Administration of p.g. 600 to sows at weaning and the time of ovulation as determined by transrectal ultrasound.

This study determined whether the interval from estrus to ovulation was altered by giving P.G. 600 to sows at weaning. Mixed-parity sows received P.G. 600 i.m. (n = 72) or no treatment (n = 65) at weaning (d 0). Beginning on d 0, sows were observed for estrus twice daily. At the onset of estrus and thereafter, ultrasound was performed twice daily to determine the average size of the largest follicles and time of ovulation. Weaning age (20.1+/-0.4 d) did not differ (P > 0.10) between treatments. More P.G. 600 sows expressed estrus within 8 d (P < 0.01) than controls (94.4% vs 78.4%, respectively). Parity was associated with expression of estrus (P < 0.02), with 78% of first-parity and 93% of later-parity sows exhibiting estrus. However, no treatment x parity effect was observed (P > 0.10). The interval from weaning to estrus was reduced (P < 0.0001) by P.G. 600 compared with controls (3.8+/-0.1 d vs 4.9+/-0.1 d). Follicle size at estrus was not affected by treatment (P > 0.10). The percentage of sows that ovulated did not differ (P > 0.10) for P.G. 600 and control sows (90.3% vs 81.5%, respectively). Time of ovulation after estrus was not affected by treatment and averaged 44.8 h. However, univariate analysis indicated that the interval from weaning to estrus influenced the interval from estrus to ovulation (r = 0.43, P < 0.0001). Further, multivariate analysis showed an effect of treatment on the intervals from weaning to estrus, weaning to ovulation (P < 0.0001), and estrus to ovulation (P < 0.04). Within 4 d after weaning, 81% of the P.G. 600 sows had expressed estrus compared with 33% of controls. However, this trend reversed for ovulation, with only 35% of P.G. 600 sows ovulating by 36 h after estrus compared with 40% of controls. The estrus-to-ovulation interval was also longer for control and P.G. 600 sows expressing estrus < or = 3 d of weaning (45 h and 58 h, respectively) than for sows expressing estrus after 5 d (39 h and 32 h, respectively). Farrowing rate and litter size were not influenced by treatment. However, the interval from last insemination to ovulation (P < 0.02) indicated that more sows farrowed (80%) when the last insemination occurred at < or = 23 to > or = 0 h before ovulation compared with insemination > or = 24 h before ovulation (55%). In summary, P.G. 600 enhanced the expression of estrus and ovulation in weaned sows but, breeding protocols may need to be optimized for time of ovulation based on the interval from weaning to estrus.

Effect of subcutaneous vs intramuscular administration of P.G. 600 on estrual and ovulatory responses of prepubertal gilts.

The effects of s.c. and i.m. administration of P.G. 600 on estrual and ovulatory responses of prepubertal gilts were investigated. One hundred eighty-four crossbred gilts between 159 and 174 d of age were assigned to receive P.G. 600 s.c. (s.c. P.G. 600) in the flank, P.G. 600 i.m. in the neck (i.m. P.G. 600), or no treatment (control). At the beginning of the study (d 0), animals were selected from a modified, open-front barn, regrouped, relocated to new pens, and exposed once daily to a mature boar to check for estrus. On d 17, ovaries were collected from all gilts and analyzed for the presence of corpora lutea (CL), cystic follicles, and cystic CL. A higher proportion of gilts expressed estrus with s.c. P.G. 600 (76%) than with i.m. P.G. 600 (52%, P < .01) or controls (15%, P < .01). The interval from initiation of treatment on d 0 to estrus was reduced (P < .01) by P.G. 600 (4.6 d) compared to controls (5.9 d), but there was no significant difference between P.G. 600 treatments. Both s.c. P.G. 600 (86%) and i.m. P.G. 600 (77%) induced more gilts to ovulate (P < .01) than controls (18%), but there was no significant difference between P.G. 600 treatments. No significant effect of treatment was detected on number of CL (17.9), number of cystic follicles (1.5), or number of cystic CL (2.1). Proportions of gilts that developed cystic follicles or cystic CL were not influenced by treatment. Results of this study indicated that s.c. administration of P.G. 600 significantly improved the induction of estrus in prepubertal gilts compared to i.m. administration.

