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Biomedical subjects

A M Dannenberg

Publications and source records attributed to A M Dannenberg.

At least 19 recordsLinked to original sources
PubMed1996

Rabbit vascular endothelial adhesion molecules: ELAM-1 is most elevated in acute inflammation, whereas VCAM-1 and ICAM-1 predominate in chronic inflammation.

Activation of the microvasculature is a major component of the inflammatory response. During inflammation the vascular endothelium not only becomes more permeable to plasma proteins but also develops adhesion molecules that initiate the local immigration of leukocytes. We describe herein the in vivo changes in the three major vascular adhesion molecules during the development and healing of two types of rabbit dermal inflammatory lesions: (1) acute lesions produced in rabbits by the topical application of 1% sulfur mustard (SM, the military irritant/toxicant); and (2) chronic (immune-mediated) lesions produced in rabbits by intradermal injections of Mycobacterium bovis (BCG), the vaccine strain of tubercle bacillus. In each case, frozen tissue sections were made from lesions of various ages and stained immunohistochemically for von Willebrand (vW) factor to measure the total functional microvasculature. The sections were also stained immunohistochemically for the vascular endothelial adhesion molecules ICAM-1, ELAM-1 (E-selectin), and VCAM-1, and for the leukocyte ligands for ICAM-1: LFA-1 (CD11a/CD18) and Mac-1 (CD11b/CD18). Infiltrating monocytes and lymphocytes expressed the LFA-1 ligand and infiltrating PMN expressed the MAC-1 ligand. The area of stained microvasculature per square millimeter of tissue section was determined with the use of a computerized image analyzer. Edema and cell infiltration spread apart the microvessels, changing the number of microvessels per square millimeter of tissue section. Three methods of assessing such changes are presented. In SM lesions, endothelial ICAM levels were decreased from normal by about 50% at 1 and 2 days (when the lesions reached their peak size) and returned to normal at 3 and 6 days (during the healing process). ELAM rose in peak SM lesions and remained high during healing. VCAM levels, however, were only elevated in the 6-day (almost healed) lesions. In BCG lesions the levels of endothelial ICAM and VCAM (and to a lesser extent ELAM) were increased at 9 days and remained so as the size of the lesions peaked at 23 days. During the healing phase at 37 days, the elevated ICAM and VCAM levels decreased but the slightly increased ELAM levels persisted. These findings indicate that ELAM plays a major role in acute inflammation and that VCAM and ICAM play major roles in chronic inflammation. VCAM is known to be monocyte and lymphocyte selective.

Acute Disease↗
PubMed1996

The cytokines NAP-1 (IL-8), MCP-1, IL-1 beta, and GRO in rabbit inflammatory skin lesions produced by the chemical irritant sulfur mustard.

Developing and healing dermal inflammatory lesions were produced in rabbits by the topical application of dilute sulfur mustard (SM), the military vesicant. In tissue sections of such lesions, cells containing the mRNA of important cytokines were identified with in situ hybridization techniques. These cytokines were neutrophil attractant/activation protein-1 (NAP-1 (also called IL-8), monocyte chemoattractant (activating) protein 1 (MCP-1), interleukin 1 (beta) (IL-1 (beta)), and GRO (a growth factor and chemokine). Mononuclear cells (mainly macrophages and activated fibroblasts) contained the mRNA of all four of these cytokines. A higher percentage of cytokine-producing mononuclear cells (macrophages and activated fibroblasts) was present in lesions at 2 days (their peak size) than at 6 days, when they were almost healed. Granulocytes emigrated from the bloodstream, passed through the lesions, and were the major constituent of the protective crust. This sequence correlated with the distribution of cells able to produce NAP-1: At 2 days and 6 days, the mononuclears that contained messenger RNA for this granulocyte chemoattractant were found mainly in the upper part of the dermis. At 2 days and 6 days, cells containing the mRNA of IL-1, a primary cytokine, were also found predominantly in the upper dermis, i.e., nearest the site of injury. In contrast, mononuclears containing the mRNA of MCP-1 (a monocyte chemoattractant), and the mRNA of GRO (a granulocyte chemoattractant) were more equally distributed throughout the dermis. SM stimulated hair follicle epithelial cells to up-regulate GRO mRNA and, to a lesser degree, NAP-1 mRNA. Apparently, the irritation produced by SM directly or indirectly induces such epithelial cells to manufacture these growth factors. In the rabbit, hair follicles are known to be the main source of new epithelial cells after the covering epithelium has been destroyed. Therefore, GRO is probably a major autocrine-paracrine stimulus for such repair. A brief review of the role of cytokines in dermal inflammation is presented.

