Brief Scientific Reports Cellularity in Breast Carcinoma ROBERT A. AMBROS, M.D. AND RONALD C. TROST, PH.D.
Tumor cellularities were observed in 23 consecutive cases of infiltrating ductal carcinoma of the breast and correlated with other morphometric, pathologic, and clinical features. The cellularity mean index (CMI), defined as the percentage of tumor volume occupied by invasive tumors cells, was found to correlate with lymph node involvement by tumor (P = 0.03) and the number of mitotic figures per ten high-power fields (P = 0.0002). The CMI also correlated with mitotic activity (P = 0.01) when the latter was expressed as the percentage of tumor cells actively in mitosis, thus correcting for differences in cellularity between individual tumors. The relationship between cellularity and mitotic activity is discussed. (Key words: Breast carcinoma; Tumor cellularity; Morphometry; Quantitative microscopy) Am J Clin Pathol 1990;93:98-100
Departments of Pathology, University of Medicine and Dentistry of New Jersey-New Jersey Medical School and University Hospital, Newark, New Jersey
Morphometric Analysis DESMOPLASIA, the formation of excessive extracellular matrix surrounding invasive tumor cells,9 is frequently significant in human breast carcinoma.1,7 The reciprocal of desmoplasia, tumor cellularity, has been shown to distinctly vary between individual cases of breast carcinoma. 12 Relationships between cellularity and other clinical and pathologic features, however, have not been defined. In the current report, tumor cellularity is determined and correlated with clinical, pathologic, and other morphometric features in 23 cases of infiltrating ductal breast carcinoma.
For morphometric analysis, slides stained with hematoxylin and eosin containing the most primary tumor were selected. An effort was made to select sections of both the tumor periphery and center. An average of two sections per case were used. In cases in which an excisional biopsy was performed before mastectomy, sections from the former were selected over sections of the residual tumor, provided the amount of tissue was sufficient for morphometric analysis. Measurements were performed with ocular grids and calibrations made by a slide micrometer. Fields of study were selected by setting the horizontal and vertical axes of the microscope stage according to random number tables.
Materials and Methods Case Selection
Percent Tumor Volume
Twenty-three consecutive cases of infiltrating ductal carcinoma in which a modified radical mastectomy was performed and the tumor examined for estrogen and progesterone receptors by biochemical assay were reviewed from the files of the Department of Pathology of University Hospital. Each case was reexamined and surgical pa-
The point-counting technique 313 was used to estimate the percentage of the tumor occupied by tumor cells. With the use of a 100-point ocular grid, the percentage of points superimposing tumor cell nuclei in 60 fields of view was recorded at a magnification of 150X. Only fields entirely occupied by tumor were used in analysis. A record was kept as to whether the tumor cell represented part of an in situ nest of tumor cells including intraductal carcinoma and cancerization of lobules' or invasive carcinoma. The percentage of tumor volume occupied by invasive tumor cell nuclei is referred to in this report as the cellularity mean index.
Received March 1, 1989; received revised manuscript and accepted for publication June 1, 1989. Address reprint requests to Dr. Trost: Department of Pathology, UMDNJ-New Jersey Medical School, 185 South Orange Avenue, Newark, New Jersey 07103.
98
Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016
thology reports reviewed. Lymph node metastases were identified in 13 of the 23 cases. Estrogen and progesterone receptor studies, performed by methods previously described,6'10 were positive (values greater than 10 fmol/mg of tissue) in nine cases and concordant in five cases.
Vol. 93 • No. I
BRIEF SCIENTIFIC REPORTS
Nuclear Parameters Under oil immersion and at a magnification of 1,000X, the largest axis and a perpendicular axis were measured from 25 nuclei by the selection technique described above. The nuclear area was then estimated by multiplying the greater by the lesser axis.4 The estimated nuclear area represented the mean value from the 25 nuclei studied. Mitotic Index
cells per field =
Table 2. Correlations Between the Cellularity Mean Index and Other Features Variable
r2
P Value
Age Tumor size Lymph node status ER receptor value* PR receptor value* Mean NA Mitotic index % Mitosisf
0.0681 0.0375 0.2087 0.0964 0.1067 0.0517 0.4806 0.2567
NS NS 0.03 NS NS NS 0.0002 0.01
NA = nuclear area; ER = estrogen; PR = progesterone; NS = not significant. * fmol/mg. | Estimated percentage of cells in mitosis. number of mitoses was corrected for differences in cellularity between tumors and expressed as the percentage of cells actively in mitosis (r2 = 0.2567). No correlation was found between the mitotic index and the percentage of tumor volume when the latter was limited to the in situ component (Table 3), nor were correlations found between cellularity and age, tumor size, receptor status, and mean nuclear areas.