Apoptosis in breast carcinomas detected with monoclonal antibody to single-stranded DNA: relation to bcl-2 expression, hormone receptors, and lymph node metastases.

Precise quantitation of apoptotic cells in solid tumors is necessary to determine the role of apoptosis in cancer growth, prognosis, and treatment. In this study, the intensity of apoptotic death was determined in 91 breast carcinomas with a novel cellular marker of apoptosis based on the staining of histological sections with a monoclonal antibody (MAb) to single-stranded DNA. Staining of apoptotic cells with the MAb reflected the decreased thermal stability of DNA induced by the digestion of nuclear proteins, as demonstrated by the elimination of staining in sections reconstituted with histones before heating. The high sensitivity and specificity of apoptosis analysis with the MAb is based on the central role of protease activation in the mechanism and control of apoptosis. Apoptotic indexes (AIs) in breast carcinomas ranged between 0 and 46%. Most of the carcinomas had relatively low AIs, whereas 29 cases were classified as carcinomas with intensive apoptosis (AI >/= 10%). The high level of apoptotic cell death was associated with negative immunostaining for bcl-2 protein, the loss of estrogen and progesterone receptors, high proportion of cells in S-phase, and increased risk of lymph node metastases. There was no correlation between AI and tumor size or p53 immunostaining. Lymph node metastases were detected in 59% of patients with high levels of apoptosis in primary carcinomas and in only 21% of patients with AIs below 10% in primary carcinomas. Thus, the high sensitivity of the MAb assay made it possible to identify a subset of breast carcinomas with intensive apoptosis and markers of poor prognosis. These results demonstrate that the measurement of apoptosis in breast carcinomas provides valuable prognostic information.

Monoclonal antibody to single-stranded DNA is a specific and sensitive cellular marker of apoptosis.

The most widely used histochemical marker of apoptosis (in situ end labeling, TUNEL) detects both apoptotic and necrotic cells and evaluates only late stages of apoptosis. Hence, a specific and sensitive cellular marker of apoptosis is needed to determine the role of apoptotic death in biology and pathology. The present study describes a novel immunohistochemical procedure for the staining of apoptotic cells using a monoclonal antibody (MAb) to single-stranded DNA. This MAb stained all cells with the morphology typical of apoptosis in etoposide-treated HL-60, MOLT-4, and R9 cell cultures, in which apoptosis was accompanied by high, moderate, and low levels of internucleosomal DNA fragmentation, respectively. TUNEL stained all apoptotic cells in HL-60 cultures, nearly 60% of apoptotic cells in MOLT-4 cultures, and only 14% of apoptotic cells in R9 cultures. Apoptotic R9 cells, which progressed into secondary necrosis, retained MAb staining and became TUNEL-positive. Necrotic cells in MOLT-4 cultures treated with sodium azide were stained by TUNEL, but were negative for MAb staining. All floating cells at a late stage of apoptosis in MDA-MB-468 cultures treated with cisplatin were stained by both MAb and TUNEL. However, among adherent cells in the early stages of apoptosis, MAb stained nearly 20 times more cells than TUNEL. In histological sections of human tumor xenografts, MAb detected clusters of apoptotic cells in viable tumor tissue, but did not stain cells in areas of central ischemic necrosis. In contrast, TUNEL stained nuclei in necrotic areas. Thus, MAb to single-stranded DNA is a specific and sensitive cellular marker of apoptosis, which differentiates between apoptosis and necrosis and detects cells in the early stages of apoptosis.

Apoptosis in human breast and gastrointestinal carcinomas. Detection in histological sections with monoclonal antibody to single-stranded DNA.

