Aggregation Of Alveolar Macrophages Research Articles

Single intratracheal administration toxicity study on safety of vapor inhalation of electrolyzed reduced water in rats.

The effects of acute intratracheal administration of electrolyzed reduced water (ERW; alkaline electrolyzed water) were investigated in rats. In this study, no deaths or near-deaths were recorded in either group, namely those treated with ERW or purified water (maximum doses of 900 mg/kg). The main symptoms observed in the rats were decreased spontaneous movements and abnormal breath sounds, which were considered to be transient symptoms caused by intratracheal administration. In addition, low values of alkaline phosphatase, total protein and lactate dehydrogenase were found in BALF tests, but these values were considered to be of low toxicological significance, since they are usually high in the presence of lung inflammation or cellular damage. This suggests that the alkalinity of ERW partially contributes to broken peptide bonds in proteins. There were no significant increases in bronchoalveolar lavage fluid protein in either group. ERW did not cause an increase in the influx of neutrophils, eosinophils, basophils, or lymphocytes, suggesting that intratracheal administration of ERW did not cause lung inflammation. ERW did not cause abnormalities in the body or pathological changes in the lungs. Aggregates of alveolar macrophages, as a measure of inflammation, were observed in both groups. These may be transient symptoms due to intratracheal administration, not due to ERW toxicity. This study confirmed the safety of intratracheal ERW infusion and demonstrated the low risk of acute toxicity for inhalation exposure to ERW aerosol or vapor. Therefore, ERW may be an effective air purifier against viruses or bacteria.

Toxicological evaluation of Ultrapure and Potent Tannic Acid (UPPTA) by inhalation exposure

The therapeutic options for coronal virus infections are limited. As SARS-CoV-2 directly targets the lungs and causes lung damage, treatment of COVID-19 with inhalants may offer more advantages over oral administration. Inhaled drug delivery provides a higher drug concentration in the target organ, where SARS-CoV-2 proliferates. In this study, we evaluated the potential systemic toxicity, relevant target organ toxicity, and toxicokinetics of Airnecflu®, Ultrapure, and Potent Tannic Acid (UPPTA) by metered-dose inhaler (MDI) inhalation to rodent and canine species once a day for 2 consecutive weeks. We further investigated the reversibility of the toxicity following a 3-week recovery period. No mortality related to the test article was observed in all the dose groups. Neither abnormalities related to the test article nor toxicologically significant changes were observed in both rodent and canine studies. In pathological examination, alveolar macrophage aggregation, perivascular/interstitial/alveolar inflammatory cell infiltration, and alveolar/bronchial epithelium hyperplasia were noted in the lung with bronchi involvement. However, after a 3-week recovery period, a substantial recovery was observed. There is limited systemic exposure to the inhalation administration. Therefore, inhalation of Airnecflu® UPPTA is safe to administer for respiratory disorders like COVID-19.

Integration of transcriptomics, proteomics and metabolomics identifies biomarkers for pulmonary injury by polyhexamethylene guanidine phosphate (PHMG-p), a humidifier disinfectant, in rats.

