Can free DNA be detected in sputum of lung cancer patients?

SUMMARY: Free DNA is present in the serum of cancer patients in a higher concentration than that in non-cancer patients. Free DNA in sputum may originate from malignant or inflammatory diseases. The aim of the study was to examine the presence of free DNA in sputum and the relationship to lung cancer. The contribution of inflammatory cells was established as well. The amount of free and cellular DNA in sputum was determined using real-time beta-globin PCR in 28 lung cancer patients and 68 controls. Free DNA was present in sputum samples of the cancer patients and controls. We found no differences in DNA concentration in sputum of patients with and without lung cancer. For all patients combined the amount of free DNA was related to the amount of inflammation. Further, we found increased hypermethylation of RASSF1A in lung cancer patients compared to controls to show that tumour related DNA is present in sputum. In conclusion, free DNA can be detected in sputum of lung cancer patients. The amount of free DNA is related to the amount of inflammation, but not to the presence of lung cancer.

Progressing imbalance between proliferation and apoptosis with increasing severity of cervical intraepithelial neoplasia.

The equilibrium between cell proliferation and protection against apoptosis was studied immunohistochemically using monoclonal antibodies against Ki-67-Ag and bcl-2, respectively, in consecutive sections from normal and metaplastic cervical epithelia and cervical intraepithelial neoplasia (CIN) lesions and cervical carcinomas. A high percentage of Ki-67-Ag positive cells was seen in the parabasal cells of normal ectocervical and mature squamous metaplastic epithelium, although the basal cells were virtually negative. In preneoplastic lesions, however, the basal cells showed high proliferative activity and an increasing frequency of Ki-67-Ag positive cells was observed in the higher epithelial layers with increasing severity of CIN. In squamous cell carcinomas, variable numbers of Ki-67-Ag positive cells were observed and in adenocarcinomas expression increased with the degree of anaplasia. bcl-2 expression was observed only in the basal cells of normal endo- and ectocervix including reserve cells. With increasing severity of CIN, staining intensity and number of bcl-2 positive cells gradually increased. Five of eight squamous cell carcinomas were variably positive. All five adenocarcinomas showed extensive bcl-2 expression. Increased expression of both Ki-67-Ag and bcl-2 with increasing severity of CIN indicates an increasing imbalance between cell proliferation and protection from apoptosis. It is therefore proposed that an increasing proliferative fraction combined with a higher number of cells protected from apoptotic cell death contributes to progression of CIN. This phenotype may identify premalignant lesions with the potential to transform to cervical cancer.

BCL-2 immunoreactivity increases with severity of CIN: a study of normal cervical epithelia, CIN, and cervical carcinoma.

The presence of the BCL-2 protein, a marker for inhibition of programmed cell death, was studied in a series of routinely processed cervical tissues, consisting of normal endocervical (n = 40) and ectocervical epithelium (n = 27), squamous metaplastic epithelium (n = 30), CIN (cervical intraepithelial neoplasia) lesions (n = 32), and cervical carcinomas (n = 13). BCL-2 was strongly expressed in the basal cell compartment of normal ectocervical squamous epithelium and in nearly all reserve cells, while in endocervical columnar cells it was moderately expressed. In immature squamous metaplastic epithelium, BCL-2 expression varied. Half of the cases showed only basal cell staining, while the other half showed staining also in suprabasal layers. BCL-2 could be detected in all premalignant lesions, showing a striking increase in the number of positive cells with increasing severity of CIN, in combination with a mild increase in staining intensity. All adenocarcinomas were positive (n = 5), while five of eight squamous cell carcinomas expressed BCL-2. Based on these results, it is hypothesized that both the larger number of cells staining with BCL-2 in higher grades of CIN and the increase in staining intensity imply an increasing protection of these neoplastic conditions against programmed cell death. This protection facilitates not only continuing proliferation, but also the induction of genetic instability in dysplastic epithelial cells; it may thus reflect the greater capacity of the more severe CIN lesions to evolve into cervical carcinoma.

The mesangium in anti-Thy-1 nephritis. Influx of macrophages, mesangial cell hypercellularity, and macromolecular accumulation.

Mesangial pathology is a hallmark of focal and segmental glomerulosclerosis (FGS). In an immunologically mediated mesangial cell (MC) injury model, we analyzed the relationship between mesangial hypercellularity, increased macromolecular uptake by the mesangium, and long-term pathologic sequelae. A single injection of monoclonal anti-Thy-1 (AT) antibody induces MC apoptosis, extensive mesangiolysis, proteinuria, MC proliferation, and hypercellularity. Immunohistologic analysis indicated influx of ED 1-positive macrophages after 24 hours, which gradually subsiding thereafter. At day 12, hypercellularity was due to smooth musclelike MCs, and macrophagelike MCs were absent. Injection of iron dextran in nephritic rats indicated that mesangial uptake of iron correlated with mesangial hypercellularity, but was independent of proteinuria. Long-term studies showed no difference after 19 weeks in FGS between nephritic and control rats. In conclusion, although mesangiolysis is accompanied by influx of macrophages, a phase of smooth musclelike MC proliferation and increased macromolecular uptake, these pathologic events do not result in chronic mesangial pathology and FGS.

A monoclonal antibody against rat platelets. I. Tissue distribution in vitro and in vivo.

In this study we describe a new monoclonal antibody (MoAb PL.1) against rat platelets. Immunohistology of various rat tissues showed staining of platelets, especially in the spleen, and staining of megakaryocytes in bone marrow and spleen red pulp. In the liver small platelet aggregates and endothelial cells were stained. After in-vivo administration of MoAb PL.1 an acute severe thrombocytopenia was observed. In general the distribution of the antibody and/or antibody-coated platelet aggregates showed the same pattern as after in-vitro incubation, i.e. staining of rat platelets and platelet aggregates in spleen red pulp, and staining of megakaryocytes in spleen and bone marrow. Platelet aggregates were observed in the liver and electron microscopy indicated that they were associated with Kupffer cells. Furthermore, liver endothelial cells were positively stained. Comparison of the molecular weight of the antigens recognized by this MoAb and by human anti-platelet MoAbs, as well as comparison of staining patterns of megakaryocytes indicated that MoAb PL.1 is probably directed to a GPIIb/IIIa complex analogue. Since MoAb PL.1 is of the non-complement-binding mouse IgG1 isotype, it can be used for studying clearance of platelet aggregates by Fc-receptors of the MPS. It also promises to be a useful tool in the study of platelet involvement in rats with experimental nephritis.