Differences in AgNOR quantity between colorectal cancer and corresponding metastases: are they useful for prognostic purposes?

In order to evaluate the relationship between primary colorectal adenocarcinomas and their metastases with regard to AgNOR proteins expression, we have performed a morphometric analysis of AgNOR silver precipitates (AgNORs) in advanced colorectal carcinomas (stage III-IV) as well as in corresponding lymph node and distant metastases. Surgical specimens studied consisted of 64 primary tumours, 56 lymph node and 43 hepatic or omental metastases; for each formalin-fixed paraffin-embedded specimen, silver staining according Ploton et al. (1986) was performed and the AgNOR area (NORA) relative to primary site, lymph node and/or distant metastases was calculated. A significant difference (P < 0.001) among the non-neoplastic colonic control mucosae and primary or metastatic neoplastic lesions was found, but no statistically significant differences were encountered among the categories of primary colorectal carcinomas and their lymph node or distant metastases. Moreover, on the basis of NORA data case per case, three groups of patients were selected. In particular, in the group 1 NORA values of metastases were higher than those of the corresponding primary carcinomas, while in the group 2, NORA values of metastases were lower than those of the corresponding primary neoplasms; no differences of NORA values were encountered in group 3. The cumulative patient survival estimated by the Kaplan-Meier method showed a worse prognosis for patients belonging to group 1, where higher AgNOR values, not only in primary carcinomas but also in lymph node and distant metastases, were found.

Immunohistochemical expression of metallothionein in normal human colorectal mucosa, in adenomas and in adenocarcinomas and their associated metastases.

The immunohistochemical distribution pattern of metallothionein, a low molecular weight protein with strong affinity for divalent heavy metal ions, has been investigated in normal and neoplastic conditions of the large bowel. Utilizing a monoclonal mouse antibody the following formalin-fixed paraffin-embedded surgical or biopsy samples were studied: tubulo-villous adenomas (8 cases); adenocarcinomas with various degree of differentiation (85), nine of which were mucinous-type; synchronous tubular or tubulo-villous adenomas separate from carcinomas (30); transitional mucosa (45); metastases in lymph nodes (43); and distant metastases (45). Twenty biopsies from the right and left colon of 10 patients affected by irritable bowel syndrome were also analyzed. Normal colonic mucosa as well as transitional mucosa showed metallothionein immunopositivity in enterocytes at the luminal surface and crypts. Evident nuclear and cytoplasmic staining was encountered in tubulo-villous adenomas; the same reactivity was noted in the basal glandular component of colorectal carcinomas-synchronous adenomas, while less intense staining was noted in the apical villous portions. A variable metallothionein immunostaining was observed in adenocarcinomas (62.3%), in lymph node (55.8%) and distant hepatic (17.2%) and omental (43.8%) metastases, although it was not always concordant with that reported in the corresponding primary tumour. Whether the metallothionein positivity observed in normal and neoplastic cells is the result of expression of a stable form of the protein or an accumulation in the nucleus and cytoplasm remains to be clarified.

Gallbladder carcinoma: a video image analysis of AgNOR distribution and its relation to tumour stage and grade.

The amount and distribution of interphase argyrophilic nucleolar organizer regions (AgNORs) was studied in 51 gallbladder surgical specimens including 32 primary carcinomas, 10 adenomas and 9 cases of chronic cholecystitis with calculi. The mean nuclear AgNOR area (NORA) and the AgNOR distribution score (NORDS), i.e. the percentage of cells carrying nucleolar aggregates with more than 6 distinct silver dots, were evaluated in 200 epithelial nuclei per specimen by means of automated image analysis and direct counting respectively. Statistical analysis (variance analysis and Student-Neuman-Keuls' test) performed on the pooled mean AgNOR values showed a significant difference (p < 0.001) between carcinomas and non-carcinomatous lesions. Both NORA and NORDS highly discriminated carcinomas with histopathological grade 4 versus cases with grade 1, 2 or 3 (p < 0.001); a less statistically significative p value (< 0.05) was encountered when NORDS values of well differentiated grade 1 carcinomas and adenomas were compared. The NORA parameter permitted the discrimination of stage IV versus stage I carcinomas (p < 0.001), while carcinomas in stage IV and those in stage II were distinguished with a p < 0.05; the NORDS parameter allowed also to distinguish stage IV from both stage I or II tumours (p < 0.001). Our results indicate that the above-mentioned AgNOR parameters may be utilized as additional, more objective quantitative criteria in the clinical-pathological assessment of the outcome of gallbladder carcinomas.

