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By R. Hogg, Y. Lee, D. E. Rannels

"Lee, Yu-Chen, Richard Hogg, and D. Eugene Rannels. Extracellular matrix synthesis by coal dust-exposed type II epithelial cells. Am. J. Physiol. 267 (Lung Cell. Mol. Physiol. 11): L365–L374, 1994.—Although integrity of the alveolar basement membrane may influence progression of lung injury induced by inhaled particulates, little is known about direct effects of coal dusts on the alveolar epithelium or its extracellu¬lar matrix (ECM). Effects of dust on synthesis of cell and ECM proteins by type II cells (T2P) was thus investigated. Three coal dusts (anthracite no. 867; bituminous no. 1451 and no. 1361) and mine dust MIT-3, of respirable size, were studied as a function of dose and time over 3 days of primary T2P culture. On day 1, 750 µg/ml of 867 were required to increase relative synthesis of ECM proteins (ECM/cell). MIT-3, 1451, and 1361 were without effect. By day 3, 867 or MIT-3 increased ECM/ cell 60-100% at 300 µg/ml; 1451 produced modest, dose-dependent stimulation, whereas 1361 remained without effect. None of the dusts caused significant cytotoxicity. The results show dose- and time-dependent effects of well-characterized coal and mine dusts to modify partitioning of newly synthesized proteins into the ECM and suggest that coal dust exposure may modulate structure or function of the subepithelial basement membrane.basement membrane; cell culture; inorganic dust; lung injury; extracellular matrix deposition; pulmonary epitheliumCHRONIC EXPOSURE to coal dust may be associated with development of pulmonary fibrosis and consequent loss of normal lung function (see Ref. 26 for review). Al¬though the pathways of inorganic dust-induced lung injury are recognized to involve an inflammatory re¬sponse, the underlying pathogenic mechanisms are not understood in detail. Acute and chronic responses to dust exposure involve a variety of interacting cells of the immune system, including macrophages, T cells, and B cells, along with pulmonary fibroblasts. These responses reflect direct effects of the dust particles, as well as indirect effects mediated by soluble factors, including cytokines and eicosanoids (see Ref. 41 for review).Studies of pulmonary macrophages and fibroblasts following silica exposure suggest that dust-induced damage at the airway surface may involve cellular injury through oxidant-mediated pathways. Coal dusts and related inorganic particulates can activate the alveolar macrophage (42) to produce fibronectin (15) and cytokines, such as interleukin-1 or tumor necrosis factor (4) and/or other soluble products which act locally (29) to cause acute cellular injury and to initiate the progression of fibrotic lung damage (16). Type I epithelial cells are typically more susceptible than type II cells to oxidant injury. Type I cell damage and subsequent death leads to denudation of the subepithelial basement membrane (2, 21) and can be associated with leakage of plasma constituents from the pulmonary vasculature into the alveolar compartment (13)."

Dec 1, 1995

By S. R. Rannels, D. E. Rannels

"I. IntroductionThe alveolar surface of the lung is covered by two populations of pneumocytes, type I and type II epithelium. The type I cell, which accounts for most of the alveolar surface area and provides a highly specialized barrier for gas exchange (Rannels et al., 1991), is poorly understood. Although approaches to type I cell purification have been discussed, they have not been applied widely because of the lack of phenotypic markers to monitor isolation procedures (Dobbs et al., 1988; Danto et al., 1992). In contrast, the type II cell has been studied extensively, largely because of the development of reliable procedures for its isolation and primary culture. Investigations based on the use of purified populations of cultured type II cells have provided convincing evidence to support a role for the pneumocytes both in the maintenance of normal lung function and in the response of the lung to injury (for review, see Rannels and Rannels, 1989).While type II cells can be isolated by a variety of approaches, the protocols used most commonly are modifications of the gradient method of Dobbs and Mason (1979) or of the panning method of Dobbs et al. (1986). Purification of the cells based on density gradient centrifugation is reliable and consistent, with yields of 30 to 40 X 106 type II cells per rat of 230 g body weight. Isolation procedures applied to a minimum of three to four rats provide the best results, as lower cell yields are common with the use of fewer animals. The protocol below is designed to obtain a maximal cell yield from the lungs of six rats.II. Materials and InstrumentationMaterialsCalbiochem: DNase I (Cat. No. 260912); Corning: culture flask, 75 cm2 #25115; Costar: culture plate, 24-well #3424; disposable serological pipette #4101; Elastin Products Inc: elastase #E-134; Falcon: cellulose nitrate membrane filter, 0.22 µm, #7105; conical tube, 50 ml, #2070; culture dish, 100 mm, #3003; culture plate, sixwell, #3046; Fisher: BaS04 , #B68-500; NaCl, #5271-3; NaHC03 , #5233-50; Fort Dodge Laboratories: pentobarbital, #1206-lE; Gibco: Dulbecco's minimal essential medium (DMEM), #430-1600EC; Hanks' balanced salt solution (HBSS), #310- 4185AG; Joklik's modified Eagle's medium (JMEM), #410-2300EC; HyClone Laboratories Inc: fetal calf serum (FCS ), # A-1115-L; I CN Biochemicals (Flow Labora tories): amphotericin B, #16-723-48; newborn calf serum (NCS), #29-121-54; try pan blue, # 19 5 5 32; Na/gene: Erlenmeyer flask, # 100-4116C; Pharmacia: Percoll, #17-0891-01; density marker beads, #17-0549-01; Sigma: gentam1cm sulfate, #G3632; penicillin/streptomycin, #P3539; trypsin inhibitor, #T9128; Tetko: HC- 160 nylon mesh, #3-160-53."