area of field X percent tumor volume estimated nuclear area
Correlations between variables were tested by linear regression models. P values of < 0.05 were considered statistically significant. Results The percentage of tumor volume occupied by invasive carcinoma, i.e., the cellularity mean index, was found to vary from 4 to 82%, with a mean of 32% (Table 1). This index correlated with both lymph node metastasis and mitotic activity (Table 2). The latter correlation was detected when mitotic activity was expressed as the mitotic index (r2 = 0.4806) and remained significant when the
Table 1. Clinical and Morphometric Values in 23 Cases of Infiltrating Ductal Carcinoma Variable
Mean
Range
Age (years) Tumor size (cm) ER receptors (fmol/mg) PR receptors (fmol/mg) Percent tumor volume In situ and invasive Invasive only (CMI) Mean NA* Mitotic index % Mitosisf
49.0 3.8 14.4 25.0
32-79 0.9-7.0 0-66 0-138
35.8 32.0 65.6 5.7 0.06
6-82 4-82 23-102 0-30 0-0.31
Discussion This study reports the association between tumor cellularity and other pathologic and clinical features in ductal carcinoma of the breast. A correlation was found between mitotic activity and percent volume of invasive but not in situ carcinoma. This relationship was aiso identified when mitotic activity was corrected for the number of tumor cells per field of view and expressed as the percentage of cells in mitosis. Although it had been postulated more than 25 years ago that tumor cells regulate their connective tissue component in breast carcinoma,8 factors that could influence the amount of desrrioplasia have not been described. The findings in the current report indicate that desmoplastic reaction in a given tumor is directly correlated with the tumor's inherit mitotic activity, i.e., Table 3. Correlations Between Mitotic Index and Clinical and Morphometric Features Variable
CMI = cellularity mean index; NA = nuclear area. * pm2. t Estimated percentage of cells in mitosis.
Age Tumor size Lymph node status ER receptor value* PR receptor value* Percent tumor volume In situ Invasive (CMI) Mean NA
P Value 0.0681 0.0022 0.1735 0.1392 0.0990
NS NS 0.04 NS NS
0.0729 0.4805 0.1026
NS 0.002 NS
NA = nuclear area; CMI = cellularity m index; ean ER = estrogen; P=R progesterone; •• NS = not significant. • fmol/mg.
Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016
The mitotic index represented the number of mitoses in ten high-power fields at a magnification of 400X. Fields of view were selected as described by Baak3; the count was started in what was subjectively thought to be the most active area and the remaining nine fields selected at random. Because the quantitative value of this index is dependent on the number of cells per field of view, an additional parameter expressing the percentage of tumor cells in mitosis was obtained by dividing the mean number of mitoses per field of view by the number of cells per field. The number of tumor cells per field of view was estimated by the following formula:
99
100
AMBROS AND TROST
References I. Azzopardi JG. Problems in breast pathology. In: Major problems in pathology, vol 11. London: WB Saunders, 1979;203-213, 244245.