Animals↗
PubMed1996

Cavitary tuberculosis produced in rabbits by aerosolized virulent tubercle bacilli.

Liquefaction of solid caseous tuberculous lesions and the subsequent cavity formation are probably the most dangerous processes in the pathogenesis of human pulmonary tuberculosis. In liquefied caseum, the tubercle bacilli grow extracellularly for the first time since the onset of the disease and can reach such large numbers that mutants with antimicrobial resistance may develop. From a cavity, the bacilli enter the bronchial tree and spread to other parts of the lung and also to other people. Of the commonly used laboratory animals, the rabbit is the only one in which cavitary tuberculosis can be readily produced. This report is the first to describe and analyze the complete course of cavitary tuberculosis, produced by aerosolized virulent bovine-type tubercle bacilli in commercially available New Zealand white rabbits. After the inhalation of 220 to 880 bacillary units, all of the rabbits were overtly well until they were sacrificed at 33 weeks. After the inhalation of 3,900 to 5,800 bacillary units, half of the rabbits died of progressive tuberculosis between 5 and 9 weeks and the other half lived until they were sacrificed at 18 weeks. Pulmonary cavities developed in both low- and high-dose groups, some beginning as early as 6 weeks. Bacilli from primary cavities sometimes caused nearby secondary cavities, but more frequently, they ascended the bronchial escalator, were swallowed, and caused secondary tubercles in the lymphoid tissue of the appendix and ileocecal junction. Histologically, and by culture, the number of bacilli found in the liquefied caseum varied from many to comparatively few. Strong tuberculin reactions at 4 weeks after infection were associated with fewer primary lesions, while strong tuberculin reactions at 33 weeks were associated with more cavitary lesions. In the tuberculous granulation tissue surrounding caseous and liquefied pulmonary foci and cavities, we found many mature epithelioid macrophages that contained high levels of the proteinase cathepsin D. Therefore, cathepsin D probably plays a major role in the liquefaction of solid caseous material and in the subsequent cavity formation.

Aerosols↗
PubMed1994

Histochemical demonstration of hydrogen peroxide production by leukocytes in fixed-frozen tissue sections of inflammatory lesions.

The production of H2O2 by cells in cold paraformaldehyde-fixed frozen sections of inflammatory lesions was histochemically demonstrated by incubating them with diaminobenzidine (DAB) for 2 to 6 h. Catalase (150 micrograms/ml, about 1400 U/ml) inhibited the reaction, indicating that H2O2 was required to produce the chromogenic DAB product. Granulocytes (PMNs and eosinophils) were the main types of cells stained by the DAB reaction. Positive staining of macrophages was less frequent. The H2O2 was produced by metabolic enzymes that were still active after cell death and mild fixation. An atmosphere of 95 to 100% oxygen enhanced the specific DAB reaction, and an atmosphere of 100% nitrogen eliminated it. The DAB histochemical reaction to detect H2O2 requires the presence of peroxidases to produce the colored reaction product. Within our tissue sections, such peroxidases were evidently present in excess, because addition of low concentrations of H2O2 significantly increased the reaction product. Although some of the H2O2 produced by the granulocytes may have been derived from the dismutation of superoxide (O2-), the NADPH oxidase pathway for O2- formation did not seem to be involved: NADPH oxidase, a rather labile enzyme, should not be active after mild fixation, and diphenyleneiodonium (100 microM), an inhibitor of flavine-requiring NADPH oxidase, did not inhibit the reaction. Reactive nitrogen intermediates were also not involved, because NG-monomethyl-L-arginine and NG-nitro-L-arginine methyl ester, inhibitors of nitric oxide synthetase, did not appreciably inhibit the reaction. We conclude that stable, non-flavine-requiring oxidases, possibly cyclooxygenases or lipoxygenases, produced the H2O2 measured histochemically by our DAB reaction. These studies were made on tissue sections of acute dermal inflammatory lesions produced in rabbits by the topical application of 1% sulfur mustard [bis(2-chloroethyl) sulfide] in methylene chloride. Both intact PMNs and disintegrating PMNs in the base of the crust produced H2O2. Despite the production of H2O2 and the presence of peroxidase activity, no tissue damage was seen microscopically near the H2O2-producing cells, which indicates that the tissues are well protected by the antioxidants present in this self-limiting inflammatory reaction.