Dec 1, 1995

By R. D. Dey, V. Castranova, J. M. Antonini, J. A. Blackford Jr.

Alveolar macrophages (AM) exposed to cytokines or bacterial lipopolysaccharide (LPS) produce the free radical nitric oxide (NO·) by an inducible nitric oxide synthase (iNOS). They also release reactive oxygen free radicals following exposure to silica dust. The purpose of the present study was to determine whether NO· is produced by rat AM and/or recruited leukocytes following the intratracheal (IT) instillation of silica. Male Sprague-Dawley rats (175 to 225 g) were IT instilled with either silica dust (10 mg/100 g body wt) or LPS (0.25 mg/100 g body wt). After 24 h, bronchoalveolar lavage cells (BALC) and lavaged lung tissue were assayed for iNOS mRNA. Cell counts of BALC and iNOS-dependent (Nw-nitro-Larginine methyl ester [L-NAME]-inhibitable) chemiluminescence generated by AM were also determined. Northern blot analysis demonstrated that the steady-state levels of BALC iNOS mRNA were significantly increased by 3-fold following IT silica and by 7-fold following IT LPS. Partially enriched fractions of either AM or leukocytes from silica-treated rats both exhibited significantly elevated iNOS mRNA in Northern analysis. iNOS-dependent chemiluminescence was significantly increased in AM by 36-fold following IT silica and by 89-fold following IT LPS. Differential counts of BALC showed that AM numbers did not change in any of the treatments; however, red blood cells increased by 30-fold following IT silica and by 23-fold following IT LPS. Total leukocytes (polymorphonuclear leukocytes plus lymphocytes) increased by 58-fold following IT silica and by 274-fold following IT LPS. The results demonstrate that AM and neutrophils iNOS steady-state mRNA levels, AM iNOS-dependent chemiluminescence, and lavagable red blood cells and leukocytes all increased following IT silica. These findings suggest that NO· production in rat AM and neutrophils is increased in response to silica. NO· has the ability to then combine with superoxide and form a highly reactive toxic species, peroxynitrite, which may be responsible in part for silica-induced lung damage.

Dec 1, 1995

By L. M. Demers, D. C. Kuhn

The alveolar macrophage (AM) responds to stimuli such as coal mine dust by releasing inflammatory mediators such as cytokines, growth factors. reactive oxygen species. and eicosanoids. Eicosanoids are synthesized by°AM through the action of cyclooxygenase and lipoxygenase enzymes and serve to modulate the proinflammatory function of this cell as part of the lungs' host defense mechanism. Reactive oxygen species can be generated by AM as a by-product in the biosynthetic pathway of the prostaglandins. AM produces primarily prostaglandin E2, thromboxane A2, and leukotriene B4 as part of the cellular response to an inflammatory stimulus. There is evidence to suggest that eicosanoid production by AM is functionally linked to both surface interaction with mine dust particles like silica and by the phagocytosis of the dust particle itself. In this report, we examined the effects of an antioxidant, vitamin E. on dust-induced synthesis of PGE2 and TXB2 in vitro and in vivo by AM obtained by bronchoalveolar lavage from rats. We also looked at the effects of the surface of silica particles on AM eicosanoid biosynthesis under conditions of calcination, a process that removes exposed hydroxyl groups from the surface of silica particles, thus reducing the likelihood of soluble hydroxyl radical formation. Treatment of AM with vitamin E in vivo and in vitro reduced the augmentation in eicosanoid production usually observed when AM are exposed to mine dusts. These results suggest that vitamin E may effectively reduce the inflammatory and fibrotic response produced by inhalation of mineral dust through an antioxidant mechanism. Silica that has been chemically altered by calcination was unable to activate AM eicosanoid production in vitro when compared to untreated, freshly fractured silica. These findings suggest that the mechanism by which dust particles can activate AM eicosanoid release may involve interaction of surface and/or soluble factors with the cell membrane. Taken together, these studies point to the involvement of AM eicosanoid production as part of the proinflammatory response of this cell to occupational inhalation of mineral dust. - Environ Health Perspect 102(Suppl 10):97-100 (1994)

Dec 1, 1995

By R. L. Grayson, W. E. Wallace, M. D. Attfield, A. Q. Wearden, P. Bolsaitis, R. D. Kennedy, M. J. Keane, J. C. Harrison

