Diagnostic Value of Silver-Stained Interphasic Nucleolar Organizer Regions in Breast Tumors

Seventy-six specimens of normal breast tissue and benign and malignant breast lesions were studied to assess the mean area occupied by silver-stained proteins of the nucleolar organizer regions (MNORA) of the nucleolus. The assessment was performed with a computer-assisted image analyzer. The results indicate that only 30% of malignant lesions have a MNORA value greater than that of normal breast tissue or benign lesions. On the other hand, MNORA values of ductal carcinoma in situ were significantly greater than those of epitheliosis (papillomatosis). MNORA values were also significantly different in grade I and grade III invasive ductal carcinomas, the latter exhibiting the highest MNORA values of all the cases observed. Evaluation of MNORA values may therefore help in differentiating benign epithelial proliferations from ductal carcinomas in situ. Furthermore, because there is evidence that MNORA values are indicative of the cell duplication rate, MNORA values may ultimately be considered an objective prognostic parameter in addition to grading for invasive ductal carcinomas.     

The expression of E-cadherin and catenins in colorectal tumours from familial adenomatous polyposis patients

Familial adenomatous polyposis patients (FAP) harbour a germline mutation of the adenomatous polyposis coli gene (APC), and APC mutations are early events in the development of sporadic colorectal neoplasms. The APC protein interacts with beta-catenin and gamma-catenin and APC mutations are believed to play a role in the altered levels of beta-catenin in colorectal tumours. Immunohistochemical studies have shown changes in the expression and distribution of E-cadherin and catenins in sporadic colorectal neoplasms. This study assessed the expression and distribution of E-cadherin and catenins in colorectal neoplasms and non-neoplastic mucosa from FAP patients. The expression and cellular distribution of E-cadherin and catenins were studied by immunohistochemistry in 61 adenomas, five carcinomas, and non-neoplastic mucosa from 18 FAP patients. mRNA levels in the carcinomas were studied by in situ hybridization. The expression of E-cadherin and catenins was increased in over 80% of the adenomas, with evident cytoplasmic immunoreactivity. There was increased expression of E-cadherin and catenin in the carcinomas, with a notable increase in the levels of mRNA, in comparison with the non-neoplastic mucosa.     

Latent transforming growth factor-beta activation in mammary gland: regulation by ovarian hormones affects ductal and alveolar proliferation

Transforming growth factor-beta1 (TGF-beta 1) is a pluripotent cytokine that can inhibit epithelial proliferation and induce apoptosis, but is also widely implicated in breast cancer progression. Understanding its biological action in mammary development is critical for understanding its role in cancer. TGF-beta 1 is produced as a latent complex that requires extracellular activation before receptor binding. To better understand the spatial and temporal regulation of its action during mammary gland development, we examined the pattern of activation in situ using antibodies selected to distinguish between latent and active TGF-beta. Activation was highly restricted. TGF-beta 1 activation was localized primarily to the epithelium, and within the epithelium it was restricted to luminal epithelial cells but absent from either cap or myoepithelial cells. Within the luminal epithelium, we noted a further restriction. During periods of proliferation (ie, puberty, estrus and pregnancy), which are stimulated by ovarian hormones, TGF-beta 1 activation decreased in some cells, consistent with preparation for proliferation. Paradoxically, other cells simultaneously increase TGF-beta 1 immunoreactivity, which suggests that TGF-beta 1 differentially restrains epithelial subpopulations from responding to hormonal signals to proliferate. These data suggest that endogenous TGF-beta 1 activation and thus activity are regulated by ovarian hormones. To determine the specific consequences of TGF-beta 1 activity, we manipulated TGF-beta 1 levels in vivo using Tgfbeta 1 knockout mice and undertook tissue recombination experiments with heterozygous tissue. In Tgfbeta 1 heterozygous mice, which have <10% wild-type levels of TGF-beta1, ductal development during puberty and alveolar development during pregnancy were accelerated, consistent with its role as a growth inhibitor. The proliferative index of Tgfbeta 1+/- epithelium was increased approximately twofold in quiescent tissue and fourfold in proliferating tissue but both ducts and alveoli were grossly and histologically normal. To test whether epithelial TGF-beta1 was critical to the proliferative phenotype, Tgfbeta 1+/+ and +/- epithelium were transplanted into +/+ mammary stroma. The outgrowth of Tgfbeta 1+/- epithelium was accelerated in wild-type hosts, indicating that the phenotype was intrinsic to the epithelium. Moreover, proliferation was 15-fold greater in Tgfbeta 1+/- than wild-type mice after ovariectomy and treatment with estrogen and progesterone, suggesting that TGF-beta 1 acts in an autocrine or juxtacrine manner to regulate epithelial proliferation. Together these data indicate that ovarian hormones regulate TGF-beta 1 activation, which in turn restricts proliferative response to hormone signaling.     