Lazaroid U-74389F attenuates phorbol ester-induced lung injury in rabbits.

We examined the effect of the antioxidant lazaroid U-74389F on acute lung injury induced in rabbits by phorbol myristate acetate (PMA). Thirty minutes after receiving either U-74389F (15 mg.kg-1 i.v.) or U-74389F vehicle, rabbits (n = 60) were given PMA (60 micrograms.kg-1 i.v.). PMA vehicle injected rabbits (n = 20) served as controls. Over a 5 h period after PMA or PMA vehicle injection, we measured arterial pH, arterial oxygen tension (Pa,O2), arterial carbon dioxide tension (Pa,CO2), and the plasma concentration of the neutrophil chemoattractant interleukin-8 (IL-8). At postmortem, lungs were inspected for macroscopic injury and examined histologically. Malondialdehyde levels were assayed in lung tissue as an index of lipid peroxidation. In bronchoalveolar lavage (BAL), total and differential cell counts, protein and IL-8 concentrations were measured. Compared to normal controls, rabbits challenged with PMA alone developed arterial acidosis, hypercapnia and hypoxaemia, accompanied by significant rise in plasma IL-8 concentration. U-74389F pretreated animals did not develop significant arterial blood gas abnormalities and had significantly lower IL-8 concentration in plasma. U-74389F did not prevent PMA-induced lipid peroxidation. However, macroscopic signs of lung injury and the degree of alveolar haemorrhage and protein extravasation were significantly less severe in pretreated rabbits than in those given PMA alone. In addition, U-74389F significantly reduced IL-8 concentration and neutrophil number in BAL. By histological assessment, 80% of lung neutrophils were localized in alveolar spaces of animals receiving PMA alone. Conversely, in U-74389F pretreated animals, 75% of neutrophils were distributed within extra-alveolar blood vessels and alveolar septa. We conclude that lazaroid U-74389F attenuates lung injury in rabbits given PMA by preventing neutrophil migration into pulmonary alveoli. This effect may, in part, be related to downregulation of IL-8 production.

The prevalence of the precancerous lesions in breasts contralateral to clinical cancer. A morphological comparison with breasts containing a benign lump.

Two small series (nine cases each) of human female breasts were collected to compare the morphological changes of mammary glandular trees contralateral to primary breast cancer and those collateral to symptomatic benign lump. Each whole mammary gland was analysed by a submacroscopic scrutiny method using a stereomicroscope. Interesting and suspicious samples were removed for routine histology. Benign subclinical lesions were indifferently present in both series: Spheric cysts (5:5), sclerosing adenosis (3:3), intraductal papillomas (1:1), fibroadenomas (3:1). On the contrary proliferative epithelial lobular lesions with various degree of atypia i.e. atypical lobules (Grades IV-V according to Wellings), were detected only in the first series (p < 0.01). These data agree completely with the hypothesis of a systemic nature of breast cancer and support indirectly the possible predictive value of atypical lobules in bioptic specimens for the subsequent development of cancer in collateral and/or contralateral breasts.

67Ga-citrate imaging in rhabdomyosarcoma.