2. Baak JPA,"Van Dop H, Kurver PHJ, Hermans J. The value of morphometry to classic prognosticators in breast cancer. Cancer 1985;56:374-382. 3. Baak JPA, Oort J. A manual of morphometry in diagnostic pathology. Berlin: Springer-Verlag, 1983. 4. Bhattacharjee DR, Harris M, Faragher EB. Nuclear morphometry of epitheliosis and intraductal carcinoma of the breast. Histopathology 1985;9:511-516. 5. Black R, Prescott R, Bers K, Hawkins A, Stewart H, Forrest P. Tumor cellularity oestrogen receptors and prognosis in breast cancer. Clinical Oncology 1983;9:311-318. 6. Clark GM, Gary M, Mcquire WL, et al. Progesterone receptors as a prognostic factor in stage II breast cancer. N Engl J Med 1983;309:22; 1343-1346. 7. Groniowski J, Walski M, Pietrow D. Observations on collagenous stroma formation and desmoplastic breast carcinoma. Annals of the Medical Section of the Polish Academy of Sciences 1975;20: 81-84. 8. Gullino PM, Grantham FH. The influence of the host and the neoplastic cell population on the collagen amount of a tumor mass. Cancer Res 1963;23:648-653. 9. Liotta LA, Rao CN, Barsky SH. Tumor invasion and the extracellular matrix. Lab Invest 1983;49:636-648. 10. Mcquire WL, De La Garza M. Improved sensitivity in the measurement of estrogen receptor in human breast cancer. J Clin Endocrinol Metab 1973;37:986-989. 11. Parham DM, Robertson AJ, Brown RA. Morphometric analysis of breast carcinoma: association with survival. J Clin Pathol 1988;41: 173-177. 12. Underwood JCE. A morphometric analysis of human breast carcinoma. Br J Cancer 1972;26:234-237. 13. Weibull ER. Principles and methods for the morphometric study of the lungs and other organs. Lab Invest 1963;12:131-155.
Fine-Needle Cytology of 292 Palpable Orbital and Eyelid Tumors ANTOINE ZAJDELA, M.D., PHILIPPE VIELH, M.D., PIERRE SCHLIENGER, M.D., AND CHRISTIAN HAYE, M.D.
Fine-needle cytology (FNC) of 292 palpable orbital and eyelid tumors was performed with a 25-gauge (0.5-mm), 3-cm needle and compared with the histopathologic findings in 286 cases. Among these 286 cases, a concordant diagnosis of malignancy and type was achieved in 249 cases (87%). False positive diagnoses were made in four cases (1.6%) and false negative diagnoses infivecases (1.8%). No complications were encountered. These results led to the conclusion that FNC is an accurate tool in the diagnosis of orbital and eyelid tumors, especially when sampling and interpretation are performed by an experienced pathologist. (Key words: Cytology; Orbital; Eyelid; Tumors) Am J Clin Pathol 1990;93:100-104 DURING the last 15 years, cytology has become an important diagnostic method in ophthalmology,15 and some
Received March 13, 1989; accepted for publication April 19, 1989. Address reprint requests to Dr. Zajdela: Institut Curie, 26 rue d'Ulm, 75231 Paris Cedex, 05 France.
Departments of Cytopathology and Radiotherapy, and the Ophthalmology Consultation, Institut Curie, Paris, France
studies 8 ' 91618-20 have reported the reliability of fine-needle cytology (FNC) in the evaluation of deep and superficial orbital tumors. This simple procedure allows a valuable assessment before surgery; a high rate of diagnostic accuracy exists when there is a close working relationship between clinician and pathologist. In addition, the use of a very fine needle, such as a 25-gauge (0.5 mm), 3-cm needle provides an adequate cytologic sample, with minimal patient discomfort. The aim of this study is to present our ten-year experience in the diagnosis of palpable orbital and eyelid tumors by FNC and to discuss possibilities and limitations of this procedure.
Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016
tumors with a high proliferative activity being associated with low stromal volume. Although a cause-effect relationship cannot be established in our study, a hypothesis can be postulated in which the degree of desmoplastic reaction is dependent on the proliferative activity of the neoplasm, i.e., the stroma "plays catch up" to the neoplastic growth. A cause-effect relationship may possibly be determined by animal models. The effect of cellularity on prognosis has not been determined. High cellularity has been associated with both good5 and poor" prognosis. Although patient outcome was not evaluated in our study, the correlation between high cellularity and both lymph node metastases and mitotic activity, a feature linked to outcome, supports findings of high cellularity to be an indicator of poor prognosis. In summary, this study confirms the previous observation that breast carcinomas vary significantly in cellularity12 and suggests a strong correlation between cellularity and mitotic activity, a feature that has been strongly linked to patient outcome. 2
A.J.C.P.. January 1990
Tumor cellularities were observed in 23 consecutive cases of infiltrating ductal carcinoma of the breast and correlated with other morphometric, pathologic, and clinical features. The cellularity mean index (CMI), defined as the percentage of tumor volume occupied by invasive tumors cells, was found to correlate with lymph node involvement by tumor (P = 0.03) and the number of mitotic figures per ten high-power fields (P = 0.0002). The CMI also correlated with mitotic activity (P = 0.01) when the latter was expressed as the percentage of tumor cells actively in mitosis, thus correcting for differences in cellularity between individual tumors. The relationship between cellularity and mitotic activity is discussed. (Key words: Breast carcinoma; Tumor cellularity; Morphometry; Quantitative microscopy) Am J Clin Pathol 1990;93:98-100
Departments of Pathology, University of Medicine and Dentistry of New Jersey-New Jersey Medical School and University Hospital, Newark, New Jersey
Morphometric Analysis DESMOPLASIA, the formation of excessive extracellular matrix surrounding invasive tumor cells,9 is frequently significant in human breast carcinoma.1,7 The reciprocal of desmoplasia, tumor cellularity, has been shown to distinctly vary between individual cases of breast carcinoma. 12 Relationships between cellularity and other clinical and pathologic features, however, have not been defined. In the current report, tumor cellularity is determined and correlated with clinical, pathologic, and other morphometric features in 23 cases of infiltrating ductal breast carcinoma.
For morphometric analysis, slides stained with hematoxylin and eosin containing the most primary tumor were selected. An effort was made to select sections of both the tumor periphery and center. An average of two sections per case were used. In cases in which an excisional biopsy was performed before mastectomy, sections from the former were selected over sections of the residual tumor, provided the amount of tissue was sufficient for morphometric analysis. Measurements were performed with ocular grids and calibrations made by a slide micrometer. Fields of study were selected by setting the horizontal and vertical axes of the microscope stage according to random number tables.
Materials and Methods Case Selection
Percent Tumor Volume
Twenty-three consecutive cases of infiltrating ductal carcinoma in which a modified radical mastectomy was performed and the tumor examined for estrogen and progesterone receptors by biochemical assay were reviewed from the files of the Department of Pathology of University Hospital. Each case was reexamined and surgical pa-
The point-counting technique 313 was used to estimate the percentage of the tumor occupied by tumor cells. With the use of a 100-point ocular grid, the percentage of points superimposing tumor cell nuclei in 60 fields of view was recorded at a magnification of 150X. Only fields entirely occupied by tumor were used in analysis. A record was kept as to whether the tumor cell represented part of an in situ nest of tumor cells including intraductal carcinoma and cancerization of lobules' or invasive carcinoma. The percentage of tumor volume occupied by invasive tumor cell nuclei is referred to in this report as the cellularity mean index.
Received March 1, 1989; received revised manuscript and accepted for publication June 1, 1989. Address reprint requests to Dr. Trost: Department of Pathology, UMDNJ-New Jersey Medical School, 185 South Orange Avenue, Newark, New Jersey 07103.
98
Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016
thology reports reviewed. Lymph node metastases were identified in 13 of the 23 cases. Estrogen and progesterone receptor studies, performed by methods previously described,6'10 were positive (values greater than 10 fmol/mg of tissue) in nine cases and concordant in five cases.
Vol. 93 • No. I
BRIEF SCIENTIFIC REPORTS
Nuclear Parameters Under oil immersion and at a magnification of 1,000X, the largest axis and a perpendicular axis were measured from 25 nuclei by the selection technique described above. The nuclear area was then estimated by multiplying the greater by the lesser axis.4 The estimated nuclear area represented the mean value from the 25 nuclei studied. Mitotic Index
cells per field =
Table 2. Correlations Between the Cellularity Mean Index and Other Features Variable
r2
P Value
Age Tumor size Lymph node status ER receptor value* PR receptor value* Mean NA Mitotic index % Mitosisf
0.0681 0.0375 0.2087 0.0964 0.1067 0.0517 0.4806 0.2567
NS NS 0.03 NS NS NS 0.0002 0.01
NA = nuclear area; ER = estrogen; PR = progesterone; NS = not significant. * fmol/mg. | Estimated percentage of cells in mitosis. number of mitoses was corrected for differences in cellularity between tumors and expressed as the percentage of cells actively in mitosis (r2 = 0.2567). No correlation was found between the mitotic index and the percentage of tumor volume when the latter was limited to the in situ component (Table 3), nor were correlations found between cellularity and age, tumor size, receptor status, and mean nuclear areas.