The cytochemical demonstration of oestrogen receptors in human breast carcinomas

A cytochemical method has been used to demonstrate oestrogen receptor in histological sections of breast carcinomas, and has given similar qualitative results to the biochemical dextran-coated charcoal method. Two conjugation techniques were employed to couple horseradish peroxidase to 17β-oestradiol-6-O-carboxymethyloxime—bovine serum albumin. The periodate conjugation method led to a greater degree of coupling but a higher concentration was required for demonstration of oestrogen receptor than that needed when the two-stage glutaraldehyde conjugation method was used, even though this resulted in a lesser degree of coupling. Care in the preparation of tissue has been found to be essential. Formalin-fixed, paraffin-embedded tissue was unsuitable; rapid freezing of fresh tissue, only brief air-drying or acetone fixation of tissue sections and short-term storage were important. A correlation has been shown between the presence of oestrogen receptor in breast carcinomas and good histological differentiation. The poorer differentiated tumours were also found to contain fewer reactive cells.     

Multiple organ engraftment by bone-marrow-derived myofibroblasts and fibroblasts in bone-marrow-transplanted mice

Myofibroblasts are ubiquitous cells with features of both fibroblasts and smooth muscle cells. We suggest that the bone marrow can contribute to myofibroblast populations in a variety of tissues and that this is exacerbated by injury. To assess this, female mice were transplanted with male bone marrow and the male cells were tracked throughout the body and identified as myofibroblasts. Skin wounding and paracetamol administration were used to assess whether myofibroblast engraftment was modulated by damage. Following radiation injury, a proportion of myofibroblasts in the lung, stomach, esophagus, skin, kidney, and adrenal capsule were bone-marrow derived. In the lung, there was significantly greater engraftment following paracetamol administration (17% versus 41% p < 0.005). Bone-marrow-derived fibroblasts were also found. We suggest that bone marrow contributes to a circulating population of cells and, in the context of injury, these cells are recruited and contribute to tissue repair.     

Progesterone control of interleukin-8 production in endometrium and chorio-decidual cells underlines the role of the neutrophil in menstruation and parturition

Interleukin-8 (IL-8) attracts neutrophils into tissues and causedthem to degtranulate. Both menstruation and parturition involveneutrophil migration into uterine tissues and therefore IL-8is a likely mediator of the tissue re-arrangements that accompanythese events. We have examined the ability of endometrium explantsand chorion cells in culture to synthesize and release IL-8and the ability of progesterone, a sunthetic progestin[medroxyprogesteroneacetate(MPA)] and dexamethasone to inhibit this production.In endo-metrium, the stage of the menstrual cycle didnot affectIL-8 production but a 10^–6 M concentration of progesteroneor dexamethasone significantly reduced the concentration ofIL-8 in medium after 24 H. After a further 24 h with lipopoly-saccharide(LPS) stimulation, only MPA and dexamethasone inhibited productionsignificantly. In chorion cells, IL-8 production was significantlydecreased by both MPA and dexamethasone in the LPS stimulatedcells but the reduction in the first 24 h was not significant.The IL-8 produced in uterine tissues might act synergisticallywith prostaglandin E (PGE), a likely site for this interactionbeing blood vessels where PGE production is also repressed byprogesterone. Such a co-operative action would maintain lowleykocyte entry into uterine tissues in the presence of progesteroneand falling steroid levels would induce leukocyte immigrationand activation with consequent tissue destruction. Such steroid-dependentinteractions are important in our understanding of the mechanismsof menstruation and paturition and could allow new approachesto the treatment of uterine pathology.     

Attenuation and speed of 10 MHz ultrasound in canine blood of various packed-cell volumes at 37°C

The attenuation coefficient and the speed of 10 MHz ultrasound were determined in canine blood at 37°C by a differential path-length technique. Blood specimens with packed-cell volumes (p.c.v) ranging from 0 to 53% were prepared by separating the cells from the plasma and mixing the two components. The mean attenuation coefficient increased linearly with packed-cell volume, the least squares regression function being α=0·992+0·039 PCV with a standard error of the estimate=0·255. The speed of 10 MHz ultrasound c in millimetres per second, increased with packed-cell volume, the regression function for a wave equation model being 1/c²=0·418+2·09×10⁻⁴ (PCV)−1·75×10⁻⁵ (PCV)² with a standard error of the estimate=0·0049 (mm/s)⁻². Both the attention coefficient and speed of 10 MHz ultrasound were greater in blood than in plasma to a degree dependent on the packed-cell volume.     

Clonal anergy to staphylococcal enterotoxin B in vivo: selective effects on T cell subsets and lymphokines

Injection of bacterial superantigens such as staphylococcal enterotoxin B (SEB) in adult mice results in initial proliferation of SEB-responsive Vβ8⁺ T cells followed by induction of a state of non-responsiveness frequently referred to as clonal anergy. We show here that SEB-induced anergy involves selective changes in lymphokine production and that it affects CD4⁺ Vβ 8⁺ and CD8⁺ Vβ 8⁺ T cells in different fashions. Whereas both CD4⁺ Vβ 8⁺ and CD8⁺ Vβ 8⁺ cells from anergic mice exhibit strongly reduced proliferative capacity and interleukin(IL)-2 production upon restimulation with SEB either in vivo or in vitro the CD8⁺ subset from SEB-injected mice produces other lymphokines (such as interferon(IFN)-γ) at normal or slightly increased levels in response to SEB. Changes in the levels of production of IL-2 and IFN-γ protein correlated well with mRNA accumulation both in vivo and in vitro. Collectively these data suggest that superantigen-induced anergy involves selective changes in signal transduction and/or gene regulation in T lymphocytes.