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Biomedical subjects

A J Kahn

Publications and source records attributed to A J Kahn.

At least 55 records · Page 3Linked to original sources
PubMed1985

Differentiation of a human leukemia cell line and expression of collagenase inhibitor.

A human collagenase inhibitor (CI) of Mr 28,500 has been extensively characterized in skin fibroblasts and identified in a variety of connective tissues. Because human alveolar macrophages synthesize and secrete both a collagenase and CI that are immunologically and functionally identical to their counterparts in fibroblasts, we studied the production of such proteins by an immature human cell line (HL60) that can be induced to differentiate along monocytic or granulocytic pathways. The cells failed to synthesize collagenase under any culture condition tested. However, upon exposure to 1,25-dihydroxyvitamin D3 or phorbol esters (PMA), both of which promote monocytic differentiation of HL60, these cells synthesized and released CI in a dose-dependent manner. Furthermore, the extent of CI expression was paralleled by the acquisition by such cells of the monocytic marker 63D3, indicating that inhibitor production and differentiation are closely correlated. This CI was immunologically and functionally identical to that produced by human macrophages and human skin fibroblasts. The quantity of CI synthesized by PMA-stimulated cells was 3- to 5-fold greater than produced by human alveolar macrophages, approximately equal to 1 microgram per 10(6) cells per day. In contrast, undifferentiated HL60 cells produced little or no detectable CI (less than or equal to 10-20 ng per 10(6) cells per day). Interestingly, when HL60 cells were stimulated to undergo granulocytic differentiation by dimethyl sulfoxide or retinoic acid, they also produced the "monocytic" CI.

Animals↗
PubMed1985

Skeletal complications of Gaucher disease.

Gaucher disease is a collection of related disorders of sphingolipid catabolism caused by the deficiency of a specific beta-glucosidase. The inefficiency of this enzyme, glucocerebrosidase, to degrade its natural substrate leads to the accumulation of the complex lipid glucocerebroside in tissue macrophages. The pathogenesis of the disease is, as yet, poorly understood. The manifestations of the disease are protean with hepatosplenomegaly and bone deterioration frequently the predominating signs. The disease most frequently causes disability because of its effects on the skeleton. This review seeks to summarize the current clinical understanding of these complications. Experience with 327 patients reveals that the bone disease in this disorder is extremely variable. The severity of the problems range from asymptomatic persons with neither radiographic, scintigraphic, nor histologic evidence of bone involvement to those whose skeleton is completely devastated by a process of osteopenia, osteonecrosis, and osteosclerosis. These severely affected individuals show the most bizarre deformities in their bones and are subject to pathologic fracture. Most patients fortunately, are less profoundly affected, but many are plagued by bone pain of an arthritic nature or by an acute prostrating bone crisis probably best described as a bone infarction. The accepted etiology that these crises are a result of vascular compromise produced by occlusion of vessels by Gaucher cells is not supported by scintigraphic or histologic studies. Moreover, the vascular hypothesis does not explain the variety of lesions of the skeleton seen in this multifocal bone disease. Preliminary metabolic and endocrinologic studies suggest that this is not a systemic disorder of metabolism which affects bone uniformly. On the contrary, the lesions are multiple and localized, and sometimes much of the skeleton is preserved. These observations suggest that bone is affected because of collections of Gaucher cells scattered throughout its substance and may be the result of a toxic process around these foci. Alternatively, the storage of glucocerebroside in tissue macrophages may disturb the generation of competent osteoclasts and thus result in a failure to maintain a healthy skeleton. Further research is needed to delineate the pathogenesis of this disorder before any effective therapy can be developed.

Adolescent↗
PubMed1984

Age dependent changes in aversive responses of mice to ethanol and their correlation with life duration.