area of field X percent tumor volume estimated nuclear area
Correlations between variables were tested by linear regression models. P values of < 0.05 were considered statistically significant. Results The percentage of tumor volume occupied by invasive carcinoma, i.e., the cellularity mean index, was found to vary from 4 to 82%, with a mean of 32% (Table 1). This index correlated with both lymph node metastasis and mitotic activity (Table 2). The latter correlation was detected when mitotic activity was expressed as the mitotic index (r2 = 0.4806) and remained significant when the
Table 1. Clinical and Morphometric Values in 23 Cases of Infiltrating Ductal Carcinoma Variable
Mean
Range
Age (years) Tumor size (cm) ER receptors (fmol/mg) PR receptors (fmol/mg) Percent tumor volume In situ and invasive Invasive only (CMI) Mean NA* Mitotic index % Mitosisf
49.0 3.8 14.4 25.0
32-79 0.9-7.0 0-66 0-138
35.8 32.0 65.6 5.7 0.06
6-82 4-82 23-102 0-30 0-0.31
Discussion This study reports the association between tumor cellularity and other pathologic and clinical features in ductal carcinoma of the breast. A correlation was found between mitotic activity and percent volume of invasive but not in situ carcinoma. This relationship was aiso identified when mitotic activity was corrected for the number of tumor cells per field of view and expressed as the percentage of cells in mitosis. Although it had been postulated more than 25 years ago that tumor cells regulate their connective tissue component in breast carcinoma,8 factors that could influence the amount of desrrioplasia have not been described. The findings in the current report indicate that desmoplastic reaction in a given tumor is directly correlated with the tumor's inherit mitotic activity, i.e., Table 3. Correlations Between Mitotic Index and Clinical and Morphometric Features Variable
CMI = cellularity mean index; NA = nuclear area. * pm2. t Estimated percentage of cells in mitosis.
Age Tumor size Lymph node status ER receptor value* PR receptor value* Percent tumor volume In situ Invasive (CMI) Mean NA
P Value 0.0681 0.0022 0.1735 0.1392 0.0990
NS NS 0.04 NS NS
0.0729 0.4805 0.1026
NS 0.002 NS
NA = nuclear area; CMI = cellularity m index; ean ER = estrogen; P=R progesterone; •• NS = not significant. • fmol/mg.
Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016
The mitotic index represented the number of mitoses in ten high-power fields at a magnification of 400X. Fields of view were selected as described by Baak3; the count was started in what was subjectively thought to be the most active area and the remaining nine fields selected at random. Because the quantitative value of this index is dependent on the number of cells per field of view, an additional parameter expressing the percentage of tumor cells in mitosis was obtained by dividing the mean number of mitoses per field of view by the number of cells per field. The number of tumor cells per field of view was estimated by the following formula:
99
100
AMBROS AND TROST
References I. Azzopardi JG. Problems in breast pathology. In: Major problems in pathology, vol 11. London: WB Saunders, 1979;203-213, 244245.
