patients in whom results of humoral mediator assessments were obtained. In the other patients, hypercalcaemia was associated in nine instances with myeloma, in five with B-cell non-Hodgkin's lymphoma (NHL), and in one with myeloid neoplasia. No evidence was obtained of a humoral mechanism involving 1.25(OH)2 vitamin D, but elevated circulating levels of PTHrP, comparable with those in humoral hypercalcaemia of malignancy, were present in 2/4 patients with NHL, and in 3/9 with myeloma. The relationship between presence or absence of elevated circulating PTHrP, and presence or absence of hypercalcaemia during the course of treatment, indicated PTHrP was involved in the production of hypercalcaemia. Such an association raises the possibility that PTHrP released by neoplastic cells in these disorders acts in a paracrine manner to produce local bone resorption, and when produced in greater amounts causes elevated circulating levels which make an additional humorally-mediated contribution to the development of hypercalcaemia.
Adult↗
Physiological levels of calcitonin regulate the mouse osteoclast calcitonin receptor by a protein kinase Alpha-mediated mechanism.
We have reported that calcitonin (CT) treatment induced downregulation of the CT receptor (CTR) in mouse osteoclast-like cells (OCLs). Here, we studied the features of homologous down-regulation of the CTR in mature mouse OCLs. Treatment with salmon CT (sCT) and human CT (hCT) reduced [125I]sCT specific binding. The decreased binding after 24 h of CT treatment was associated with a decrease in the cell surface receptor concentration. The extent of CT-induced down-regulation in 24 h was dose-dependent, and the ED50 value was 3.6 +/- 4.1 (mean +/- SD; n = 3) x 10(-13 M for sCT and 4.9 +/- 3.3 x 10(-11) M for hCT. These values were very similar to those for the CT inhibition of the bone-resorbing activity of OCLs. The data suggest that these two distinct actions of CT may be mediated by a common intracellular pathway. Treatment of OCLs with activators of protein kinase A (PKA) mimicked the effect of CT on CTR downregulation, whereas neither activation of protein kinase C nor elevation of intracellular Ca2+ did so. Attenuation of CT-induced CTR down-regulation by the competitive cAMP antagonist, RpcAMP, and high concentrations of H-7, but not by protein kinase C-specific inhibitors (sphingosine, staurosporine, and a lower concentration of H-7), suggested that the PKA pathway is primarily involved in homologous regulation of the CTR. The changes in CTR messenger RNA confirm the findings in binding studies and demonstrate that CT treatment of OCLs results in decreased CTR synthesis through the PKA pathway. The low concentrations of hCT that result in CTR regulation are very close to the physiological range, providing new insights into a dynamic relationship between circulating levels of CT and CTR expression in osteoclasts.
Animals↗
Calcitonin receptor down-regulation relates to calcitonin resistance in mature mouse osteoclasts.
We have reported that short calcitonin (CT) treatment of mature mouse osteoclast-like cells (OCLs) in culture induced prolonged down-regulation of the CT receptor (CTR) and desensitization to CT rechallenge, at the level of adenylate cyclase activity. In this study, we have extended those studies to examine the bone resorbing activity of OCLs pretreated with CT. OCLs, which formed on gelled type I collagen, were pretreated with salmon CT (sCT)(10(-9)M, 1 h) and 24 h later were replated onto plastic dishes or dentine slices after removal from the gel by collagenase digestion. The number and population of either mononuclear or multinuclear OCLs that adhere to either surface was not affected by sCT pretreatment. It was found that OCLs pretreated with sCT regained reduced but significant bone resorbing capacity, which was quantitated as the surface area resorbed by OCLs on dentine slices. However, compared with control, the number of resorption pits produced by sCT- pretreated OCLs was slightly reduced, and the total pit area was decreased by approximately 40-50%. The distribution of individual pit sizes was altered by sCT-pretreatment so that the number of larger pits was predominantly reduced, suggesting that short sCT treatment may produce a long lasting decrease in osteoclast mobility. sCT was able to inhibit bone resorption activity of CT-pretreated OCLs (ED50:10(-13)-10(-12)M). Importantly, the ED50 of sCT inhibition of bone resorption in sCT-pretreated OCLs was approximately 100-fold greater than for control, indicating resistance of the OCLs to CT rechallenge. Consistent with these results, treatment of OCLs with sCT greatly decreased the expression of CTR messenger RNA, whereas no significant effect was observed on the tartrate-resistant acid phosphatase messenger RNA expression, a marker of resorptive capacity of osteoclasts. These results indicate, therefore, that an important component of escape of osteoclastic resorption from CT inhibition is CT resistance of mature osteoclasts, which regain bone resorbing function.
Animals↗
Cell-specific and regulator-induced promoter usage and messenger ribonucleic acid splicing for parathyroid hormone-related protein.
