Darrel Whitaker

Mesothelial regeneration is not dependent on subserosal cells

It has been proposed that after mesothelial injury, resident cells within the subserosal connective tissue proliferate, differentiate, and migrate to the serosal surface. The aim of this study was to examine the temporal and spatial changes of proliferating cells in a murine model of testicular mesothelial healing and assess the potential of submesothelial cells to reconstitute the damaged mesothelium. Histology and autoradiography were employed to determine the number of cells within the submesothelial connective tissue, as well as the proportion of cells undergoing DNA synthesis on and beneath the injured serosa. Mesothelial cells surrounding the wound demonstrated maximal DNA synthesis 48 h after injury (27.82±5.64% SEM, compared with 0.17±0.16% 3H‐TdR labelled cells for resting mesothelium), whereas a significant increase in proliferating submesothelial cells was not seen until day 4 post‐injury (7.79±3.31% compared with 0.85±0.64% 3H‐TdR labelled cells at day 2). Furthermore, this small number of dividing submesothelial cells must include cells other than the proposed mesothelial precursors, indicating a very low proportion of precursor cells in the submesothelial cell population. As large numbers of mesothelial cells were seen at the wound centre by 3–4 days after injury, it is unlikely that submesothelial cells contributed significantly to the repopulation of the injured mesothelium. It is hypothesized that regenerating mesothelium is more likely to originate from the surrounding uninjured mesothelial cell population. Copyright

Stimulation of Mesothelial Cell Proliferation by Exudate Macrophages Enhances Serosal Wound Healing in a Murine Model

Examination of thermally induced serosal lesions in mice displayed collections of inflammatory cells, predominantly macrophages, on and surrounding the wound within 48 hours of injury. Furthermore, by 2 days a large number of uninjured mesothelial cells adjacent to the wound were synthesizing DNA. From these findings, it was hypothesized that macrophages play a major role in serosal repair by stimulating mesothelial cell proliferation. Again, using a murine model of mesothelial regeneration, depletion of circulating monocytes significantly delayed serosal healing whereas addition of peritoneal exudate cells to the wound site 36 hours before injury increased the healing rate. In vivo assessment of mesothelial cell proliferation using tritiated thymidine incorporation and autoradiography demonstrated that peritoneal exudate cells stimulated mesothelial cell proliferation (12.44 +/- 1.63% labeling index, compared with controls in which medium only was used 4.48 +/- 0.71%). The mesothelial proliferation was predominantly because of macrophage-secreted products with molecular weights of 36 to 53 kd or 67 to 100 kd. These data support the hypothesis that macrophages play an important role in serosal healing by stimulating mesothelial cell proliferation.

Comparison of measures of exposure to asbestos in former crocidolite workers from Wittenoom Gorge, W. Australia

Determinations of exposure-response relationships between crocidolite and the major asbestos-related diseases in the Wittenoom cohort have previously depended on the validity of estimates of airborne exposure to asbestos. This work aims to validate the airborne exposure measurements by obtaining measurements of the concentrations of uncoated crocidolite fibers and asbestos bodies retained in the lungs of individual workers, and to estimate the half-life of crocidolite fibers in the lungs. Samples of lung tissue, excluding tumor, of all former Wittenoom workers known to have died in Western Australia (WA) were sought from teaching hospitals, pathology departments, and the Coroners pathologist. The lung specimens were processed using Pooleys method with TEM for counts of fibers of all types and using Smith and Naylors method with conventional light microscopy for asbestos bodies (AB). Multiple linear regression was utilized to examine the associations between crocidolite concentrations in the lung and duration of employment at Wittenoom, time since last employed at Wittenoom, nature of job, estimated average fiber concentration at the worksite, and estimated cumulative crocidolite exposure (CCE) in fiber-years/ml for each subject. Lung tissue from 90 cases was processed and there was good agreement between counts of crocidolite fibers, asbestos bodies, and CCE. Correlations were 0.77 for AB and fibers, 0.54 for AB and CCE, and 0.58 for CCE and fibers, after log transformation. The half-life of crocidolite fibers in the lung was estimated at 92 months (95% CI 55-277 months). Previous estimates of airborne exposure to Wittenoom crocidolite have been reasonably reliable. The relatively simple technique of light microscopy for counting ABs in lung tissue also provides a useful and reliable indication of the level of past occupational exposure to crocidolite in subjects whose exposure has been only to crocidolite. The half-life of crocidolite fibers in the lungs of former Wittenoom workers is about 7-8 years.

Detection of tissue CEA-like substance as an aid in the differential diagnosis of malignant mesothelioma

Summary Sections of various adenocarcinomas and malignant mesotheliomas were tested for carcinoembryonic antigen (CEA) localized in tissues by the immunoperoxidase technique; epithelial mucin was demonstrated with the PAS technique. While CEA and mucin were found in many adenocarcinomas, both were absent in the 43 cases of malignant mesothelioma we investigated. In the problem of distinguishing between adenocarcinoma and mesothelioma, the CEA‐test in combination with conventional stains for mucin is a useful technique and clearly identifies most adenocarcinomas. A dual negative result for CEA and mucin, although not proving that a given lesion is a mesothelioma, adds considerable support to this histological diagnosis.