Polyhexamethylene guanidine phosphate (PHMG-p) was used as a humidifier disinfectant in Korea. PHMG induced severe pulmonary fibrosis in Koreans. The objective of this study was to elucidate mechanism of pulmonary toxicity caused by PHMG-p in rats using multi-omics analysis. Wistar rats were intratracheally instilled with PHMG-p by single (1.5mg/kg) administration or 4-week (0.1mg/kg, 2 times/week) repeated administration. Histopathologic examination was performed with hematoxylin and eosin staining. Alveolar macrophage aggregation and granulomatous inflammation were observed in rats treated with single dose of PHMG-p. Pulmonary fibrosis, chronic inflammation, bronchiol-alveolar fibrosis, and metaplasia of squamous cell were observed in repeated dose group. Next generation sequencing (NGS) was performed for transcriptome profiling after mRNA isolation from bronchiol-alveoli. Bronchiol-alveoli proteomic profiling was performed using an Orbitrap Q-exactive mass spectrometer. Serum and urinary metabolites were determined using 1H-NMR. Among 418 differentially expressed genes (DEGs) and 67 differentially expressed proteins (DEPs), changes of 16 mRNA levels were significantly correlated with changes of their protein levels in both single and repeated dose groups. Remarkable biological processes represented by both DEGs and DEPs were defense response, inflammatory response, response to stress, and immune response. Arginase 1 (Arg1) and lipocalin 2 (Lcn2) were identified to be major regulators for PHMG-p-induced pulmonary toxicity based on merged analysis using DEGs and DEPs. In metabolomics study, 52 metabolites (VIP > 0.5) were determined in serum and urine of single and repeated-dose groups. Glutamate and choline were selected as major metabolites. They were found to be major factors affecting inflammatory response in association with DEGs and DEPs. Arg1 and Lcn2 were suggested to be major gene and protein related to pulmonary damage by PHMG-p while serum or urinary glutamate and choline were endogenous metabolites related to pulmonary damage by PHMG-p.

Subchronic inhalation study of stone wool fibres in rats.

Pathology results after subchronic inhalation in rats of three separate fibres representing the new biosoluble high-aluminium low-silica HT type stone wool are given, and the results were compared with the results from a similar study done with the traditional stone wool MMVF21. Male Wistar rats were exposed at one exposure level by nose-only inhalation to well-characterized fibre test atmospheres. The fibres had been size selected to be largely rat respirable. The target dose was an exposure to 150 long fibres/ml (length>20 microm) in each group, and this dose was achieved for all the fibres. The negative control groups were exposed to filtered air. The exposure duration was 6 h/day, 5 days/week for 3 months, with a subsequent non-exposure period lasting 3 months. The rats were killed 1 week after the last exposure and additional post-exposure kills were performed at 1.5 and 3 months to monitor the progression of pulmonary change and fibre numbers in the lung. The assessments included bronchoalveolar lavage fluid (BALF) for evaluation of inflammatory response (e.g. protein content, enzymes, increase in polymorphonuclear leucocytes) and measurement of cell proliferation, assessment of early fibrosis through histological examination and comparison of body weight and lung lobe weights. After exposure of rats to the new biosoluble fibres no biologically significant effects were observed except that a statistically significant increase in lung weight was observed up to 1.5 months post-exposure in all three treatment groups. At 3 months post-exposure, the small increase was no longer significant. The increase in lung weight was still present in the MMVF21 group at the 3 months post-exposure kill. After 3 months exposure, lung retention of long fibres (length>20 microm) varied from 0.4 to 5.2 x 10(6) per lung for the biosoluble fibres. At 3 months post-exposure, the long fibre concentration in the lungs had decreased to 1-7% of this figure. The fibre with the relatively highest biopersistence (RIF41001) showed the highest fibre retention. The retention of the more biopersistent traditional stone wool MMVF21 was 5.7 x 10(6) per rat lung after 3 months exposure and had decreased to 64% of this figure at 3 months post-exposure. There was no clear difference in the bronchoalveolar lavage fluid cell concentration and percentage of cells between MMVF21 and the HT groups. Fibre inhalation caused a significant increase after 3 months in all the biochemical parameters measured in the BALF. Cell proliferation was enhanced at the end of exposure for MMVF21 for all three labelling indices, but only for the bronchiolar epithelium in the RIF41001 group and for alveolar parenchymal cells in the RIF43006-1 group. At the termination of the 3 month exposure period, as well as after 1.5 and 3 month recovery periods, minimal morphological changes were diagnosed in the biosoluble fibre groups. These changes included alveolar macrophage aggregation and/or microgranulomas at the bronchiolar-alveolar junction in the few rats affected. No fibrogenic potential was noted for any of the three fibres. No clear-cut difference between the different biosoluble fibre types was noted. In the MMVF21 group, minimal interstitial fibrosis was observed that gradually decreased after the 1.5 and 3 month non-exposure periods. In this study, the pathological changes found in the lungs of exposed rats were in accordance with the pathology previously reported from full lifespan inhalation studies. This may indicate that for fibres belonging to the man-made vitreous fibres group a well conducted biopersistence study is sufficient to predict possible pathogenic effects for new fibre types. The biological parameters examined in a 90 day study may indicate little additional information to contribute to the prediction of the outcome of carcinogenicity studies.