PTH-related protein (PTHrP) is the principle mediator of the syndrome of humoral hypercalcemia of malignancy and has potential paracrine actions on smooth muscle, epithelial cell growth, and placental calcium transport. The human PTHrP gene is complex: a combination of three promoters, one 5' alternative splicing event and alternative 3' splicing, which produces three PTHrP isoforms (139, 141, or 173 amino acids), results in multiple PTHrP messenger RNA (mRNA) species. We employed the RT-PCR technique to identify promoter usage and splicing patterns in a range of human cell lines. Cell line-specific utilization of the promoters and the 3' alternative splicing pathways was detected among bone, breast, kidney, and lung cell lines, although each cell line could potentially produce the three PTHrP isoforms. We also determined whether some of the known regulators of PTHrP differentially modulate promoter usage or splicing patterns. Dexamethasone decreased the abundance of each of the alternative mRNA species. In contrast, epidermal growth factor and transforming growth factor-beta treatment increased the abundance of each PTHrP mRNA species, with particularly marked effects on promoter 1- and promoter 2-initiated transcripts, especially those containing exon VII or VIII. Epidermal growth factor treatment was found to alter PTHrP splicing patterns in a manner consistent with increased transcription from promoters 1 and 2 and stabilization of exon VII- and IX-containing transcripts.
Base Sequence↗
A type I collagen substrate increases PTH/PTHrP receptor mRNA expression and suppresses PTHrP mRNA expression in UMR106-06 osteoblast-like cells.
We have previously shown that the response of osteoblasts to parathyroid hormone (PTH) can be influenced at the receptor level by growth on the physiological substrate, type I collagen, or by treatment with retinoic acid. We have also shown differential expression of genes when cells of the osteoblast lineage are grown on type I collagen. The aim of this study was therefore to examine the effect of retinoic acid and growth on type I collagen on PTH/PTH-related protein (PTHrP) receptor mRNA expression in the osteosarcoma osteoblast-like cell line UMR 106-06. PTH/PTHrP receptor mRNA levels, as assessed by Northern blot, of cells grown on collagen were increased up to 2-fold compared with cells on plastic and in a concentration-dependent manner with respect to collagen. An increase was seen as early as 6 h and was maintained over a 24 h period. This was not due to increased mRNA stability. Retinoic acid decreased the level of receptor mRNA on both plastic and collagen at each time but did not alter mRNA stability. For all treatments PTH/PTHrP receptor mRNA abundance, relative to glyceraldehyde-3-phosphate dehydrogenase, increased steadily over 24 h after subculture of cells. In contrast, PTHrP mRNA levels were reduced in cells on collagen, compared with plastic. PTH-stimulated cAMP levels of cells grown on collagen were increased compared with plastic at 24 h, but not earlier. Consistent with the mRNA data, retinoic acid decreased the amplitude of cAMP responses in cells on plastic and collagen. There was no evidence for changes in adenylate cyclase per se, since forskolin-induced cAMP levels did not change with either treatment. This study shows that known modulators of osteoblast maturation also affect signal transduction in these cells by regulating gene expression of the PTH/PTHrP receptor as well as the PTHrP ligand.
Animals↗
Regulation of osteopontin expression by type I collagen in preosteoblastic UMR201 cells.
When UMR201 cells, phenotypically preosteoblastic, were placed onto a type I collagen gel, expression of osteopontin (OP) mRNA and protein were strongly upregulated, compared to cells plated onto plastic. This upregulation was dose-dependent, with respect to the concentration of collagen gel, and was observable within hours of cells having attached and spread on the substrate. Retinoic acid (RA) acted synergistically with type I collagen at each concentration to induce a much greater increase in OP mRNA than in cells on plastic. In addition, RA increased the phosphorylation of secreted OP. The exogenous collagen substrate inhibited the growth of UMR201 cells, with the extent and duration of inhibition dependent on the collagen concentration. The effect of type I collagen was specific; plating cells on fibronectin, laminin or vitronectin did not upregulate OP expression. In contrast to the effects on OP expression, the strong RA induction of alkaline phosphatase (ALP) mRNA in cells on plastic was attenuated in cells plated on type I collagen. Growth on type I collagen did not change OP mRNA stability or transcription rate, although there was decreased stability of the ALP mRNA in cells on collagen.
Alkaline Phosphatase↗
Bilateral ptosis due to mesencephalic lesions with relative preservation of ocular motility.
Three cases of bilateral ptosis with relatively normal ocular motility are presented. In two of the patients, neuroimaging demonstrated lesions in the region of the third cranial nerve subnuclei. These unusual clinical presentations are due to isolated involvement of the central caudal nucleus supplying the bilateral levator muscles, with, virtually complete sparing of other third cranial nerve structures.
Aged↗
Interleukin (IL)-6 induction of osteoclast differentiation depends on IL-6 receptors expressed on osteoblastic cells but not on osteoclast progenitors.