"Respirable high-silica particles from four coal mines. a clay mine and a tunnelling site were analysed for silicon vs aluminum composition by performing scanning electron microscopy-energy dispersive X-ray analysis (SEM-EDS) at electron accelerating voltages of 20 kiloelectron volts (keV). The same particles were analysed at 5 keV to determine possible differences in surface vs bulk composition. In some cases high-silica particles exhibited a decrease in the ratio of silicon to aluminium K-alpha line intensities with decreasing electron beam accelerating voltage. This behaviour is consistent with aluminosilicate surface contamination or occlusion of a silica particle. Some 20 high-silica particles were analysed from each of 10 samples. Powdered aluminosilicate glass was used as a control for the behaviour of silica with a homogeneous distribution of aluminium in the particle. Significant differences were found between the control and sample means for an Illinois bituminous coal mine dust, two western and one central Pennsylvania bituminous coal mine dust low-temperature ashes. and a clay mine and mill dust sample, but not for an anthracite coal mine dust and two central Pennsylvania bituminous coal mine dust low-temperature ashes. INTRODUCTIONRESPIRABLE dust quartz content commonly is measured on bulk samples by using i.r. spectroscopy or X-ray diffraction. For research purposes, individual particle-by particle SEM-EDS analysis at 20-30 keV electron beam accelerating potential may be used also. Such measurements have not provided an unambiguous association of quartz content with occupational pulmonary disease prevalence in exposed workers (WALTON et al., 1977). Rather, there is evidence that coal rank modifies the relationship between dust exposure and pneumoconiosis development (WAL TON et al., 1977; ROBOCK and BAUER, 1990; ATTFIELD and MORRING, in press). SEM-EDS analysis was performed to measure the silicon fraction, Si/(Si +Ai), at 20 and at 5 keV on high-silica particles to determine if some respirable quartz particles generated by mining exhibit heterogenous composition with depth (WALLACE et al., 1990), which might affect the bioavailability of the quartz surface."

Dec 1, 1995

By S. L. Weber, D. E. Banks

"Silicosis was first reported by the ancient Greeks and is apparently as old as human history. The prevalence of this illness peaked in the last half of the 19th century and the early part of this century, when mechanized industry was developing and the relationship between dust exposure and disease was less well understood. Yet, even today in devel-oped countries, sporadic outbreaks of silicosis occur when workers are consistently exposed to silica particles of respir-able size (0.5 to 5.0 µm in diameter) at levels exceeding those recognized to be safe.Silicon dioxide or silica. is the earth's most abundant mineral. Silica is considered to be either free (i.e.. unbound to other minerals) or combined. Minerals with high amounts of free silica include quartz (including granite) flint. Chert, opal, chalcedony, and diatomite. Combined forms of silica are called silicates and include, among others, asbestos talc and kaolin.In the crystalline form, silica exists as quartz, cristobalite, and tridymite (Fig. 28). Of these quartz is ubiquitous in the environment and occurs as sand and as a part of many rocks. Cristobalite and tridymite occur naturally in lava and can be formed by the heating of quartz or amorphous silica. Cristo-balite is more tibrogenic than quartz. Amorphous silica is noncrystalline and, therefore, is relatively nontoxic and does not cause silicosis. It occurs as diatomite (skeletons of pre-historic marine organisms) or as vitreous silica (glass), the result of carefully melting and then quickly cooling free crystalline silica. Heating diatomite with or without alkali (a process known as calcining), forms cristobalite and adds the respiratory toxicity of free crystalline silica.There are many opportunities for occupational exposure to respirable crystalline free silica. Some of the commonly en-countered occupations in which the risk of silicosis exists are reported in Table 20.Silicosis. the pulmonary disease caused by the inhalation of silica particles of respirable size, is most commonly cate-gorized by findings on the chest radiograph. The most typical presentations simple silicosis and progressive massive fibrosis appear different radiologically but are grouped under the category of classic silicosis because they are a part of the radiographic spectrum of this illness. Radiographically, simple silicosis appears as small (less than 10 mm in diameter), rounded opacities, typically predominant in the upper lung zones. Progressive massive fibrosis is the result of the coalescence of these small nodular opacities and appears as larger opacities (more than 10 mm in diameter), again typicallydistributed in the upper lung zones."

Dec 1, 1995

By S. P. Singh

The purpose of the cut is to provide a free face to which the remainder of a development or breasting round can break. The results of field testing in drifts and at a quarry face were used as a basis for a description of the rock breakage and flushing mechanism in the cut. It was concluded that the important features of a cut design are proper hole spacing, sufficient relief volume, drilling accuracy and relief holes slightly longer than the planned length of the round. The void space provided by the relief holes should be about 15% of the cut volume, with more required in weak, highly fissured rocks. The importance of drilling accuracy increases with round length, an error being magnified by the small blasthole spacing. The first hole pulls only about 20% of the round length, 4-6 holes having to be fired before the cut is pulled close to its design depth. The first four blastholes should be fired with a minimum inter-hole delay of 100 ms to provide progressive relief of burden and avoid the superimposition of ground vibrations. The earliest-firing charge should be closest to the relief area and act laterally, while the first of the vertically-fired charges to be detonated should be the uppermost. Problems during cut-pulling include sympathetic detonation, dynamic pressure desensitisation, charge disruption and freezing of the cut