A 22-year-old female patient with a swelling in the right forearm near the elbow underwent a 67Ga-citrate scan because of paraesthesia on the ulnar side of the right hand. The 67Ga total body scan showed intense focal uptake on the forearm in the same position as the swelling. At surgery a histological diagnosis of alveolar rhabdomyosarcoma was made. Five months later, a repeat 67Ga scan was normal. Eight months after the diagnosis, the patient complained of pain in the left lumbar region extending to the hypogastric area. A third 67Ga scan showed intense uptake near the spinal column which at surgery was found to be a metastasis of the primary tumor.

Comparison of monoclonal immunocytochemical and immunoenzymatic methods for steroid receptor evaluation in breast cancer.

The production of monoclonal antibodies against estrogen receptor (ER) and progesterone receptor (PR) has permitted the development of the enzyme immunoassay (EIA) and immunocytochemical assay (ICA) for steroid receptor determination. The results obtained with these two techniques, using the same monoclonal antibodies, were compared in a large series of breast carcinomas (187 for ER and 100 for PR). The correlation between these methods was significant for ER (rs = 0.54) and PR (rs = 0.55) (P less than 0.001) but was lost when the receptor concentrations determined by EIA were less than or equal to 15 and less than or equal to 30 fmol/mg protein for ER and PR, respectively. When these values are considered as cutoffs, the concordance between the two methods was 84.5% for ER and 73% for PR. An analysis of discordant results revealed that low epithelial cellularity generally was present in ICA-positive, EIA-negative specimens, whereas only focal positivity with ICA, or positivity of only normal peripheral mammary ducts and lobules, frequently was found in ICA-negative, EIA-positive tumors. In conclusion, there is good correlation between the results obtained by EIA and ICA methods for detection of ER and PR. The authors suggest that biochemical and histochemical methods for steroid receptors could be considered complementary and used together for the analysis of breast cancer.

Estrogen, progesterone receptors and proliferating activity evaluated by immunocytochemistry in breast cancer.

The correlation of the most important prognostic indicators was evaluated in 75 breast cancer cases. Estrogen-progesterone receptors and proliferating activity were analyzed by immunocytochemical methods (ER-ICA, PR-ICA, Ki-67). Both steroid receptors were inversely correlated with the proliferating activity (ER-ICA vs Ki-67, p less than 0.003; PR-ICA vs. Ki-67, p less than 0.0001). No correlation was found between steroid receptors or cell kinetics and tumor size or lymph node status. These findings confirm the relevance of biochemical and kinetic parameters as independent markers in breast cancer and suggest a routine use of the simple immunocytochemical methods in assessing the biological behavior of tumors.

Fibrocystic condition and "at risk" lesions in asymptomatic breasts: a morphologic study of postmenopausal women.

A series of postmenopausal women who had died without noticing any clinical breast disease in their anamnesis (100 cases, age range 46-90 years, average age 62 years) were submitted to bilateral subcutaneous mastectomy during autopsy in order to evaluate the morphologic profile of asymptomatic mammary glands, at different ages. Submacroscopic changes were found and removed to be processed for histology. Results were as follows: a) 46% of cases did not show any change; b) 54% of cases showed benign changes, namely a fibrocystic condition; c) 14% of cases had in addition epithelial lobular hyperplasia with low grade atypia and d) 3% of cases showed atypical borderline lobules (ABL), i.e., terminal ductal-lobular units characterized by severe epithelial atypia. Such lesions cannot be easily distinguished from "in situ" carcinoma, and are currently considered at morphologic risk for subsequent cancer when found in breast biopsies. Our data show that: 1) ABL do not represent a common finding in women who never complained of breast pathology during life; 2) ABL are not related to older age; 3) Fibrocystic condition is quite frequent at subclinical levels also in asymptomatic aging women. The latter statement confirms the opinion that fibrocystic condition should be considered as a common "functional" change. On the contrary, the rarity of ABL gives us a further indirect evidence of their possible precancerous significance. The risk of subsequent development of cancer from the collateral mammary gland could be theoretically higher when ABL are found in breast biopsies of fertile and premenopausal women, who have a longer period of life expectation.

Cancerous versus noncancerous breasts. A comparative morphological analysis of the entire glandular tree of the breast.