Alveolar macrophages are the main source for tumour necrosis factor-alpha in patients with sarcoidosis.

Tumour necrosis factor (TNF)-alpha is known to play a major role in the formation of noncaseating granuloma, a hallmark of sarcoidosis. The main cellular source in situ is still ambiguous. Serial sections of transbronchial biopsies from 14 patients with and 12 without sarcoidosis were studied, using immunohistochemistry (IHC), for TNF-alpha, T-cells (CD3), macrophages (CD68), and epithelial cells (MNF116). TNF-alpha spontaneously released (sr-TNF-alpha) by freshly isolated bronchoalveolar lavage cells, isolated from the same patients and cultured without any stimulus over a 24-h period was measured using an enzyme-linked immunosorbent assay. IHC revealed colocalisation of TNF-alpha with CD68 cells only. Cases with TNF-alpha tissue immunoreactivity exhibited higher sr-TNF-alpha (1,667 +/- 504 pg x mL(-1)) than cases without tissue immunoreactivity (211 +/- 60 pg x mL(-1)). In an explorative approach, a subgroup of patients could be identified and characterised by the presence of alveolar macrophage aggregates. It was found that sr-TNF-alpha was highest in this subgroup (2,700 +/- 769 pg x mL(-1)) compared with patients with normal histology (221 +/- 61 pg x mL(-1)) or with prominent granuloma (460 +/- 137 pg x mL(-1)), whereas in most clinical parameters this subgroup was intermediate. The findings from this study strongly corroborate the view that alveolar macrophages are the main cellular source for tumour necrosis factor-alpha in the initial phase of sarcoidosis. The authors suggest that in these patients, aggregates of alveolar macrophages may represent at least predecessors to granulomas if not granulomas in statu nascendi.

Homotypic Aggregation of Rat Alveolar Macrophages Stimulated by Ultrafine Carbon Black is Associated with Up-Regulation of Intercellular Adhesion Molecule-1

This study aimed to determine the effect of ultrafine carbon black (uf-CB) on alveolar macrophage (AM) adhesion molecule function. Rat AM were cultured for 4 and 21 h with either uf-CB (diameter 14 nm) or media control. Functionally relevant changes in surface adhesion molecules were determined using homotypic aggregation, and the contribution of intercellular adhesion molecule-1 (ICAM-1) and its counter-receptor CD18 were identified by blocking monoclonal antibodies (mAbs). Semiquantitative polymerase chain reaction was used to determine changes in ICAM-1 mRNA. At 1.2 µg/mm 2 , uf-CB increased AM aggregation at 4 and 21 h (P < 0.05 versus media control, n = 4), whereas 0.8 µg/mm 2 increased aggregation at 21 h only. Ultrafine-CB-stimulated aggregation at 21 h was completely blocked by mAbs to CD18 and ICAM-1 (n = 6). Semiquantitative PCR of uf-CB stimulated AM showed a mean 2.18 increase in the ICAM-1/β-actin ratio at 40 cycles over media control (n = 3). We conclude that uf-CB increases AM adhesion in vitro via an ICAM-1/CD18 interaction, and that increased adhesion is associated with increased ICAM-1 mRNA.