Age dependent changes in voluntary responses to an ethanol solution were observed in CF1 male control mice, and an experimental group provided 5% sucrose (w/v) as their only liquid for ten days beginning at the age of four weeks. This was a life duration study that began when the mice were ten weeks of age. An untreated group of C57/B1 mice was similarly observed. A 3% ethanol solution (v/v) was offered the CF1 mice, and for a comparable demonstration of aversion to ethanol in C57/B1 mice, an 18% ethanol solution was required. In all three groups, the curves of decreasing proportions of behaviorally aversive mice that occurs with advancing age closely parallels the survivorship curves. In the CF1 experimentals a more rapid development of behavioral aversion during the early portion of the life cycle, and an earlier onset of its decline phase occurs. Mean life span was significantly shorter in this group than in controls. The age dependent changes in another parameter, total fluid intake per gram body weight, were compared for short-lived and long-lived subgroups of the C57/B1 mice. This ratio is relatively high early in life, decreases to significantly lower levels in adulthood, and later increase with advancing age. In the short-lived contingent lowest values are attained around the age of 40 weeks compared to 100 weeks in long-lived mice; and the subsequent phase of increasing values is similarly shifted.

Age Factors↗
PubMed1984

Glucocorticoids modulate macrophage surface oligosaccharides and their bone binding activity.

The circumstantial evidence that indicates that glucocorticoids (GC) may stimulate osteoclastic resorption in vivo has recently found support in observations that demonstrate that these compounds effectively increase the activity of isolated resorptive cells (osteoclasts, macrophage polykaryons, and elicited macrophages [MO] ) in vitro. Data are presented here that indicate that this stimulation by GC is due to an enhancement of the initial stage of the resorption process, the attachment of cells to bone, and that this is caused by alterations of cell surface oligosaccharides. Specifically, dexamethasone and cortisol enhance by 80% the attachment of MO to bone surfaces in a dose dependent manner but do not alter or reduce the binding of these cells to other surfaces (plastic, collagen, and hydroxyapatite crystals). The effect of GC on cell-bone attachment is blocked by the glycosylation inhibitor, tunicamycin, and the glycosylation modifier, swainsonine; this demonstrates that asparagine-linked oligosaccharides are involved in the stimulatory process. Flow cytometric analysis of GC-treated cells using a panel of fluoresceinated lectins confirms this by indicating a selective, enhanced exposure of plasma membrane-associated N-acetylglucosamine and N-acetylgalactosamine residues, sugars we have previously shown to be pivotal in MO-bone binding. Finally, progesterone, a known GC antagonist, blocks GC-stimulated resorption, macrophage-bone binding, and membrane oligosaccharide modification, presumably by competing for the GC receptor. Progesterone alone alters none of these processes. Thus, GC stimulates the resorptive activity of macrophages by enhancing the initial events in the degradative process (cell-bone binding) and does so, apparently, via receptor-mediator alteration of cell surface glycoproteins.

Animals↗
PubMed1983

Induction of monocytic differentiation and bone resorption by 1,25-dihydroxyvitamin D3.

1,25-Dihydroxyvitamin D3 [1,25(OH)2D3] stimulates bone resorption in man and other vertebrates, in part, by increasing the number of osteoclasts, the principal resorbing cells of bone. Because osteoclasts are very likely derived from a member(s) of the mononuclear phagocyte family, we determined if 1,25(OH)2D3 promotes maturation of these cells by studying its effects on the human promyelocytic leukemia cell line HL-60. Of the vitamin D3 metabolites tested, only 1,25(OH)2D3, at 10(-10) to 10(-7) M, induces the differentiation of HL60 into mono- and multinucleated macrophage-like cells. Phenotypic change is evident within 24 hr and reaches a plateau between 72 and 96 hr of incubation. The changes are metabolite-specific and include (i) adherence to substrate, (ii) acquisition of the morphological features of mature monocytes, (iii) a 4- to 6-fold enhancement in lysozyme synthesis and secretion, (iv) increase in the fraction of alpha-naphthyl acetate esterase-positive cells from approximately 2% to 100% of the population, and (v) the acquisition of several monocyte-associated cell surface antigens. More importantly, treated HL-60 cells acquire the capacity to bind and degrade bone matrix, two of the essential, functional characteristics of osteoclasts and related bone-resorbing cells. These results, considered together with the reported action of 1,25(OH)2D3 on nontransformed mononuclear cells, are consistent with the view that vitamin D3 enhances bone resorption and osteoclastogenesis in vivo by promoting the differentiation of precursor cells.