Dec 1, 1995

By M. J. Reasor, M. DiMatteo, J. M. Antonini, K. Van Dyke, Z. Ye

"The inhalation of silica has been shown to produce a dramatic inflammatory and toxic response within the lungs of humans and laboratory animals. A variety of cellular and biochemical parameters are used to assess the silica-induced lung injury. The purpose of this paper is to introduce the use of luminok:Jependent chemiluminescence as a new method to study inflammation in both phagocytic cells and lung tissue recovered from silicaexposed animals. Chemiluminescence, or the emission of light, accompanies the release of reactive forms of oxygen when phagocytic cells are challenged. In this study, male Fischer 344 rats were intratracheally instilled with either silica (10 mg/100 g bw) or saline vehicle. One day after the instillations. a marked increase in the chemiluminescence was observed in the lung tissue and bronchoalveolar lavage cells recovered from the silica- treated animals when compared with the saline controls. The light reaction was markedly decreased by either superoxide dismutase or N-nitro-Larginine methyl ester hydrochloride. Superoxide dismutase is involved in the enzymatic breakdown of superoxide anion. while N-nitro-L-arginine methyl ester hydrocholoride, a nitric oxide synthase inhibitor, prevents the formation of nitric oxide. When superoxide anion and nitric oxide react. they form the highly oxidizing substance peroxynitrite. This study then implicates peroxynitrite as an agent that may be responsible for some of the oxidant lung injury that is associated with silica exposure. The use of luminol-dependent chemiluminescence may prove valuable as a method to measure the earliest events in the inflammatory process. and may be an adjunct in studying the mechanisms that produce inflammation. - Environ Health Perspect 102(Suppl 10):37-42 (1994) IntroductionResearch on inflammation has not produced a method that links the initial events of the process to the controlling events. Rather, a variety of parameters are measured that are the result of and not the basis of the inflammatory reaction. Parameters such as tissue weight, edema, blood flow, enzyme and protein release, or production of metabolites (prostaglandins, leukotrienes) have been used extensively. A method is needed that could be applied to cells of a particular type, groups of cells of different types, and even tissue, in an attempt to study the kinetics of the inflammation as well as being sensitive enough to monitor the early events of the process . Since inflammation is so complicated, the relationship between the control of the initial events and the oxidation process has never been explained nor studied in any derail."

Dec 1, 1995

By T. R. Taubert, D. H. Carlson, J. H. Johnson

"ObjectiveDevelop a portable emissions measurement apparatus to sample and measure tailpipe concentrations of diesel particulate matter (DPM), carbon monoxide (CO), carbon dioxide (COJ, nitric oxide (NO), and nitrogen dioxide (NOJ under steady-state engine load and speed conditions during a 1-minute test.BackgroundThe U.S. Bureau of Mines (USBM) has a comprehensive research program to develop ways to reduce exposure of miners to diesel exhaust pollutants. Engine maintenance has been shown to have important effects on such exposure.Accurate measurements of tailpipe emissions are becoming an essential part of a good maintenance program for underground mines that use diesel-powered vehicles. Past studies of particulates in U.S. underground coal mines have shown that DPM alone may be present at concentrations greater than 1 milligram per cubic meter (mg/m3), half the 2-mg/m3 limit for respirable coal mine dust. A poorly tuned (smoking) diesel engine may be the sole cause for respirable dust concentrations exceeding the limit. Although there is currently no standard specifically for DPM in U.S. mines, regulatory steps are underway, and significant reductions of DPM levels in both coal and metal mines will most likely be required.Tailpipe emission measurements can provide diagnostic information to identify engine malfunctions such as clogged air cleaners, faulty injectors, or incorrect injection timing. In some cases, mine vehicle tailpipe DPM concentrations have been reduced up to 50 percent by engine maintenance and adjustments.An accurate measuring instrument would also be useful for determining the effects of exhaust control devices on tailpipe emissions. Little information is currently available about the overall effectiveness of exhaust controls after long-term use. Raw exhaust concentrations with and without the control device can be measured in underground mines, and from these data a control efficiency can be calculated. Knowledge of the effectiveness of exhaust control devices over time is needed to make intelligent decisions on bow to use and maintain these devices, and when to replace them."

Dec 1, 1995

By S. L. Weber, V. Castranova, D. E. Banks, J. K. H. Ma, Y. Rojanasakul, J. Y. C. Ma, M. Bhat

"The mechanism of bleomycin-induced pulmonary fibrosis is not ye: clear. Recent studies have shown that alveolar macrophages (AM) can be stimulated by bleomycin in vitro releasing inflammatory cytokines, suggesting that the interaction of bleomycin with AM is an important step in the drug-induced fibrotic process. Bleomycin is known to bind DNA and generate oxygen radicals through complexation with Fe2+ and oxygen. To provide,more insight into the cellular oxidative property of bleomycin, v.s have developed a fluoromicroscopic method using 2',7'-dichlorofluorescin diacetate (DCFHDA) as an oxidative fluorescence probe to study the deomycin-induced intracellular oxidation in rat AM and the inhibition of the oxida¬tion by taurine, a compound known to inhibit the bleomycin-induced fibrosis. Bleomycin at 5 to 20 ug/ml has a moderate stimulatory effect (1.87- to 2.66-fold) on the secretion of superoxide anion. A high concentration of bleomycin (20 ug/ml), however, inhibits cell response to zymosan-induced secretion of superoxide anion. At 4 ug/ml, bleomycin has no effect on cell membrane integrity or morphology but results in a significant increase in intracellular oxidation. This oxidative process is Fe2-,-dependent and is accompanied by an increase in intracellular calcium (35 nM). Both the intracellular oxidation and calcium rise induced by internalized bleomycin are inhibited by pretreatment of cells with varying concentrations of taurine (25, 125, and 187.5 uM). The inhibitory effect on intracellular oxidation .vas found to be 36, 57, and 60%, respectively. These results demonstrate a stimulation-inhibition relationship between bleomycin and taurine on re cellular oxidation at a subcytotoxic dose of bleomycin, suggesting that the oxidative effect of the intracellular bleomycin-Fe2 complex is important in the initiation of the fibrotic process. — Environ Health Perspect 102 (Suppl10):91-96 (1994) IntroductionThe pulmonary system is most susceptible to oxygen radical-induced diseases due to its constant exposure to the highest oxygen tension, and the potential encounters with oxidant gases such as ozone, nitrogen oxides, and toxic doses of dust particles. The lung is also a well-perfused organ and thereby vulnerable to the effect of circulating xenobiotics such as paraquat, and, bleomycin, which is known to cause pulmonary injuries (1). Alveolar macrophages (AM), which act as a means of defense by phagocytizing inhaled dust particles and microorganisms while increasing production of reactive oxidative species, have been shown to play an important role in mechanisms leading to oxidative lung injury and fibrosis (2-4)."