Bronchiolo-fiberscopic, histological and ultrastructural studies of peripheral lung tissues in bronchial asthma patients

Bronchiolar findings in B8 and 9 of the right lung were taken with a BF-1. 8T bronchiolo-fiberscope in a total of 11 patients with bronchial asthma (6 male, 5 female, average age 59 years). In these patients air pollution was considered responsible for bronchial asthma. Approximately one month after the bronchoscopic examination, transbronchial lung biopsies were performed in the same portion of the lung. Histological and electron microscopic findings were correlated with the above results. Abundant secretion of a serous nature was constantly noted in the peripheral airways of each patient, which was consistent with the histological finding of goblet cell hyperplasia. Dark greenish pigmentation over the edematous mucosa was identified as anthracotic pigments, with or without iron deposit in the tissues. All patients showed occasional aggregates of alveolar macrophages with various amounts of carbon-mixed mineral dusts in their cytoplasm. Six of them formed in the alveolar interstitium micro-granuloma which consisted of epithelioid cells and multinucleated giant cells. These monocyte-derived cells were obviously adherent to one another by development of paired or unpaired subplasmallemmal linear densities along their cell membranes. Nine patients showed regeneration of bronchiolar cuboidal and type II alveolar epithelial cells along the proximal alveolar sacs. They frequently showed nuclear inclusions. This proliferative tendency is suggestive of previous bronchiolo-alveolar damage due possibly to the air pollutants in these patients.

The Effects of Tobacco Modification on the Biological Response to 13-Week Cigarette Smoke Inhalation in Inbred Syrian Hamsters

Twice daily exposure of 12–24 male inbred Syrian BIO 15.16 hamsters for 13 weeks to maximally tolerated doses of smoke from 12 types of investigational cigarettes under experimental conditions and with previously described instrumentation (Bernfeld et al., 1979) resulted in the following response: increased occurrence and aggregation of alveolar macrophages, increased occurrence of tracheal squamous metaplasia, and increased frequency and severity of laryngeal hyperplasia. The statistical significance of these changes was evaluated by means of loglinear models. The effect of varying the nicotine level in tobacco leaf between 0.2 and 1.5% caused no statistically significant changes in the subchronic response. Smoke from Bright tobacco with average tar yields of from 22.4 to 26.2 mg per cigarette caused significantly more alveolar macrophages and laryngeal hyperplasia but less tracheal squamous metaplasia than did smoke from Burley tobacco, with from 9.3 to 10.5 mg tar per cigarette. Deproteinization of tobacco did not change the response of the hamsters to the resulting smoke. Maleic hydrazide-field treatment of tobacco significantly reduced the alveolar macrophages, but spiking of tobacco with maleic hydrazide increased the response. Previous work had suggested that the tumorigenic response to cigarette smoke in chronic inhalation studies in hamsters may be predicted from subchronic inhalation studies in the same animal model (Bernfeld et al., 1983a). The present results then suggest that smoke from low-nicotine cigarettes is not less tumorigenic in the hamster than that from high-nicotine cigarettes, that smoke from high-tar Bright tobacco is more carcinogenic than that from low-tar Burley tobacco, that deproteinization of tobacco does not affect the tumorigenicity of the resulting smoke, and that maleic hydrazide-field treatment of tobacco does not increase the tumorigenicity of the resulting smoke, whereas spiking of tobacco with maleic hydrazide might do so.

Effect of Micropolyspora faeni cells and cell wall fractions on rabbit alveolar macrophages.

The reactivity of alveolar macrophages (AM) to cells and cell wall fractions (CWF) of Micropolyspora faeni was investigated. Exposure of cultured AM to M. faeni and its CWF caused the AM to form clumps or aggregates which remained attached to the culture dish surface. Other gram-positive and gram-negative bacteria as well as yeast, zymosan, latex microspheres, and isolated peptidoglycan from Listeria monocytogenes did not cause this response. The response was independent of species source and antibody content of the serum used in culture. The use of heat-inactivated sera negated the role of complement activation in the aggregation of AM. AM cultures required a period of culture before exposure to cells or CWF for this response to occur. This response was both time and dose dependent. Rabbit peritoneal macrophages also exhibited the clumping response. Degradation of a purified CWF, fraction 3, with lysozyme greatly diminished the clumping response. Chemical purification of fraction 3 with periodate, formamide, or trichloracetic acid also decreased this activity. These data suggest that the major active component causing this response is peptidoglycan but that other materials associated with the cell wall may also be important. A soluble-factor chemotactic for normal rabbit AM was found in the culture fluid of AM exposed to fraction 3. M. faeni cells and CWF also caused normal rabbit AM to chemiluminesce.