Amitrole↗
PubMed1994

Roles of cytotoxic delayed-type hypersensitivity and macrophage-activating cell-mediated immunity in the pathogenesis of tuberculosis.

The tubercle bacillus is a facultative intracellular parasite that grows well in non-activated macrophages. When large numbers of these bacilli have grown intracellularly within such macrophages, a cytotoxic immune response, herein called tissue-damaging (or necrotizing) delayed-type hypersensitivity (DTH), kills the macrophages (and usually some of the surrounding tissue), forming the caseous center of the developing tubercle. In solid caseum, tubercle bacilli may survive, but do not multiply. When bacilli escape from the edge of the caseum, they are rapidly ingested by nearby viable macrophages. If these macrophages have not been activated, the bacilli again multiply intracellularly, and the cytotoxic immune response kills the bacilli-laden macrophages (and surrounding tissue), thus enlarging the caseous center. In hosts that develop poor activation of macrophages, this process is repeated until much of the lung is destroyed. In hosts that can develop good activation of macrophages (by cytokines from antigen-specific T cells), herein called cell-mediated immunity (CMI), the caseous centers become surrounded by these activated macrophages, which ingest and destroy the bacilli escaping from the caseum. This process can arrest the disease. Unfortunately, the caseous center may liquefy in such resistant hosts. In the liquefied menstruum, the bacilli may grow extracellularly (for the first time during the course of the disease), reaching tremendous numbers. The cytotoxic immune response to these numerous bacilli and their tuberculin-like products causes much tissue necrosis, including erosion of the walls of small bronchi, which results in cavity formation. From such cavities, the bacilli spread to other parts of the lung and to the environment. The extracellular multiplication of tubercle bacilli in the liquefied caseum is the main reason why tuberculosis perpetuates itself in mankind. It is also the reason why antimicrobial drug-resistant bacillary strains develop. To elucidate the various mechanisms involved in macrophage activation, caseation, and liquefaction is a major challenge for tuberculosis researchers today.

Animals↗
PubMed1993

Immunopathogenesis of pulmonary tuberculosis.

Antimicrobial therapy quickly eradicates susceptible bacilli, but the fight against drug-resistant disease must be waged entirely by host defenses. Knowledge of the two main types of immune response against tuberculosis and of how to manipulate those mechanisms--leading to precisely designed recombinant BCG vaccines--is essential to mounting an effective attack on the current epidemic.

Animals↗
PubMed1993

Immediate and delayed (late-phase) dermal contact sensitivity reactions in guinea pigs. Passive transfer by IgG1 antibodies, initiation by mast cell degranulation, and suppression by soybean proteinase inhibitor.

Specific immediate and delayed (24-hour) local dermal reactivity to oxazolone and dinitrochlorobenzene (DNCB) was passively transferred to naive guinea pigs by the intradermal injection of immune guinea pig serum and its IgG1 fraction. In both actively sensitized and passively sensitized animals, neutrophils were the major cells present in the immediate reaction to the specific contactant. Basophils and mononuclear cells were the major cells present in the delayed reaction to the contactant. This late-phase reaction is, therefore, a cutaneous basophil hypersensitivity (CBH) response. The serum factor that passively transferred this CBH response was stable when heated at 56 degrees C for 1-4 h. Since transfer factor, cutaneous basophil hypersensitivity factor and guinea pig IgE are heat-sensitive, these factors probably made little or no contribution to this response. Because proteinase inhibitors are known to inhibit mast-cell degranulation in vitro, we tested the effect of soybean proteinase inhibitor in vivo. This inhibitor suppressed both the immediate and the delayed skin reactivities mediated by intradermal contactant-specific IgG1. These studies support the following concept: IgG1 in guinea pigs (and IgE in human beings) sensitize mast cells to specific antigens. Such antigens degranulate mast cells, releasing histamine and other mediators for the immediate hypersensitivity reaction, and cause mast cells to produce cytokines that recruit basophils, eosinophils and mononuclear cells for the late-phase (CBH) reaction.