Animals↗
PubMed1983

Saccharides mediate the attachment of rat macrophages to bone in vitro.

Macrophages (M phi) are multipotential cells capable of giving rise to osteoclasts and of resorbing bone. Since both of these processes are ultimately dependent upon the attachment of cells to a mineralized bone surface, we have examined in this study the mechanism by which such attachment is achieved. The data show that elicited rat peritoneal M phi bind to bone in a temperature-dependent and -saturable manner with half-maximal attachment occurring within 10 min at 37 degrees C and reaching a plateau by approximately 60 min. The kinetics of binding are essentially the same whether devitalized bone particles or viable calvaria are used as a substrate. The attachment of M phi to bone is inhibited by some sugars (e.g., N-acetyl-galactosamine, thiogalactoside, beta-lactose), fetuin and asialofetuin, and by pretreating the bone with periodate. Binding is also significantly reduced when M phi are preincubated with tunicamycin and swainsonine at nontoxic concentrations sufficient to inhibit or alter glycosylation. On the other hand, exposing the cells to neuraminidase increases the capacity of M phi to bind to bone. Collectively, our observations indicate that the attachment of M phi to bone is a highly regulated process and is mediated, at least in part, by saccharides located on both the cell and the bone surface.

Animals↗
PubMed1983

Defective binding of macrophages to bone in rodent osteomalacia and vitamin D deficiency. In vitro evidence for a cellular defect and altered saccharides in the bone matrix.

In the osteomalacic as well as normal skeleton, few osteoclasts are associated with osteoid-covered bone surfaces. The reason for this particular cellular deficit is not clear, but may relate to the inability of osteoclasts and/or osteoclast precursors (monocyte-macrophages) to attach to immature, unmineralized bone matrix, a step apparently essential for normal resorptive activity and osteoclast differentiation. In this study, we have examined cell-bone binding using macrophages (M phi) and bone isolated from vitamin D-deficient rats and hypophosphatemic, osteomalacic mice and from their normal counterparts. The data show that M phi-bone attachment is greatly reduced (P less than 0.001) in both vitamin D deficiency and hypophosphatemia, but that the mechanisms responsible for this reduction are apparently different in the two disorders. In hypophosphatemia, the reduction in binding appears solely attributable to the absence or inaccessibility of bone matrix oligosaccharides or glycoproteins essential to the attachment process. In vitamin D deficiency, on the other hand, not only is the bone matrix defective as a binding substrate, but the M phi, per se, is limited in its capacity to attach to normal, vitamin D-deficient, and hypophosphatemic bone.

Animals↗
PubMed1983

Calcitonin response in circulating human lymphocytes.

The concentration of cAMP increases in human mononuclear leukocytes after exposure to salmon calcitonin (SCT). This response is lost when the cells are separated into adherent (monocytic) and nonadherent (lymphocytic) cells, although the appropriate response to prostaglandin E2 remains in both groups. Adherent and nonadherent cells, each cultured alone for 16 h, do not regain the SCT response. Coculturing adherent and nonadherent cells together for 16 h restores the SCT response in the lymphocytes. The addition of a cyclooxygenase inhibitor to this culture system prevents development of the SCT response. The SCT response may be induced in nonadherent cells by culturing them for 16 h in medium previously conditioned by the growth of mixed mononuclear leukocytes.

Calcitonin↗
PubMed1983

Multinucleation enhances macrophage-mediated bone resorption.