Dec 1, 1995

By A. W. Khair, L. D. Gehl

"This paper describes the mechanism of coal and rock cutting from quasi-static indentation through dynamic indentation to rotary cutting. The results indicate that during quas~static indentation, a crushed zone, and subsurface fractures develop. The crushed zone is a potential source of dust generation. Dynamic indentation produces a crushed zone due to the frictional resistance offered by the coal or rock during penetration of the bit. Confining pressure changes the mode of failure from tension to mixed tension and shear. The wider bit causes larger crushed zone and more dust generation. The rotary cutting is a combination of quasi-static and dynamic indentation which results in chip formation. 1. INTRODUCTIONThe trend from conventional mining to continuous machine-cutting of coal, using more productive machinery is inexorable. Intolerable levels of dust generated at the face is an indication of inefficient coal or rock cutting by these machines. The Federal Government of the United States paid about 11.7 billion dollars workman compensation to more than 470,000 miners with Coal Workers Pneumoconiosis (Newmeyer, 1981). Bieniawski and Zipf (1985) reported that annual benefit payments were about 2 billion dollars. The need for research m improving coal cutting 1s therefore paramount. The enactment of Health and Safety Act in 1969 in the United States triggered the initial investigation into coal cutting mechanism and cutting tool design. The European countries were also engaged in the same area of research, however, their emphasis was in the use of wedge shaped bits. In 1981, the United States Bureau of Mines commissioned the South West Research Institute to investigate the effect of operating parameters mainly on airborne respirable dust in order to provide a healthier and safer atmosphere for coal mine workers.A research program by the Department of Mining Engineering, West Virginia University, USA was initiated to study the fragmentation mechanism during coal/ rock cutting. The research program was divided into three areas of research, namely quasi-static indentation, dynamic indentation and rotary cutting."

Dec 1, 1995

By M. J. Reasor, J. M. Antonini, L. Wu, K. Van Dyke, Z. Ye

The inhalation of silica has been shown to produce a dramatic inflammatory and toxic response within the lungs of humans and· laboratory animals. Currently, no effective treatment exists for workers who may have been exposed to the inhalation of silica. The objective of this study was to develop an animal model in which we could evaluate the effect that anti-inflammatory steroids have on the acute silica-induced pulmonary inflammatory response. Male Fischer 344 rats were pretreated with either dexamethasone (2 mg/kg) or saline vehicle (i.p.) on days 1, 3, and 5. On day 6, the animals from the two groups were then intratracheally instilled with either silica (20 mg/0.5 ml saline vehicle) or saline vehicle (0.5 ml). Twenty-four hours after the instillations in the non-steroid group, significant increases occurred in total protein, total number of cells, neutrophils, and lymphocytes recovered from the lungs of animals treated with silica compared to saline controls. Silica also caused dramatic increases in the luminol-dependent chemiluminescence (LDCL) of lung tissue and bronchoalveolar lavage (BAL) cells. The LDCL reaction was markedly decreased by either superoxide dismutase (SOD) or N-nitro-L-arginine methyl ester hydrochloride (L-NAME). SOD is involved in the enzymatic breakdown of superoxide anion, while L-NAME, a nitric oxide (NO) synthase inhibitor, prevents the formation of NO. When the superoxide anion and NO react, they form the highly oxidizing substance peroxynitrite. This study then implicates peroxynitrite as an agent which may be involved in the silica-induced oxidant lung injury. When the animals were pretreated with the steroid dexamethasone, there was a complete protection against the biochemical, cellular, and chemiluminescent indices of damage caused by silica. The mechanism in which the steroid protects the lung from damage may be due to the ability of dexamethasone to block the induction of NO synthase. With further study in animals, the anti-inflammatory steroids may be useful in the treatment of silicainduced lung injury.

Dec 1, 1995

By H. Netuka, J. V. Horak, A. W. Khair

"A general review of major problems concerning mathematical modelling of roof bolting problem is given. A mathematical model (MM) of interactive set Bolt - Resin - Rock and its discretization based on the Finite Element Method (FEM) is developed to calculate numerically course of displacement, stress and strain fields in this set. This new MM enables to get also course of the normal and tangential components of contact stress vector, normal displacement and then mutual tangential slip at the bolt - resin and resin - rock interfaces, respectively.Behavior of system on all interfaces is modelled in the MM as a ""unilateral contact"" governed by so called non-classical boundary condition of Signorini's type including Coulomb's law of friction between resin and rock, and ""a given friction"" law between bolt and resin (see Duvaut, Lions (1972) for mathematical formulation).Numerical example and some values of resulting stresses are shown and remarks on necessary and sufficient conditions for solvability of the problem are also given.IntroductionAs is well known roof bolting has become the primary support system in almost all underground coal mines. But the ""theoretical"" design criteria for roof bolt patterns are rather problematic because: "" ... real mechanisms by which the roof bolts reinforce the immediate roof of underground coal mine entry are still relatively unknown"", see Peng (1986).The aim of this contribution is to present brief description of fundamentals of mathematical theory (see Hlavacek et al., 1988) which could be, according to our opinion and knowledge, very helpful for numerical modelling of a such type of problems. For the sake of brevity we restrict ourselves here to just one of the simplest possible problem: analysis of the behavior of interactive set composed by only single full - length - grouted bolt, resin (or cement) and suitable part of domain of a ""immediate"" neighborhood of bolt which should represent physical action of real rock material."