Animals↗
PubMed1992

Antigen-specific IgG1-mediated epidermal cell injury: a component of contact hypersensitivity reactions in guinea pigs, measurable in vitro in full-thickness skin explants.

Guinea pigs were sensitized by the topical application of either dinitrochlorobenzene (DNCB) or oxazolone on days 1, 2, 3, and 10. Seventeen days after the first treatment with the sensitizer, full-thickness 1.0-cm2 explants of untreated areas of skin were topically exposed in vitro to these contactants. Compared to the response of skin from control guinea pigs, skin from specifically sensitized animals showed a dose-related increase in the number of epidermal cells containing vacuoles. A specific increase in epidermal microblistering paralleled the increase in epidermal vacuolization. In addition, skin explants from sensitized animals (exposed to the contactant) showed a specific decrease in the incorporation of [14C]leucine. Full-thickness skin explants from unsensitized guinea pigs were sensitized in vitro by the intradermal injection of serum IgG1 fraction from oxazolone-sensitized guinea pigs. In such passively sensitized explants, the specific contactant produced an increase in the number of epidermal vacuoles, an increase in the amount of microblistering, and a decrease in the number of mast cells detectable by Giemsa staining. To elicit this specific response, the concentration of the specific contactant had to be mildly injurious, as well as antigenic. This requirement for nonspecific injury could be met by topically exposing skin explants to a nonspecific irritant followed by a sub-threshold concentration of the specific contactant. In contrast to vacuole formation and blistering, contactant-specific degranulation of mast cells (measured by the decrease in their number) did not require irritant levels of the contactant. These studies show that several components of contact sensitivity reactions can be reproduced in vitro by the passive transfer of sera containing antigen-specific immunoglobulins. Banks of such sera might, therefore, be useful in identifying (in human populations) many pre-existing sensitivities to chemical compounds.

Animals↗
PubMed1991

Delayed-type hypersensitivity and cell-mediated immunity in the pathogenesis of tuberculosis.

It is widely believed that cell-mediated immunity and the associated ability of macrophages to destroy or inhibit the bacillus are all that is required to control pulmonary tuberculosis. However, although cell-mediated immunity is a major host defense against the tubercle bacillus, it is fully effective only in one of the four stages of the disease. Here, Arthur Dannenberg describes the entire pathogenesis of tuberculosis, with illustrations from the rabbit model of M.B. Lurie. In addition, he documents that the delayed-type hypersensitivity reaction (producing tissue necrosis) greatly benefits the host by arresting the logarithmic growth of bacilli within immature macrophages.

Animals↗
PubMed1990

Physiologic oxygen tensions limit oxidant-mediated killing of schistosome eggs by inflammatory cells and isolated granulomas.

Explanted hepatic granulomas, eosinophils obtained from the peritoneal cavity of schistosome-infected mice, schistosome egg granuloma macrophages, alveolar macrophages, and activated peritoneal macrophages obtained from Listeria-infected mice were miracidicidal when cultured at 21% oxygen. This activity was markedly attenuated at physiologic oxygen concentrations (1-15%). Catalase and superoxide dismutase blocked the miracidicidal activity of inflammatory cells but did not prevent granuloma-mediated egg killing. However, the biomimetic superoxide dismutase, copper (II) [diisopropyl salicylate]2, inhibited granuloma-mediated egg killing in a dose-dependent, apparently nontoxic manner. Thioglycollate-elicited macrophages did not kill schistosome egg miracidia even when cultured in 21% oxygen, unless pretreated with lipopolysaccharide. Isolated schistosome eggs initiated an oxidative burst in macrophages, as measured by superoxide anion production. This burst was suppressed at reduced oxygen concentrations. Thus schistosome egg miracidia can be killed nonspecifically by macrophages through the release of cytotoxic reactive oxygen intermediates triggered by the egg. This activity is not supported by the oxygen concentrations found in most tissues, with the possible exception of the lung. Schistosoma mansoni eggs, injected intraveneously and lodged in the pulmonary vasculature of mice, were killed rapidly, with a half life of 3.5 days. Eggs, injected into the mesenteric veins and lodged in the liver, remained fully viable for several weeks. The data suggest that the high oxygen tension of the lung allows for the increased production of reactive oxygen intermediates (ROI) by local inflammatory cells, which in turn increases their miracidicidal efficiency. Conversely, the relatively hypoxic environment of the liver decreases ROI production by local inflammatory cells and decreases their miracidicidal efficiency.