Mutagenic and tumourigenic properties of the spores of Aspergillus clavatus.

Spore walls of a sputum-derived isolate of Aspergillus clavatus yielded mutagen(s) when their extracts were fractionally precipitated with ethanol following alkaline hydrolysis. After spores were given by nasal inoculation to 6-8-week-old CF-1 mice, light and electron microscopy of lung sections showed that they had been readily phagocytozed by the polymorphonuclear leucocytes and alveolar macrophages mobilized during early allergic alveolitis in immunized mice. The formation of phagosomes was followed in thioglycollate-stimulated peritoneal macrophages grown in vitro. Unimmunised mice showed a comparable lung reaction, attributed to pulmonary mycotoxicosis, and revealed a rising incidence of lung tumours, from 25% at 2 months from inoculation, to 27.3% at 6 and to 55.5% at 8. Mean numbers of tumours per lung rose from 1.0 to 2.2. Total tumours, including lymphomas, reached a final incidence of 77.7% at 8 months, when control animals were tumour-free. Tumour development correlated with the retention of apparently intact spores within giant cells probably derived from aggregates of alveolar macrophages. The implications of these findings in the light of the known history of human exposure to such spores is discussed.

Pneumoconiosis in Animals Exposed to Poly(Vinyl Chloride) Dust

Rats, guinea pigs and monkeys were exposed by inhalation (6 hr/day, 5 days/week) for up to 22 months to a 13 mg/m3 concentration of PVC dust. Autopsies on rats and guinea pigs were performed after 12 months of exposure and on monkeys after 22 months after 22 months of exposure. Lung function tests were performed on monkeys after 9, 14 and 22 months of exposure. Aggregates of alveolar macrophages containing PVC particles were found in the lungs of all animals. These aggregates were more numerous in the monkey lungs. No fibrosis or significant cellular infiltrates were present in or near these cellular aggregates. No significant effects on pulmonary function could be demonstrated in the monkeys exposed to PVC. Under the conditions of this experiment, inhaled PVC produced a benign pneumoconiosis.

Pneumoconiosis in animals exposed to poly(vinyl chloride) dust.

Rats, guinea pigs and monkeys were exposed by inhalation (6 hr/day, 5 days/week) for up to 22 months to a 13 mg/m3 concentration of PVC dust. Autopsies on rats and guinea pigs were performed after 12 months of exposure and on monkeys after 22 months after 22 months of exposure. Lung function tests were performed on monkeys after 9, 14 and 22 months of exposure. Aggregates of alveolar macrophages containing PVC particles were found in the lungs of all animals. These aggregates were more numerous in the monkey lungs. No fibrosis or significant cellular infiltrates were present in or near these cellular aggregates. No significant effects on pulmonary function could be demonstrated in the monkeys exposed to PVC. Under the conditions of this experiment, inhaled PVC produced a benign pneumoconiosis.

Proteoglycans, proteases, chemotaxis, and aggregation of inflammatory, cells.

A trypsin-like protease activity present in rabbit alveolar macrophages was shown to be activated by hyaluronic acid (HA) at 10(-9) M, but not by other proteoglycans. This activation of proteases by HA was accompanied by aggregation of alveolar macrophages and by the release of chemotactic activity. This chemotactic activity was destroyed by proteases and was less active for polymorphonuclear leukocytes than for alveolar macrophages. These effects of HA appear to be sepcific for alveolar macrophages, because no similar effects of HA on human polymorphonuclear leukocytes or lymphocytes could be observed. These observations suggest that HA, which is secreted into airways, plays a specific role in the macrophage-mediated chronic inflammatory processes in the distal lung.