2. Baak JPA,"Van Dop H, Kurver PHJ, Hermans J. The value of morphometry to classic prognosticators in breast cancer. Cancer 1985;56:374-382. 3. Baak JPA, Oort J. A manual of morphometry in diagnostic pathology. Berlin: Springer-Verlag, 1983. 4. Bhattacharjee DR, Harris M, Faragher EB. Nuclear morphometry of epitheliosis and intraductal carcinoma of the breast. Histopathology 1985;9:511-516. 5. Black R, Prescott R, Bers K, Hawkins A, Stewart H, Forrest P. Tumor cellularity oestrogen receptors and prognosis in breast cancer. Clinical Oncology 1983;9:311-318. 6. Clark GM, Gary M, Mcquire WL, et al. Progesterone receptors as a prognostic factor in stage II breast cancer. N Engl J Med 1983;309:22; 1343-1346. 7. Groniowski J, Walski M, Pietrow D. Observations on collagenous stroma formation and desmoplastic breast carcinoma. Annals of the Medical Section of the Polish Academy of Sciences 1975;20: 81-84. 8. Gullino PM, Grantham FH. The influence of the host and the neoplastic cell population on the collagen amount of a tumor mass. Cancer Res 1963;23:648-653. 9. Liotta LA, Rao CN, Barsky SH. Tumor invasion and the extracellular matrix. Lab Invest 1983;49:636-648. 10. Mcquire WL, De La Garza M. Improved sensitivity in the measurement of estrogen receptor in human breast cancer. J Clin Endocrinol Metab 1973;37:986-989. 11. Parham DM, Robertson AJ, Brown RA. Morphometric analysis of breast carcinoma: association with survival. J Clin Pathol 1988;41: 173-177. 12. Underwood JCE. A morphometric analysis of human breast carcinoma. Br J Cancer 1972;26:234-237. 13. Weibull ER. Principles and methods for the morphometric study of the lungs and other organs. Lab Invest 1963;12:131-155.
Fine-Needle Cytology of 292 Palpable Orbital and Eyelid Tumors ANTOINE ZAJDELA, M.D., PHILIPPE VIELH, M.D., PIERRE SCHLIENGER, M.D., AND CHRISTIAN HAYE, M.D.
Fine-needle cytology (FNC) of 292 palpable orbital and eyelid tumors was performed with a 25-gauge (0.5-mm), 3-cm needle and compared with the histopathologic findings in 286 cases. Among these 286 cases, a concordant diagnosis of malignancy and type was achieved in 249 cases (87%). False positive diagnoses were made in four cases (1.6%) and false negative diagnoses infivecases (1.8%). No complications were encountered. These results led to the conclusion that FNC is an accurate tool in the diagnosis of orbital and eyelid tumors, especially when sampling and interpretation are performed by an experienced pathologist. (Key words: Cytology; Orbital; Eyelid; Tumors) Am J Clin Pathol 1990;93:100-104 DURING the last 15 years, cytology has become an important diagnostic method in ophthalmology,15 and some
Received March 13, 1989; accepted for publication April 19, 1989. Address reprint requests to Dr. Zajdela: Institut Curie, 26 rue d'Ulm, 75231 Paris Cedex, 05 France.
Departments of Cytopathology and Radiotherapy, and the Ophthalmology Consultation, Institut Curie, Paris, France
studies 8 ' 91618-20 have reported the reliability of fine-needle cytology (FNC) in the evaluation of deep and superficial orbital tumors. This simple procedure allows a valuable assessment before surgery; a high rate of diagnostic accuracy exists when there is a close working relationship between clinician and pathologist. In addition, the use of a very fine needle, such as a 25-gauge (0.5 mm), 3-cm needle provides an adequate cytologic sample, with minimal patient discomfort. The aim of this study is to present our ten-year experience in the diagnosis of palpable orbital and eyelid tumors by FNC and to discuss possibilities and limitations of this procedure.
Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016
tumors with a high proliferative activity being associated with low stromal volume. Although a cause-effect relationship cannot be established in our study, a hypothesis can be postulated in which the degree of desmoplastic reaction is dependent on the proliferative activity of the neoplasm, i.e., the stroma "plays catch up" to the neoplastic growth. A cause-effect relationship may possibly be determined by animal models. The effect of cellularity on prognosis has not been determined. High cellularity has been associated with both good5 and poor" prognosis. Although patient outcome was not evaluated in our study, the correlation between high cellularity and both lymph node metastases and mitotic activity, a feature linked to outcome, supports findings of high cellularity to be an indicator of poor prognosis. In summary, this study confirms the previous observation that breast carcinomas vary significantly in cellularity12 and suggests a strong correlation between cellularity and mitotic activity, a feature that has been strongly linked to patient outcome. 2
A.J.C.P.. January 1990