Animals↗
PubMed1990

Full-thickness human skin explants for testing the toxicity of topically applied chemicals.

This report describes a model organ-culture system for testing the toxicity of chemical substances that are topically applied to human skin. In this system, the viable keratinocytes in the full-thickness skin explants are protected by the same keratinized layer as skin remaining on the donor, and toxicity can be assessed microscopically and/or biochemically. The human skin specimens were discards from a variety of surgical procedures. They were cut into full-thickness 1.0-cm2 explants, and briefly exposed to the military vesicant sulfur mustard (SM), which was used as a model toxicant. The explants were then organ cultured in small Petri dishes for 24 h at 36 degrees C. In the 0.03-1.0% dosage range, a straight-line dose-response relationship occurred between the concentration of SM applied and the number of paranuclear vacuoles seen histologically in the epidermis. Within the same SM dosage range, there was also a proportional decrease in 14C-leucine incorporation by the explants. Thus, the number of paranuclear vacuoles reflected decreases in protein synthesis by the injured epidermal cells. The epidermis of full-thickness untreated (control) human skin explants usually remained viable for 7 d when stored at 4 degrees C in culture medium. During storage, a relatively small number of paranuclear vacuoles developed within the epidermis, but the explants were still quite satisfactory for testing SM toxicity. Incubation (for 4 or 24 h at 36 degrees C) of such control skin explants reduced (often by 50%) the small number of paranuclear vacuoles produced during 4-7 d of storage. This reduction was probably caused by autolysis of many of the vacuolated cells. Two types of paranuclear vacuoles could be identified by both light and electron microscopy: a storage type and a toxicant type. The storage type seemed to be caused by autolysis of cell components. The toxicant type seemed to be caused by an invagination of the plasma membrane. Only toxicant-type vacuoles increased appreciably in number when skin explants were exposed to mustard, and to other toxicants.

Administration, Topical↗
PubMed1990

Extracellular collagenase, proteoglycanase and products of their activity, released in organ culture by intact dermal inflammatory lesions produced by sulfur mustard.

Peak (1 and 2 d) and healing (3, 6, and 10 d) inflammatory lesions were produced in rabbits by the topical application of the military vesicant, bis(2-chloroethyl)sulfide, commonly called sulfur mustard (SM). SM produces an acute sterile dermal inflammatory reaction with little or no necrosis, except in the epidermis, which dies during the first day. After an animal was killed, its lesions were excised intact, as full-thickness 1.0-cm2 explants. They were then organ-cultured for 3 d in order to maintain the viability of both local and infiltrating cells. The extracellular fluid in each lesion equilibrated with the culture fluid, which was collected daily and analyzed for collagenase and proteoglycanase activities. These metalloproteinase activities were measured after we had i) destroyed the alpha-macroglobulin inhibitors with KSCN, ii) destroyed the tissue inhibitor of metalloproteinases (TIMP) by reduction and alkylation, and iii) activated the latent proteinase activity with aminophenylmercuric acetate (APMA). Hydroxyproline-containing peptides and glycosaminoglycans (GAG) released into the culture fluids were also measured as indicators of local collagenase and proteoglycanase activity within the inflammatory lesions. In general, the levels of both the metalloproteinases and the products of their activity were higher in second- and third-day culture fluids than in first-day culture fluids, and higher in fluids from SM lesions than in those from normal skin. The activated fibroblast was apparently the major cell type producing the collagenase and proteoglycanase. The hydrolysis of collagen and ground substance occurs pericellularly. An excess of inhibitors exists outside the pericellular region. The daily change in culture fluids apparently decreased such inhibitors, so that by the second and third day of culture we could detect the changes in pericellular enzyme activity that were not detectable on the first day of culture. As the inflammatory lesions healed, the extracellular enzyme products (hydroxyproline and GAG) increased more than the enzymes that produced these products. With healing, a decrease occurs in the extravasation of all serum components, especially the large ones such as the alpha-macroglobulin inhibitors. We propose that during healing, the decrease in these inhibitors allows the metalloproteinases to begin the remodeling process, and that during the peak phase of inflammation, these same inhibitors protect extracellular matrix against hydrolysis by such proteinases.