The degradation of mineralized connective tissues in vivo is a function of multinucleated giant cells ('clasts) derived from and belonging to the mononuclear phagocyte family. However, it is not known whether the attainment of the multinucleated state is causally or only coincidentally related to 'clast degradative activity. To address this issue, 'clast-like polykaryons were produced by exposing elicited rat macrophages to human serum (10% for 7 to 14 days). Pure populations (90%) of multinucleated cells and their mononuclear macrophage counterparts were then assessed for their ability to bind and degrade 45Ca-labeled, devitalized bone particles in vitro. Macrophage polykaryons bind bone particles significantly more effectively than ordinary elicited mononuclear macrophages (approximately 40% attachment versus approximately 25%; p less than 0.001). Similarly, 'clast-like polykaryons resorb both large (24 to 43 microns diameter) and small (less than or equal to 24 microns) particles more efficiently than age- and culture condition-matched mononuclear cells (treated to control ratios, 1.3 to 4.2:1, p less than 0.001). This difference in degradative activity remains highly significant whether resorption (45Ca release) is expressed per microgram of DNA, per 100 micrograms of protein, or per total cell surface area. We conclude that multinucleation produces qualitative changes in the resorptive capacity of macrophagic cells. The data suggest that the formation of polykaryons is a physiologically important event in 'clast development.

Animals↗
PubMed1982

An improved method for the retrieval of cultured macrophages.

We report a simple procedure for the maintenance and retrieval of rodent peritoneal macrophages on dialysis membrane supports. Culture units are easily constructed from plastic petri dishes and dialysis membranes. Macrophages adhere readily but are detached easily from the dialysis membrane by mild mechanical agitation. Cell viability remains high and the morphologic and functional characteristics of the macrophages are maintained.

Animals↗
PubMed1982

Inductive specificity of mineralized bone matrix in ectopic osteoclast differentiation.

The present report describes the first in a series of studies designed to identify the factor or factors responsible for eliciting osteoclast differentiation. Particles of mineralized and demineralized bone, hydroxyapatite (HA), and eggshell were grafted onto the chorioallantoic membranes (CAMs) of chick embryos. After 3 of 6 days, portions of CAMs with associated grafts were harvested, processed for light and electron microscopy, and examined for the presence of multinucleated giant cells with the morphological characteristics of osteoclasts. Light microscopic examination revealed that, within only 3 days, many particles of mineralized materials had become surrounded or engulfed by multinucleated giant cells. Ultrastructurally, all such cells possessed a vacuolated and mitochondria-enriched cytoplasm, but they differed in the nature of the contacts formed at the cell-particle interface. With eggshell, the cells developed filopodia but lacked clear zones and ruffled membranes. With HA, clear zones were evident but cytoplasmic extensions and membrane ruffling were absent. Implants of mineralized bone, however, elicited the formation of giant cells with prominent clear zones and ruffling of the plasma membrane like that observed in bonafide osteoclasts. In contrast, grafts of demineralized bone did not evoke giant cell formation but rather recruited two cell types morphologically akin to either fibroblasts or macrophages. We conclude that the factor(s) responsible for osteoclast differentiation resides specifically within bone matrix and is intimately associated with the mineral phase. Further, in response to such a factor(s), osteoclast differentiation can occur ectopically, outside of the developing vertebrate body.

Allantois↗
PubMed1982

Recruitment of osteoclast precursors by purified bone matrix constituents.

The osteoclast, the multinucleated giant cell of bone, is derived from circulating blood cells, most likely monocytes. Evidence has accrued that is consistent with the hypothesis that the recruitment of monocytes for osteoclast development occurs by chemotaxis. In the present study, we have examined the chemotactic response of human peripheral blood monocytes and related polymorphonuclear leucocytes to three constituents of bone matrix: peptides from Type I collagen, alpha 2-HS glycoprotein, and osteocalcin (bone gla protein). The latter two substances are among the major noncollagenous proteins of bone and are uniquely associated with calcified connective tissue. In chemotaxis assays using modified Boyden chambers, Type I collagen peptides, alpha 2HS glycoprotein, and osteocalcin evoke a dose-dependent chemotactic response in human monocytes. No chemotaxis is observed on PMNs despite their ontogenetic relationship to monocytes and their documented sensitivity to a broad range of other chemical substances. Our observations are consistent with the view that osteoclast precursors (monocytes) are mobilized by chemotaxis, and suggest that the chemoattractants responsible for this activity are derived from the bone matrix or, in the case of collagen and osteocalcin; directly from the osteoblasts which produce them.