We report application of a novel immunohistochemical procedure for the staining of apoptotic (AP) cells in paraffin sections using monoclonal antibody (MAb) to single-stranded DNA. MAb differentiated between apoptosis and necrosis and in contrast to in situ end labelling specifically stained only AP cells. AP carcinoma cells stained with the antibody were detected in 32 of 58 infiltrating human breast carcinomas and in 9 of 15 colon adenocarcinomas. Stromal cells stained with the MAb were observed in all carcinomas, including those in which no AP carcinoma cells were detected. There was a strong positive correlation between the presence of AP cells, loss of hormone receptors and a high proliferation rate in breast carcinomas. AP cells were present in 80-87% of receptor-negative carcinomas, while most of receptor-positive breast carcinomas did not contain AP cells. Apoptosis in tumor cells was detected significantly more frequently among breast carcinomas with high, than among carcinomas with low S-phase fraction. AP cells were present in 93-95% of breast carcinomas which were receptor-negative and had a high S-phase fraction. Immunostaining demonstrated a strong positive correlation between the loss of bcl-2 protein and intensive apoptosis in breast carcinomas. Association between apoptosis and markers of poor prognosis in breast cancer (loss of hormone receptors, intensive proliferation, loss of bcl-2 protein) indicates that apoptotic cell death is typical of more aggressive carcinomas.

Atypical squamous cells of undetermined significance. Current laboratory practices of participants in the College of American Pathologists Interlaboratory. Comparison Program in Cervicovaginal Cytology.

OBJECTIVE: To evaluate current laboratory practices and rates for atypical squamous cells of undetermined significance (ASCUS), a category of epithelial cell abnormality in the Bethesda System. DESIGN: Questionnaire surveys were mailed in December 1993 and March 1994. SETTING: Cytopathology laboratory participants in the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology (PAP). RESULTS: Most responding laboratories (82.5%) limited the use of "atypia" terminology to abnormalities of undetermined significance. Nearly half of the laboratories employed only the term ASCUS for squamous epithelial changes in this category. The median rate of ASCUS in 1993 was 2.8%, with 10% of laboratories reporting rates greater than 9.0%. The median squamous intraepithelial lesion rate was 2.0%, with a median ASCUS-squamous intraepithelial lesion ratio of 1.3. The majority of laboratories qualified a portion of ASCUS cases and issued recommendations for follow-up when appropriate. Fifty-six percent of laboratories surveyed included patients diagnosed with ASCUS in follow-up programs. Laboratories estimated that about 20% (median response) of patients with ASCUS smears had a squamous intraepithelial lesion or equivalent diagnosis made within a year's follow-up. CONCLUSIONS: The ASCUS category is used by the majority of laboratories as recommended by the Bethesda System, but reporting rates vary. The results of this survey and associated surveys provide laboratories with useful benchmark figures for interlaboratory comparison of ASCUS practices.

Synergistic induction of apoptosis in breast cancer cells by tamoxifen and calmodulin inhibitors.

Breast cancer cells are relatively resistant to the induction of apoptosis (AP) and drug regimens which readily activate apoptotic death, may enhance the antitumor effect. Rapid and intensive induction of apoptosis was observed in estrogen receptor positive and negative breast cancer cell cultures treated with tamoxifen (TMX) combined with the calmodulin antagonists trifluoperazine (TFP) or W7. TMX (1-5 microM) alone or calmodulin antagonists alone did not induce apoptosis. Importantly, intensive apoptosis was also induced by TMX and TFP in the cells obtained from primary human breast carcinomas. Inhibition of the Ca2+ calmodulin signaling pathway is an effective way to activate apoptotic death in epithelial cells. Combination of TMX with non-toxic calmodulin inhibitors may increase the preventive and therapeutic effects of TMX.

Caring for children in an adult intensive care unit--Part I.