Cancerous and clinically normal autopsy obtained breasts were collected in order to compare the physiopathological profile of both types of glandular tree. Each breast was visualized by whole thin sections and observed under a stereomicroscope with removal of the more interesting changes for histology. The comparison was made between 67 atrophic cancerous breasts and 88 atrophic control breasts. The results were as follows: 25% of the cancerous breasts versus 47% of control breasts showed no changes, atypical lobules, microfoci of "in situ" and/or infiltrating cancer were present in 46% of cancerous breasts and in 16% of control breasts, showing a significant correlation with clinical cancer. All other types of functional and proliferative changes, variously associated each other, were found in 29% of cancerous and in 37% of control breasts. Our morphological data agree completely with the statements in follow-up studies carried out on benign breast biopsies. The significant differences in the physiopathological profile of the glandular tree between "normal" and cancerous breasts, confirms that some changes are causally related to clinical cancer.

Orientation and frequency dependence of backscatter coefficient in normal and pathological breast tissues.

The backscatter coefficient was measured on five groups of normal and pathological breast specimens: (1) as a function of frequency (in the range 4-14 MHz) and (2) at a single fixed frequency (10 MHz), as a function of the angle of incidence between the beam and the specimen (approximately 60 degrees). The results of the study are discussed in relation to the content of cells and collagen fibers in breast tissues. The absolute value of backscatter coefficient is larger in tissues with a prevalence of collagenous fibers in comparison to tissues with only cells. In fibrofatty tissue, the inhomogeneity of the specimens is probably responsible for the highest backscatter value. The power law frequency dependence of the backscatter coefficient is of a diffractive nature in tissues characterized by only cells; in tissues with a strong prevalence of collagenous fibers, the power law frequency dependence increases. Periodicities in the angular patterns have been quantified by the autocorrelation functions for each group of specimens. The results of the study suggest a means for assessing tissue structure in normal and pathological breast tissue.

The use of frequency histograms of ultrasonic backscatter amplitudes for detection of atherosclerosis in vitro.

This study was designed to determine whether a quantitative analysis of integrated backscatter amplitude distribution is potentially useful in characterizing the atherosclerotic lesion. One hundred measurements (10 X 10 array) were made in fresh aortic regions (2 cm X 2 cm) of nine normal and 19 atherosclerotic arterial walls. A 10 MHz transducer was used. The integrated backscatter distinguished normal from atherosclerotic specimens (-56.7 +/- 4.3 vs -42.5 +/- 8.9 dB, p less than .01). The shape of the integrated backscatter amplitude distribution was analyzed by calculation of skewness and kurtosis of each arterial region. Both skewness values (0.134 +/- 0.325 vs -0.193 +/- 0.491 in normal and atherosclerotic segments, respectively, p = NS) and kurtosis values (0.055 +/- 0.765 vs -0.610 +/- 0.379, p less than .01) discriminated between the two groups. When only the six atherosclerotic specimens with mostly fatty and fibrofatty sites were considered, skewness and kurtosis still distinguished normal from atherosclerotic regions (0.134 +/- 0.325 vs -0.404 +/- 0.232, p less than .05 and 0.055 +/- 0.765 vs -0.558 +/- 0.337, p less than .05, respectively), while integrated backscatter values did not (-56.7 +/- 4.5 vs -52.3 +/- 6.1 dB, p = NS). In conclusion, atherosclerosis may be detected in vitro by the quantitative analysis of integrated backscatter distribution. This variable could also be of help in the identification of less obvious forms of atherosclerotic disease that are not distinguishable on the basis of integrated backscatter amplitude.

Evaluation of frequency dependence of backscatter coefficient in normal and atherosclerotic aortic walls.