Bone Matrix↗
PubMed1982

Differential action of the bisphosphonates (3-amino-1-hydroxypropylidene)-1,1-bisphosphonate (APD) and disodium dichloromethylidene bisphosphonate (Cl2MDP) on rat macrophage-mediated bone resorption in vitro.

The bisphosphonates (3-amino-1-hydroxypropylidene)-1,1-bisphosphonate (APD) and disodium dichloromethylidene bisphosphonate (Cl(2)MDP) effectively inhibit the accelerated bone resorption associated with some skeletal disorders, e.g., Paget's disease. However, it has not been established whether these compounds exert their inhibitory effect by rendering the bone mineral more resistant to degradation, by diminishing the activity of resorbing cells, or through some combination of both activities. In this study, we have tested these possibilities using an in vitro resorption assay system consisting of elicited rat peritoneal macrophages co-cultured with particles of (45)Ca-labeled, devitalized rat bone. This assay system permits the quantitative assessment of the action of APD and Cl(2)MDP on the two major phases of bone resorption (cell-substrate attachment and osteolysis) under circumstances where the drugs are present continuously or, most importantly for the issues in question, after the separate pretreatment of the particles or the resorbing cells. Our data indicate that (a) Both APD and Cl(2)MDP at concentrations >/=5 x 10(-6) M diminish macrophage-mediated (45)Ca release (i.e., bone resorption) in a log dose-dependent fashion. (b) A 10-min pretreatment of bone particles with either bisphosphonate (P-C-P) similarly inhibits resorptive activity, but is most pronounced with Cl(2)MDP. However, only APD is effective in reducing resorption when cells are preincubated (for 24 h) with P-C-P. (c) In cultures containing both labeled and unlabeled bone, significant inhibition occurs only when the labeled particles are coated with P-C-P (indicating that the action of P-C-P-treated bone is highly localized). (d) P-C-P does not diminish cell-bone particle attachment, an essential step in the resorptive process. On the other hand, delaying the addition of P-C-P until after cell-bone attachment is completed significantly reduces the resorption-inhibiting effect of these compounds. (e) Cl(2)MDP reduces culture DNA content in proportion to its inhibitory effect on resorption, and both the inhibitory and cytotoxic actions of this P-C-P are dependent upon the presence of bone. On the other hand, APD is cytotoxic only at very high concentrations (10(-4) M), acts independently of the presence of bone, and inhibits resorption without killing cells. We conclude that the mechanisms of action of APD and Cl(2)MDP are markedly different. Cl(2)MDP is a potent cytotoxin in the presence of bone and apparently exerts its inhibitory effect in this manner. APD is noncytotoxic at levels adequate to suppress resorption and, therefore, must inhibit macrophage activity by some other mechanism. Neither P-C-P appears to limit resorption by decreasing the solubility of mineralized bone matrix.

Animals↗
PubMed1982

Alterability of development of hemoglobin concentration in mice: transmission of changes to the next generation.

The development of hemoglobin concentration was studied in CF1 mice. Significant variations from control values appeared in the offspring of mothers kept in small poorly ventilated cages during gestation; and by providing the pregnant mothers with a solution of RNA as only liquid source. Experimental and control lines were studied for three successive generations. Results were similar in both sets of experiments. The first generation experimentals had significantly higher hemoglobin than controls. The 2nd and 3rd generation experimentals had significantly lower values than their controls. In cross-matings between control females and first generation experimental males, the offspring showed significant differences from the 2nd generation control group. Transmission of the alteration of hemoglobin development persisted for one generation only when an experimental line was no longer exposed to experimental conditions. It is hypothesized that regulatory mechanisms account for these phenomena, and regulatory genes of gametes may participate in this transmission.

Animals↗