Various reports draw attention to the deficiency of services available for critically ill children. In December 1993 the British Paediatric Association published a report by a multidisciplinary working party on intensive care. The Minister of Health has apparently promised to act on the recommendations the working party made. In fact, purchasers are being encouraged to act and ask questions about the provision of paediatric intensive care--but no extra funding is available to provide it. However, this will all take time and until more resources are available children are still being cared for on adult intensive care units by nurses who may not be experienced in caring for sick children. One of the main areas of concern nurses have when caring for children is the physiological differences between an adult and a child. This paper is designed to assist in identifying the differences and consequent nursing implications. In an attempt to prioritize the nursing care a systems/modified model has been used over 2 papers. The review of the systems demonstrates the differences between adults and children and, where possible, highlights the nursing care and medical treatment a child requires. All aspects of care have been included as well as the psychological problems (see Part 2 which will be published in the next issue) encountered by the family of a critically ill child. Where possible easy reference tables have been included and the author hopes to introduce the package into the existing orientation programme for established staff and staff new to the unit in the hopes of decreasing the stress when a child is admitted. Paediatric resuscitation has not been covered intentionally, as the information on the physiological differences seemed to grow like 'Jack's beanstalk'. Therefore, a flow chart for basic and advanced life support is included as an appendix in Part I. It is hoped this will stimulate readers' interest for more in-depth study.

Monoclonal antibody KS1/4-methotrexate immunoconjugate studies in non-small cell lung carcinoma.

The antigen reactive with murine monoclonal antibody (MAb) KS1/4 is expressed on epithelial malignancies and some normal epithelial tissues. Studies were undertaken to evaluate KS1/4-methotrexate (KS1/4-MTX) immunoconjugate in patients with advanced non-small cell carcinoma of the lung. Eleven patients in two different groups received KS1/4-MTX in two different escalating dose infusion schedules with a maximal tolerated dose of 1,750 mg/M2 and a cumulative dose of MTX of 40 mg/M2. Toxicities were similar in both groups and included fever, anorexia, nausea, vomiting, diarrhea, abdominal pain, guaiac positive stool, and hypoalbuminemia. Two patients had an associated aseptic meningitis. One patient had a 50% decrease in two lung nodules without a change in lymphangitic infiltrates. This patient received a second course of treatment and developed an immune complex-mediated arthritis and serum sickness. Four patients mounted a human antimouse antibody response. Post-treatment tumor biopsies documented binding of MAb KS1/4. These studies document the feasibility and potential usefulness of a MAb directed against tumor-associated antigens with the targeting of chemotherapeutic drugs in patients with non-small cell lung carcinoma.

An overview of ventilator observations.

Nursing staff new to intensive care can often be apprehensive when observing, recording and interpreting ventilator observations. This review of ventilation and the necessary observations involved offers a description of the observations performed, with a problem-solving approach to cause, rationale and possible action that may be required. Calculations related to spontaneous and mechanical respiratory function are discussed, with recommendations for individual requirements. Explanation of available settings, demonstrating the functions and delivery methods of different ventilators is included, and a discussion on alarm settings and rationale. For the purpose of assessing the ventilatory support in use there is a guide to interpretation of arterial blood gases with suggestions and rationale for necessary changes of parameters.

The distinction of small cell and non-small cell lung cancer by growth in native-state histoculture.

Histological analysis remains the primary method of distinguishing between small cell (SCLC) and non-small cell lung cancer (NSCLC). This distinction has significant impact therapeutically because of their relative difference in chemoresponsiveness (J.D. Minna et al., Principles and Practice of Oncology, pp. 396-474, 1981). Yet for at least 10% of lung tumors, pathologists will disagree upon the classification (A.R. Feinstein et al., Am. Rev. Respir. Dis., 101: 671-684, 1970). Furthermore, current neuroendocrine markers lack specificity for SCLC although the presence of these markers may help predict chemosensitivity (S.L. Graziano et al., J. Clin. Oncol., 7: 1375-1376, 1989; S.B. Baylin, J. Clin. Oncol., 7: 1375-1376, 1989; C.L. Berger et al., J. Clin. Endocrinol. Metab., 53: 422-429, 1981; A.F. Gazdar et al., Cancer Res., 45: 2924-2930, 1985). In vitro growth characteristics may more accurately reflect biological properties of aggressiveness and susceptibility to chemotherapy. In this study, 3-dimensional gel-histoculture was used to retrospectively distinguish between NSCLC and SCLC. Tumor explants from 78 patients with NSCLC and 13 patients with SCLC were grown in gel-supported histocultures with an overall success rate of 92%. These 2 tumor types were distinguishable by their 3-dimensional in vitro tissue architecture. In addition, proliferation rates were measured by histological autoradiography after 4-day incorporation of [3H]dThd. The percentage of cells labeled in the most proliferatively active regions of the autoradiograms was termed the growth fraction index (A.F. Gazdar et al., Cancer Res., 45: 2924-2930, 1985; R.A. Vescio et al., Proc. Natl. Acad. Sci. U.S.A., 84: 5029-5033, 1987; R.M. Hoffman et al., Proc. Natl. Acad. Sci. U.S.A., 86: 2013-2017, 1989). The mean growth fraction index for pure small cell lung cancer was 79 +/- 10%, differing markedly from that of 35 +/- 19% for mixed small cell/large cell tumors, adenocarcinoma (38 +/- 16%), large cell undifferentiated carcinoma (40 +/- 18%), and squamous cell carcinoma (33 +/- 15%) (P less than 0.001 in each case). We therefore conclude that 3-dimensional gel-histoculture is a useful means of distinguishing pure SCLC from NSCLC, which may improve treatment decision making.