Medullary carcinoma of the thyroid

A preoperative diagnosis of medullary carcinoma of the thyroid (MCT) allows for the investigation of associated multiple endocrine neoplasia/pheochromocytoma, and definitive surgery without the need for frozen section. Criteria for cytodiagnosis are well known but variable patterns of presentation may cause diagnostic difficulty.

CHANGES IN THE CONCENTRATION OF MICROVILLI ON THE FREE SURFACE OF HEALING MESOTHELIUM ARE ASSOCIATED WITH ALTERATIONS IN SURFACE MEMBRANE CHARGE

The luminal plasmalemma of regenerating mesothelial cells was examined by transmission electron microscopy and the concentration of microvilli at various stages of healing was quantified. Charged tracer and lectin binding techniques were also employed to investigate electrostatic and chemical changes in mesothelial glycocalyx. In uninjured mesothelium and at all stages of healing, the concentration of microvilli at the cellular periphery was greater than over the main cell mass (P<0·05). Furthermore, there was an increase in the concentration of microvilli in all regions by day 4, which reached a maximum at day 6, then at days 10–15 returned to values closer to uninjured mesothelium (P<0·01). These changes were associated with an alteration in surface charge. In all lesions, the surface charge on microvillar membranes was greater than for flat membranes, but the difference was only significant at days 4, 6, and 15 (P<0·001). The changes in surface charge may reflect a differential expression of mucopolysaccharides on the surface membrane. In addition, concanavalin A bound avidly to mesothelial surface membranes, suggesting the presence of α‐methyl‐d‐mannoside residues. These findings suggest an association between microvillar formation and surface charge, the former protecting the healing mesothelium by enhancing entrapment of serosal fluid and its contents.

Culturing mouse peritoneal mesothelial cells.

Obtaining normal cells has become increasingly important for use in comparative genetic analytical techniques to examine alterations in gene expression during transformation and progression into malignancy. Normal mesothelial cells are not currently available in cell banks and are essential for comparison of genetic expression analysis in current mouse mesothelioma models. The purpose of this investigation was to extract normal mouse peritoneal mesothelial cells using minimal culture techniques to obtain sufficient cells for gene expression analysis. Mesothelial cells were collected from the mouse peritoneal cavity in vivo with minimal contamination of lymphocytes and macrophages. The cells were cultured for approximately eight days until they were just confluent and retained normal mesothelial phenotype and cytokeratin immunoperoxidase staining.

Pleural fluid CEA levels in the diagnosis of malignant mesothelioma

&NA; A study was undertaken to investigate whether the levels of carcino‐embryonic antigen (CEA) in serous effusions helped distinguish between primary mesothelioma and metastatic adenocarcinoma in a body cavity. The investigation was designed to assess the role of the assay only when cytological analysis of the fluid had already shown the presence of malignant cells. No case of mesothelioma studied had levels of CEA above 2.9 ng/ml, whereas 67% of the adenocarcinomas tested were above 15 ng/ml. The test is helpful, therefore, in differentiating between these two types of malignant effusions.

Interaction between dactinomycin and tumor necrosis factor in mesothelioma. Cachexia without oncoiysis

There is no effective therapy for human malignant mesothelioma, and its susceptibility to recombinant cytokines has not been studied extensively. Recombinant human tumor necrosis factor α (rHuTNFα) was evaluated for its in vitro and in vivo antitumor activity using a human malignant mesothelioma cell line [DeH128(m)], both in culture and heterotransplanted in nude mice. In vitro, rHuTNF alone had no direct antimesothelioma activity assessed using the 3‐[4,5‐dimethylthiazol‐2‐yl]‐2,5‐diphenyl‐tetrazolium bromide assay, but in combination with the transcription inhibitor, dactinomycin (AD), mesothelioma cell metabolic activity was inhibited (80% of control). The effects of this combination of agents were studied on DeH128(m) cells heterotransplanted as subcutaneous tumors in nude mice. In vivo there was no significant inhibition of tumor growth by combined rHuTNFα and AD therapy, but the combination produced marked cachexia in doses at which each component (rHuTNF alone or AD alone) was well tolerated. The authors conclude that the well‐described in vitro interaction between AD and rHuTNF also operates in vivo to produce cachexia and that the combination of these two agents is likely to have a low therapeutic index in malignant mesothelioma.

A case of filariasis diagnosed on gastric cytology.

&NA; Cytological examination of gastric washings or brushing is a widely accepted technique used as a diagnostic aid in the detection of gastric malignancy. Occasionally, cytology may also provide valuable information concerning a variety of non‐neoplastic disorders. This brief report is of the exceptional experience of finding numerous microfilariae of Loa loa type in a gastric lavage specimen. This observation led to appropriate further investigation and successful treatment.