Dec 1, 1995

By R. L. Grayson, W. E. Wallace, J. W. Stephens, M. D. Attfield, P. S. Brower, V. Vallyathan, M. J. Keane, J. C. Harrison

Pure respirable quartz is a known pathogenic agent for lung disease. However, for mixed dusts, neither mineralogical nor in vitro cytotoxicity assays unequivocally predict the contribution of quartz to their pathogenic potential.First, although Coal Workers' Pneumoconiosis (CWP) and Progressive Massive Fibrosis (PMF) prevalence have been found to correlate with total respirable coal mine dust exposure, and with coal rank (Attfield and Morring, 1992), epidemiological studies generally have not found the quartz component of coal mine dust as measured by X-ray diffraction or infrared spectroscopy (each of which essentially measures the mass-percent quartz in a bulk sample) to be clear predictors of CWP prevalence or incidence (Walton et al., 1971). Second, the properties which distinguish the extreme fibrogenic potential of quartz are not revealed by conventional in vitro bioassay comparison with other silicate mineral dusts. As an example, quartz and kaolin (aluminosilicate) clay in vitro cytotoxicities are comparable for lavaged primary pulmonary macrophages.The studies discussed here address these two anomalies from a general hypothesis that the pathogenic activity of respired quartz particles is associated with properties of, and interactions with, the particle surfaces. One factor in the expression of toxic activity by a particle surface is the biological availability of that surface. Anomalous lack of correlation of disease prevalence with mass percent quartz in exposures to some mixed composition respirable-sized dusts suggests that individual particle surface features, invisible by bulk sample mass analysis, may alter the biological availability of a fraction of quartz particles in a dust. For pure-surfaced mineral particles, the comparably strong prompt membranolytic activities of quartz and kaolin in vitro suggest that upon deposition of such particles in the lung there must be other interactions with particle surfaces which distinguish quartz. This might be due to toxic surface-associated sites specific to silica, or to interactions of other sites on other mineral particles which affect the biological availability of their toxic functional groups. For example, alurninol groups on clays affect the availability or activity of adjacent silanol groups. Surface activities which distinguish quartz pathogenici ty might be manifest in one or more of the sequential interactions upon deposition in the lung; first with the pulmonary surfactant system, then with extracellular and phagolysosomal enzymatic digestive processes, or with subsequent cytotoxic processes.

Dec 1, 1995

By D. E. Banks, J. K. H. Ma, Y. Rojanasakul, J. Y. C. Ma, C. J. Malanga

There is evidence to suggest that cell injury induced in alveolar macrophages (AM) following phagocytic activation by silica particles may be mediated through changes in intracellular free calcium [Ca 2+}. However, the mechanism of silicainduced cytotoxicity relative to [Ca2+], overloading is not yet clear. To provide a better insight into this mechanism, isolated rat AMs were exposed to varying concentrations of crystalline silica (particle size< 5 µm in diameter) and the fluctuation in their [Ca2+]; and cell integrity were quantitatively monitored with the fluorescent calcium probe, Fura-2 AM, and the membrane integrity indicator, propidium iodide (Pl). Results from this study indicate that silica can rapidly increase [Ca2+]; in a dose-dependent manner with a characteristic transient calcium rise at low doses (<0.1 mg/ml) and an elevated and sustained rise at high doses (>0.1 mg/ml). Depletion of extracellular calcium [Ca 2+]0 markedly inhibited the [Ca2+]; rise (˜90%), suggesting that Ca2+ influx from extracellular source is a major mechanism for silica-induced [Ca2+]; rise. When used at low doses but sufficient to cause a transient [Ca2+]; rise, silica did not cause significant increase in cellular Pl uptake during the time of study, suggesting the presevation of membrane integrity of AMs under these conditions. At high doses of silica, however, a marked increase in Pl nuclear fluorescence was observed. Depletion of [Ca 2+] 0 greatly inhibited cellular Pl uptake, induced by 0. 1 mg/ml or higher doses of silica. This suggests that Ca 2 + influx, as a result of silica activation, is associated with cell injury. Indeed, our results further demonstrated that the low dose effect of silica on Ca 2+ influx is inhibited by the Ca2+ channel blocker nifedipine. At high doses of silica (>0.1 mg/ml), cell injury was not prevented by nifedipine or extracellular Ca2+ depletion, suggesting that other cytotoxic mechanisms, i.e., nonspecific membrane damage due to lipid peroxidation, are also responsible for the silica-induced cell injury. Silica had no significant effect on cellular ATP content during the time course of the study, indicating that the observed silicainduced (Ca2 ... ); rise was not due to the impairment of Ca2 + -ATPase pumps, which restricts Ca2+ efflux. Pretreatment of the cells with cytochalasin B to block phagocytosis failed to prevent the effect of silica on [Ca2+]; rise. Taken together, these results suggest that the elevation of [Ca2+]; caused by silica is due mainly to Ca2+ influx through plasma membrane Ca2+,channels and nonspecific membrane damage (at high doses). Neither ATP depletion nor Ca2+ leakage during phagocytosis was attributed to the silica-induced [Ca2+]; rise.

