In situ amplification of 5-lipoxygenase and 5-lipoxygenase-activating protein in allergic airway inflammation and inhibition by leukotriene blockade.

Leukotrienes are important mediators of the eosinophilic influx and mucus hypersecretion in the lungs in a murine model of asthma. We used in situ PCR in this model of human asthma to detect lung mRNA for 5-lipoxygenase (5-LO) and 5-LO-activating protein (FLAP), key proteins necessary for leukotriene synthesis. Lung tissue was obtained on day 28 from mice treated with i.p. (days 0 and 14) and intranasal (days 14, 25, 26, and 27) OVA or saline. After fixation, the tissue sections underwent protease- and RNase-free DNase digestion, before in situ RT-PCR using target-specific cDNA amplification. 5-LO and FLAP-specific mRNA was visualized by a digoxigenin detection system, and positive cells were analyzed by morphometry. 5-LO and FLAP-specific mRNA and protein were associated primarily with eosinophils and alveolar macrophages in the airways and pulmonary blood vessels in OVA-sensitized/challenged mice. 5-LO and FLAP protein expression increased on a per-cell basis in alveolar macrophages of OVA-treated mice compared with saline controls. Pulmonary blood vessel endothelial cells were also positive for 5-LO, FLAP mRNA, and protein. 5-LO inhibition significantly decreased 5-LO and FLAP-specific mRNA and protein expression in the lung inflammatory cells and endothelial cells. These studies demonstrate a marked increase in key 5-LO pathway proteins in the allergic lung inflammatory response and an important immunomodulatory effect of leukotriene blockade to decrease 5-LO and FLAP gene expression.

Breast tumor characteristics as predictors of mammographic detection: comparison of interval- and screen-detected cancers.

BACKGROUND: Although mammographic screening is useful for detecting early breast cancer, some tumors are detected in the interval between screening examinations. This study attempted to characterize fully the tumors detected in the two different manners. METHODS: Our study utilized a case-control design and involved a cohort of women undergoing mammographic screening within the defined population of a health maintenance organization (the Group Health Cooperative of Puget Sound). Women were classified as having "interval" or "interval-detected" cancers (n = 150) if their diagnosis was made within 24 months after a negative-screening mammogram or one that indicated a benign condition. Cancers were classified as "screen detected" (n = 279) if the diagnosis occurred after a positive assessment by screening mammography. Tumors from women in each group were evaluated for clinical presentation, histology, proliferative characteristics, and expression of hormone receptors, p53 tumor suppressor protein, and c-erbB-2 protein. RESULTS: Interval-detected cancers occurred more in younger women and were of larger tumor size than screen-detected cancers. In unconditional logistic regression models adjusted for age and tumor size, tumors with lobular (odds ratio [OR] = 1.9; 95% confidence interval [CI] = 0.9-4.2) or mucinous (OR = 5.5; 95% CI = 1.5-19.4) histology, high proliferation (by either mitotic count [OR = 2.9; 95% CI = 1.5-5.7] or Ki-67 antigen expression [OR = 2.3; 95% CI = 1.3-4.1]), high histologic grade (OR = 2.1; 95% CI = 1.2-4.0), high nuclear grade (OR = 2.0; 95% CI = 1.0-3.7), or negative estrogen receptor status (OR = 1.8; 95% CI = 1.0-3.1) were more likely to surface in the interval between screening examinations. Tumors with tubular histology (OR = 0.2; 95% CI = 0.0-0.8) or with a high percentage of in situ components (50%) (OR = 0.5; 95% CI = 0.2-1.2) were associated with an increased likelihood of screen detection. CONCLUSIONS: Our data from a large group of women in a defined population indicate that screening mammography may miss tumors of lobular or mucinous histology and some rapidly proliferating, high-grade tumors.

Telomerase activity and expression of telomerase RNA component and telomerase catalytic subunit gene in cervical cancer.

Telomerase, a ribonucleoprotein complex that includes the telomerase RNA component (hTR) and the telomerase catalytic subunit gene (hTERT) product, has been shown to be activated in the majority of cancer tissues and immortalized cells. To study telomerase activation during the progression of cervical cancer, the expression of hTR and hTERT RNAs in tissues of various stages of cervical cancer was analyzed using the in situ hybridization method and compared with proliferative activity as estimated by Ki-67 immunostaining. To test whether expression of these components is reflected in enzyme activity, we determined the levels of the RNAs in cervical cancer and normal tissues and in primary and immortal keratinocytes by reverse transcription-polymerase chain reaction and RNase protection assays and compared the results to telomerase activities as detected by telomeric repeat amplification protocol assay. In situ hybridization signals of hTR and hTERT were present not only in carcinoma tissues but also in normal epidermal layers. In many adenocarcinoma and fewer squamous cell carcinoma tissues, both signals were focally increased where high proliferative activity was present at the stages of dysplasia/metaplasia, in situ carcinoma, and invasive carcinoma. The level of bTERT, as quantitated by RNase protection assay, was not different between cancer and control tissues or immortal and a subset of primary keratinocytes and did not correlate with telomerase activity. These results indicate that expression of hTR and bTERT is up-regulated in at least a subset of neoplastic cells at an early stage of carcinogenesis and that unidentified factors, such as the modulation or coordination of its protein level with other products, may contribute to the activation of telomerase in cervical cancer.