For five groups of aortic specimens (normals and with different degrees of atherosclerosis) the ultrasonic backscatter coefficient was measured as a function of frequency in the range 4-15 MHz. The results of the study are related to two classes of structure, connective and fatty tissue (whose relative amount in the arterial wall is related to the different stages of atherosclerosis), as the main determinants of the scattering from aorta. The structure of connective tissue in the aorta produces a power law frequency dependence of the backscatter coefficient typical of small scale structures (Ka much less than 1). Fatty tissue introduces a frequency dependence of the backscatter coefficient typical of structures of intermediate scale (Ka approximately equal to 1). Biochemical composition and structure of normal and atherosclerotic aorta therefore, specifically affect the employed acoustical parameter.

Fatal peripheral airway cystic disease in a young woman.

Diffuse cystic transformation of both lungs was rapidly fatal in a 33-year-old woman. This disorder, a rare congenital condition in infants, has apparently not been described in adulthood.

Multicentricity in breast cancer: a submacroscopic study.

One hundred human female breasts surgically removed by radical mastectomy for clinical cancer were analyzed under a dissecting microscope to search for other submacroscopic (less than 0.5 cm) foci collateral to and independent from the clinical one. These foci were found in 36 percent of cases, either in situ (17 percent), or infiltrating (12 percent), or both in situ and infiltrating (7 percent). This high percentage of multiple cancer confirms previous data, suggesting the wide intramammary distribution of breast cancer disease. Multicentricity appears to be significantly associated with the incidence of familial breast cancer (p less than 0.05) and advanced patient age in the range between 71 and 80 years (p less than 0.005). Axillary lymph node metastases are significantly associated with collateral foci of in situ cancer (p less than 0.025), but not with submacroscopic foci of infiltrating cancer and the general character of the mammary glandular parenchyma, tumor size or histologic type of the clinical neoplasm. The critical question is whether multicentricity makes a difference clinically and especially to women who do not have their entire breast removed. Radical mastectomy results in a severe cosmetic and functional problem for patients. According to many authors, the goal of the treatment should be the removal of breast cancer by conservative surgical techniques (lumpectomy, subcutaneous mastectomy, quadrantectomy), using adjuvant radiotherapy and/or chemotherapy. The use of radiotherapy as primary treatment of early breast cancer has been also suggested. There is disagreement about surgical management of breast cancer. In fact, some investigators emphasize that the natural biologic history of multicentric cancers has not been documented by any adequate follow-up series in women who do not have their entire breast removed. Thus far, no difference has been seen in disease-free or overall survival between groups of patients with early breast cancer treated by an alternative therapeutic procedure and patients treated by radical mastectomy. However, Veronesi et al. (1981) refer to 4 second primary tumors of the ipsilateral breast in the 352 cases of small cancers treated by quadrantectomy, axillary dissection, and adjuvant radiotherapy. Moreover, Hellman et al. (1980), using radiation therapy without mastectomy for the primary treatment of 176 patients with early breast cancer, found 1 case of new cancer in a separate quadrant. Further evaluation is necessary to establish the long-term results of the alternative treatments of breast cancer and for the understanding of the clinical significance of microscopic multifocal tumor in the mammary gland.

Atherosclerosis detection by ultrasounds. A comparative histologic study on aortic specimens.

The aim of the study was to establish whether ultrasonic tissue characterization may be an useful tool in atherosclerosis detection. Ultrasonic measurements were performed on fresh aortic specimens taken from autopsy. Four hundred aortic regions were studied in vitro and histologically classified in four groups of 100 samples each: normal walls, fibrous, fibro-fatty and calcified plaques. Two different indices were measured, one derived from the Fourier transform of the echo produced by a specular reflector placed behind the specimen under study and named integrated attenuation index (IAI); the other derived from the Fourier transform of the echo reflected from the specimen under study and named integrated backscatter index (IBI). Results obtained from ultrasonic and histologic analyses showed that both attenuation and backscatter indices increase in atherosclerotic specimens where a deposition of biochemical components such as collagen and calcium salts takes place. This is not true for fatty tissue that reduces the echogenicity of the atherosclerotic profile of the arterial wall.