We reported that interleukin (IL) 6 alone cannot induce osteoclast formation in cocultures of mouse bone marrow and osteoblastic cells, but soluble IL-6 receptor (IL-6R) strikingly triggered osteoclast formation induced by IL-6. In this study, we examined the mechanism of osteoclast formation by IL-6 and related cytokines through the interaction between osteoblastic cells and osteoclast progenitors. When dexamethasone was added to the cocultures, IL-6 could stimulate osteoclast formation without the help of soluble IL-6R. Osteoblastic cells expressed a very low level of IL-6R mRNA, whereas fresh mouse spleen and bone marrow cells, both of which are considered to be osteoclast progenitors, constitutively expressed relatively high levels of IL-6R mRNA. Treatment of osteoblastic cells with dexamethasone induced a marked increase in the expression of IL-6R mRNA. By immunoblotting with antiphosphotyrosine antibody, IL-6 did not tyrosine-phosphorylate a protein with a molecular mass of 130 kD in osteoblastic cells but did so in dexamethasone-pretreated osteoblastic cells. Osteoblastic cells from transgenic mice constitutively expressing human IL-6R could support osteoclast development in the presence of human IL-6 alone in cocultures with normal spleen cells. In contrast, osteoclast progenitors in spleen cells from transgenic mice overexpressing human IL-6R were not able to differentiate into osteoclasts in response to IL-6 in cocultures with normal osteoblastic cells. These results clearly indicate that the ability of IL-6 to induce osteoclast differentiation depends on signal transduction mediated by IL-6R expressed on osteoblastic cells but not on osteoclast progenitors.
Animals↗
Digitalis-induced visual disturbances with therapeutic serum digitalis concentrations.
OBJECTIVE: To assess the role of digitalis in the development of visual symptoms severe enough to warrant ophthalmologic consultation in patients who received digitalis and who had no other clinical or laboratory evidence of digitalis toxicity. DESIGN: Clinical case study. SETTING: Neuro-ophthalmology referral practice. PATIENTS: Six elderly patients (aged 66 to 85 years) who received digitalis were referred to ophthalmologists for evaluation of photopsia (five patients) or decreased visual acuity (one patient). No patient had chromatopsia or nonvisual clinical manifestations of digitalis intoxication at the time of examination. MEASUREMENTS: All patients had serum digitalis concentrations within or below the therapeutic range. In most patients, the electroretinographic cone b-wave implicit time was longer than normal. RESULTS: Discontinuation of digitalis therapy, which was possible in five patients, was followed by resolution of visual symptoms and by shortening of the b-wave implicit time. Characteristic features of digitalis-induced photopsia were its dependence on illumination and its tendency to be localized in peripheral visual fields. CONCLUSIONS: In an elderly patient receiving digitalis, the development of photopsia characterized by innumerable points of light in the peripheral visual fields or a decrease in visual acuity raises the possibility that the patient's visual disturbance may have been digitalis induced. Digitalis-induced visual disturbances other than chromatopsia or disturbances of color vision may occur in elderly patients who have no other clinical manifestations of digitalis intoxication and who have a serum digitalis concentration within or below the therapeutic range.
Aged↗
Modulation of the signal recognition particle 54-kDa subunit (SRP54) in rat preosteoblasts by the extracellular matrix.
Rat preosteoblastic cells, UMR201, develop a more mature phenotype when subcultured onto a type I collagen gel when compared with their growth on plastic. Basal osteopontin mRNA expression is up-regulated, whereas retinoic acid-induced alkaline phosphatase expression is reduced in cells on collagen when compared with cells plated onto plastic. We have used differential display polymerase chain reaction (PCR) of mRNA to identify other mRNA species that are regulated by collagen and/or retinoic acid in UMR201 cells. A number of differentially expressed PCR products were isolated, whose sequences did not correspond to known sequences in the data bank. However, one species which was up-regulated by growth on collagen showed 95 and 94% homology to the murine and canine 54-kDa subunit of the signal recognition particle (SRP54), respectively. In time course experiments, using reverse transcription PCR, it was found that SRP54 mRNA was up-regulated in UMR201 cells as early as 1 h after subculture onto collagen, when compared with cells subcultured onto plastic, and levels remained elevated after 48 h. The increased expression of SRP54 paralleled the increased expression of a known secreted protein, osteopontin. SRP54 recognizes signal sequences of proteins destined for secretion and retards them for further elongation in the endoplasmic reticulum. The increased expression may correlate with the synthesis of specific extracellular matrix molecules in differentiating osteoblasts.
Amino Acid Sequence↗
Metabolic bone diseases.
Metabolic bone diseases often present in old age and some are more easily treatable than others. Osteoporosis is best managed by prevention, with maximisation of peak bone density and reduction of subsequent bone loss. Although hormone replacement therapy is most useful in prevention, it also has a role in established osteoporosis. Other treating agents include calcium, calcitriol, calcitonin and bisphosphonates. Osteomalacia in the elderly mainly results from vitamin D deficiency and supplementation should be considered in those at risk. The newer bisphosphonates show great promise in the treatment of Paget's disease, while surgery remains the only treatment option in primary hyperparathyroidism.
Aged↗