Nov 1, 1995

By P. R. Miles, J. K. H. Ma, J. Y. C. Ma, L. Bowman

"The bisbenzylisoquinoline alkaloids are compounds which can be isolated from plants and which exhibit a broad range of pharmacological activities. Perhaps the best known alkaloid of this group is tetrandrine (TT), a compound isolated from the root of a traditional Chinese medicinal herb, Stephania tetrandra. It has been tested for its effects on the cardiovascular system, tumors, and the lungs. For example, tetrandrine has been shown to act as a calcium entry blocker in cardiac sarcolemmal vesicles (King et a/., 1988) and has been used in China as a treatment for angina and hypertension. Its possible effects as a tumor-inhibitory substance have also been investigated (Kupchan and Atland, 1973). In addition, there are several lines of evidence to suggest that it is a possible therapeutic agent for the treatment of silicosis. Yu et a/. (1983) have shown that TT inhibits the development of experimental silicosis in rats when it is administered orally after intratracheal instillation of quartz. Other studies have shown that the alkaloid inhibits the synthesis of collagen by fibroblasts and the formation of silicotic nodules in rats exposed to silica (Huang et al., 1981; Liu et al., 1983). In fact, tetrandrine has been used clinically in China as an antifibrotic agent and has been shown to improve both the appearance of chest radiographs and the measurements of diffusing capacity in patients with silicosis (Li et al., 1981).Since tetrandrine may have a potential use as a therapeutic agent for fibrotic lung diseases, its effects on various pulmonary cells and tissues should be studied. Alveolar type II cells are located in the alveolar regions of the lungs and are important because they are responsible for the synthesis and secretion of pulmonary surfactant (Dobbs et al., 1982; King, 1984), regeneration of the alveolar epithelium after lung injury (Kauffman et al., 1974), and metabolism of foreign compounds (Bend et al., 1985). Furthermore, in silicosis, for which TT has been suggested as a treatment, there are increased amounts of pulmonary surfactant produced (Dethloff et al., 1986a,b) as a result of increased rates of surfactant synthesis by type II cells (Dethloff et a/., 1989; Miller et al., 1988). Because type II cells play a critical role in maintaining normal lung function and because type II cells are affected in silicosis, for which 'TT has been used as a treatment, we studied the effects of tetrandrine on energy metabolism in these cells in vitro. Energy metabolism was chosen because it is a basic cellular function which is neces-sary for other cellular processes to occur. Since type II cells depend on mitochondrial metabolism for energy production (Fisher et al., 1980), we examined the effects of tetrandrine on cellular and mitochondrial ATP production and oxygen consumption. Additional experiments were performed in an effort to determine if intracellular calcium levels may be involved in the TT-induced effects. A preliminary report of those results has appeared previously (Ma et al., 1991)."

Nov 1, 1995

By K. Clifton, K. Kar, M. Gautam, J. T. Jurewicz

"This paper describes the history of LDV measurements employed in research involving gas-solid flows. Much of the discussion deals with measurement techniques employed to obtain separate and simultaneous solid and gas phase velocities. Also, LDV based measurement techniques that arc used to obtain particle size and concentration information in conjunction with the velocity measurements arc presented.INTRODUCTIONThere are numerous examples of industrial processes in which the motion of a solid or droplet dispersed phase in a suspending gas phase is critically important. The applications range from spray droplets injected into combustion chambers to the understanding of the time release mechanisms of particles generated by dissolving pharmaceutical tablets. In many of these processes, the heat, mass, and/or momentum transfer between the dispersed phase and the suspending fluid arc the items of interest. In the past, much of the phenomena were characterized by empirical models derived from flow averaged measurements. Gradual developments in laser Doppler velocimetry (LDV), have provided researchers with an unobtrusive measurement technique capable of obtaining the particle size velocity and concentration with good spatial resolution.In the two decades that LDV instrumentation has been available, most of the uses have concentrated on single phase fluid flow measurements. The light scattered from seed particles small enough to follow complex fluid motions, including turbulence, is converted into velocity information. While the utility of this instrument in single phase flows has been demonstrated beyond question, the use of LDV in gas-solid multiphase flows has had a more gradual development. The measurements ·of interest in these flow regimes are usually the velocity of the suspending fluid as well as that of the dispersed particulate phase. Early investigators were quick to see the potential of this instrument for measurements in gas-solid flows. The interest soon expanded to the development of instruments that could also simultaneously measure particle size and concentration within a two-phase flow field.This paper tracks the historical development of LDV technology as applied to gas-solid flows. The techniques to measure particle velocity, size, and concentration arc treated in separate sections although much of the development occurred simultaneously. Finally, some unique applications conducted at West Virginia University and other locations will be discussed in the application section."