Animals↗
PubMed1988

Sources of extracellular lysosomal enzymes released in organ-culture by developing and healing inflammatory lesions.

Developing and healing inflammatory lesions were topically produced in the skin of rabbits by sulfur mustard (SM). After the rabbits were sacrificed, the various lesions were removed and organ-cultured. The organ-culture fluids extracted the extracellular lysosomal enzymes (acid phosphatase, beta-glucuronidase, beta-galactosidase, and lysozyme), so that they could be measured biochemically along with lactic dehydrogenase (LDH), an enzyme marker for cell death. In tissue sections, the number and types of cells were counted, and their lysosomal enzyme content evaluated histochemically. The culture fluids from peak lesions contained much lower levels of all five enzymes than did culture fluids from healing lesions. When histological-histochemical-biochemical correlations were made, serum, macrophages (MN), and activated fibroblasts (but not tissue PMN) appeared to be major sources of extracellular lysosomal enzymes in peak lesions; and the dead PMN in the crusts and the activated fibroblasts in the tissues appeared to be major sources in healing lesions. The high lysosomal enzyme content of the crusts covering the lesions suggests that this passive barrier may also play an active role in promoting healing and in protecting against invasion by microorganisms.

Animals↗
PubMed1988

Proteases released in organ culture by acute dermal inflammatory lesions produced in vivo in rabbit skin by sulfur mustard: hydrolysis of synthetic peptide substrates for trypsin-like and chymotrypsin-like enzymes.

The purpose of these studies was to identify some of the extracellular proteolytic enzymes associated with the development and healing of acute inflammatory lesions. Lesions were produced in the skin of rabbits by the topical application of the military vesicant, sulfur mustard (SM). Full-thickness, 1-cm2 central biopsies of the lesions were organ-cultured for one to three days, and the culture fluids were assayed for proteases with a variety of substrates. When compared to culture fluids from normal skin, the culture fluids from both developing and healing SM lesions had three to six times the levels of proteases hydrolyzing two synthetic peptide substrates: (1) t-butyloxycarbonyl-Leu-Gly-Arg-4-trifluoromethylcoumarin-7-amide(Boc-Leu -Gly- Arg-AFC, herein abbreviated LGA-AFC), and (2) N-benzoyl-phenylalanine-beta-naphthyl ester (BPN). LGA-AFC is a substrate for trypsin, plasmin, plasminogen activator, thrombin, kallikrein, and the C3 and C5 convertases; BPN is a chymotrypsin and cathepsin G substrate. The culture fluids did not consistently hydrolyze four other synthetic peptide substrates or the proteins [14C]-casein and [14C]elastin. In order to determine the likely sources of LGA-AFCase and BPNase activity, we counted the number of granulocytes (PMNs), macrophages (MNs) and activated fibroblasts in histologic sections of developing and healing SM lesions, and we measured the levels of these enzymes in serum, in culture fluids of PMN and MN peritoneal exudate cells, and in culture fluids of two fibroblast cell lines. In SM lesions, serum and fibroblasts seemed to be the major source of LGA-AFCase, and serum alone the major source of BPNase. Tissue PMNs and MNs seemed to be only minor sources. The crusts of healing lesions, which were full of dead PMNs, seemed to be a rich source of both enzymes. In the SM lesion culture fluids, whether LGA-AFC and BPN were hydrolyzed by endopeptidases or only by exopeptidases could be determined by evaluating complex formation with alpha-macroglobulin proteinase inhibitors (alpha M). Endopeptidases, but not exopeptidases, are entrapped and inhibited by alpha M, because an internal peptide band in alpha M must first be hydrolyzed before molecular rearrangement (required for proteinase inhibition) occurs. The catalytic site of endopeptidases that are entrapped and inhibited by alpha M is known to remain active on (and reachable by) small synthetic peptide substrates such as LGA-AFC and BPN.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1987

Inflammatory mediators and modulators released in organ culture from rabbit skin lesions produced in vivo by sulfur mustard. III. Electrophoretic protein fractions, trypsin-inhibitory capacity, alpha 1-proteinase inhibitor, and alpha 1- and alpha 2-macroglobulin proteinase inhibitors of culture fluids and serum.