The future anatomic pathology laboratory for molecular diagnosis and prognosis: circa 1990-95.

Molecular biology provides analytical tools for the study of human disease. These clinical assays can help make a diagnosis, predict a prognosis, or elucidate the pathogenesis of poorly understood diseases. The detection and/or quantitation of specific genes, messenger RNA's, and their protein products provide diagnostic and prognostic information for physicians. Illustrative applications of some of these techniques in human virus infection and breast cancer will suggest the future direction that an anatomic pathology laboratory should consider if it is to provide optimal and competitive service to physicians.

Phase I clinical comparative study of monoclonal antibody KS1/4 and KS1/4-methotrexate immunconjugate in patients with non-small cell lung carcinoma.

A Phase Ia clinical trial was undertaken to evaluate and compare murine monoclonal antibody KS1/4 and KS1/4-methotrexate immunoconjugate in patients with Stage IIIB or IV non-small cell carcinoma of the lung. Six patients received KS1/4 alone and five patients received KS1/4-methotrexate conjugate. The maximal total dose received per patient in both groups was 1661 mg. Mild to moderate side effects in both groups included fever, chills, anorexia, nausea, vomiting, diarrhea, anemia, and brief transaminasemia. One patient who received antibody alone had an apparent acute immune complex-mediated reaction. Ten of 11 patients had a human anti-mouse response. Posttreatment carcinoma biopsies revealed binding of monoclonal antibody KS1/4 and deposition of C3d and C4c complement fragments. Monoclonal antibody binding and complement deposition correlated with increasing doses of infused antibody. There was one possible clinical response.

Cancer biology for individualized therapy: correlation of growth fraction index in native-state histoculture with tumor grade and stage.

There is a need for individualization of all aspects of cancer therapy. Because of significant heterogeneity within a tumor class, there is a need to develop an in vitro test to accurately gauge tumor aggressiveness. Such a measurement would greatly aid treatment decision making. Current methodologies such as flow cytometry, which lacks unambiguous interpretation of cell-proliferative data, and determination of the thymidine-labeling index, which measures nucleotide uptake in a nonphysiological state, have not reproducibly attained this goal. We have developed an in vitro native-state three-dimensional gel-supported histoculture system that allows the growth of all human solid tumor types for relatively long time periods. The native-state system was used to identify the percent of cells capable of incorporating [3H]thymidine over a 4-day period, which we term the growth fraction index (GFI). We have compared the ability of cancer tissue to proliferate in native-state culture to the stage and histological grade of four major types of human carcinomas: breast, ovarian, colon, and lung. Eighty percent of tumor explants could be evaluated, even when sent from across the country. We have determined that the GFI correlates with tumor stage and grade for breast and ovarian carcinoma. In colon carcinoma, there is a trend toward higher GFIs in tumors of more advanced stage and grade. In non-small cell lung carcinomas, GFI, stage, and grade do not correlate. These results suggest the applicability of gel-supported three-dimensional native-state histoculture for prognostic purposes in patients with breast and ovarian cancers and demonstrate the clinical relevance of the native-state histoculture system.