Nov 1, 1995

By R. V. Ramani, R. L. Grayson, R. L. Frantz, V. A. Marple, W. R. Myers, K. L. Rubow, Mutmansky. J. M., T. J. Stobbe

"An investigation of protocols for research performed from 1953 to 1988 on pneumoconioses (with emphasis on diseases related to coal and silica) was conducted in the Generic Mineral Technology Center for Respirable Dust. In addition, interviews were conducted with prominent U.S. medical researchers, aerosol scientists and engineers in government, industry, and universities to determine dust sources, dust preparation methods, laboratory protocols, and determinant variables related to in vitro and in vivo research and assess the comparability of results.Since the original report on the protocol project, additional research regarding free radical existence in freshly produced dust particles and the shape and contamination of particles generated for research purposes has further heightened scientists' concerns. As indicated by current research, fresher dust particles and those with more angular shapes may have a different level of reactivity in biological systems than do older or more rounded particles. Therefore, research on dust preparation methods for biomedical studies was undertaken and will be outlined here.Results of protocol review and interviews are summarized. Research on size reduction methods and their particle characteristics plus some recommendations concerning dust preparation methods and reporting in biomedical publications are also presented. Recent evidence indicates that these recommendations may be of importance in comparing and interpreting research results."

Nov 1, 1995

By R. H. Reist, R. D. Dey, V. Castranova, J. P. Durham, Y. Rojanasakul

"Silicosis is a fibrotic pulmonary disease produced by inhalation of silica-containing dusts (Peters, 1986). Chronic silicosis may develop 20-40 years after initial exposure to crystalline silica, and the disease may progress from a simple to a complicated form. Few symptoms are evident in simple silicosis with pulmonary function being relatively normal. However, chest radiographs reveal small rounded ·opacities in the upper lobes of the lungs. These silicotic nodules consist of collagen arranged in a distinctive circular pattern. As fibrotic changes become more severe, the disease progresses to complicated silicosis (progressive massive fibrosis). Complicated silicosis is associated with shortness of breath and decreases in pulmonary function characteristic ofrestrictive lung disease. Chest x rays reveal numerous large silicotic nodules which may be associated with scar emphysema. Several mechanisms have been proposed to explain the cellular events which result in silica-induced lung damage and fibrosis (Lapp and Castranova, 1993). They include: ( l) direct cytotoxicity of silica, (2) activation of oxidant and enzyme release from alveolar macrophages, (3) recruitment and activation of polymorphonuclear leukocytes, and (4) stimulation of fibroblast proliferation and collagen synthesis.Tetrandrine is a bisbenzylisoquinoline alkaloid derived from the Chinese herb Radix Stephania tetrandra. It has a molecular weight of 622.73 and an empirical formula of C38H420 6N2. The chemical structure of tetrandrine has been published previously by our laboratory (Castranova et al., 1991 a). Its structure is characterized by methoxy groups at C7 and C12 , uncharged nitrogens at N2 and N’2 and two 17-carbon ring members connected by a double oxygen bridge between Ca-C7 and C11-C12. Tetrandrine has been shown to reduce both silica-induced fibrosis and the elevation of lung collagen content in rats (Huang et al .. 1981; Yu et al., 1983; ldel, 1987). Furthermore, tetrandrine has been used as an antifibrotic agent in Chinese clinical trials (Li et al.. 1981 ). Preliminary results suggest that administration of tetrandrine to patients with silicosis results in substantial improvement of symptoms, increases in diffusion capacity, and decreases in the size of shadows on lung x rays."

Nov 1, 1995

By N. S. Dalal, A. H. Hoffman, X. Shi, D. E. Banks, J. K. H. Ma, Y. Rojanasakul

Excessive production of reactive oxygen radicals by alveolar macrophages is proposed to play an important role in oxidative lung injury. A major product of oxygen radical formation is the highly reactive hydroxyl radical (•OH) generated via a biologic Fenton reaction. In addition to its known ability to induce lipid peroxidation, recent studies have suggested that the •OH may exert its cytotoxic effect through the alteration of [Ca2+], homeostasis. To test this potential mechanism as well as to investigate the relationship between •OH and Ca" overloading in cytotoxic injury, isolated rat alveolar macrophages were exposed to externally generated radical system, H2O2 (0.01 to 1 mM) and Fe (1 mM) and their [Ca2+] levels and cell injury were monitored using quantitative fluorescence microscopy with the aid of the specific Ca" indicator, Fura-2, and membrane integrity indicator, propidium iodide. Electron spin resonance measurements using the spin trap 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) confirmed the production of the .OH radical by this system. Upon the addition of the radicals, the macrophages displayed a rapid initial rise in [Ca2+] which was followed by a slower but more pronounced [Ca2+], elevation that reached a level 3 to 5 times higher than the basal level. This process preceded cell death as evident by nuclear propidium iodide fluorescence. Depletion of extracellular Ca" inhibited both the [Ca2+], response and cell injury. Preincubation of the cells with the Ca" channel blocker verapamil or .0H radical scavenger mannitol similarly inhibited the [Ca2+], rise and loss of viability. Firefly luciferase assay of cellular ATP content demonstrated that the alterations in [Ca2+], following .0H treatment preceded the depletion of ATP. Changes in membrane potential caused by Na* substitution with K* had no significant effect on either the unstimulated or stimulated [Ca"], levels. Taken together, these measurements indicate that .OH-induced injury in alveolar macrophages is associated with the alteration in [Ca2+], homeostasis. The mechanism of •OH-induced [Ca2+] loading is not due to nonspecific membrane damage or energy depletion, but mainly due to an increase in Ca2+ influx through Ca2+ selective channels. This Ca2+ over-load state as well as energy depletion caused by the oxidant may activate certain cellular degradative processes leading to cell injury and death.

Nov 1, 1995