This is the third report in a series on the inflammatory mediators and modulators released in organ culture from skin lesions of various ages, which were produced in vivo in rabbits by the military vesicant, sulfur mustard (SM). It describes the electrophoretic protein fractions and trypsin-inhibitory capacities of the various culture fluids and the amounts of alpha 1-proteinase inhibitor and alpha-macroglobulin proteinase inhibitors in these fluids. With one-dimensional electrophoresis, the albumin and beta-globulin fractions of protein in culture fluids varied little with the development and healing of the SM lesions. These fractions proportionally resembled the corresponding fractions found in serum. The alpha 1-globulin fraction was proportionally smaller than the corresponding fractions of serum as the lesions healed. The alpha 2-globulin fraction was proportionally smaller than the corresponding fractions of serum at all stages of lesion development and healing. The gamma-globulin fraction was proportionally larger as the lesions healed. With two-dimensional electrophoresis, about 68%, 46%, and 35% of the protein spots in culture fluids from representative 1-day and 6-day SM lesions and normal skin, respectively, matched those from serum. In each case, the large, diffuse, serum albumin spot represented about two-thirds of the protein present. Thus, gravimetrically, in normal skin and in both developing and healing lesions, the extracellular proteins were 80-90% of serum origin. The trypsin-inhibitory capacity (TIC) per milligram protein in the culture fluids of healing lesions was markedly less than the TIC per milligram protein in the fluids of peak lesions. This decrease correlates well with the decrease found in the alpha 1-globulin fraction, which contains alpha 1-antiproteinase (alpha 1-PI) (and alpha 1-macroglobulin [alpha 1M] in rabbits). The alpha 1PI and the alpha 1M-alpha 2M proteinase inhibitors were identified in the culture fluids by means of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Western blots, specific antibodies, and the immuno-peroxidase technique. The levels of both free and proteinase-complexed alpha 1PI and alpha M inhibitors in the culture fluids decreased as the lesions healed. In both developing and healing lesions, at least half of the alpha 1PI and alpha M inhibitors seemed to be complexed with proteinases. Thus, serum seems to be a major source of unbounded extracellular protein within acute inflammatory lesions, and serum proteinase inhibitors seem to be the host's major defense against local damage by proteinases from serum, infiltrating leukocytes, and activated fibroblasts.

Aging↗
PubMed1986

Interstitial fibrosis and collateral ventilation.

Interstitial fibrosis may increase resistance to collateral flow (Rcoll) because of decreased lung volume and destruction of collateral channels or it may decrease Rcoll because of emphysematous changes around fibrotic regions. In addition, if interstitial fibrosis involves a small region of lung periphery, interdependence from surrounding unaffected lung should produce relatively large changes in volume of the fibrotic region during lung inflation. We studied the effects of interstitial fibrosis on collateral airflow by measuring Rcoll at functional residual capacity (FRC) in nine mongrel dogs before and 28 days after the local instillation of bleomycin into selected lung segments. In six of these dogs Rcoll was also measured at a higher lung volume (transpulmonary pressure = 12 cmH2O above FRC pressure). Rcoll increased in fibrotic lung segments following local treatment with bleomycin. With lung inflation (high transpulmonary pressure) Rcoll fell a similar proportion in fibrotic and nonfibrotic lung regions. These observations suggest that collateral resistance increases in fibrotic segments because lung volume decreases or because collateral pathways are involved directly in the fibrotic process. Compensatory increases in collateral communications do not occur. In addition, pulmonary interdependence does not cause disproportionate increases in volume and decreases in Rcoll of the fibrotic region during lung inflation.

Airway Resistance↗