Supervision and staff training for children's group psychotherapy: general principles and applications with cumulatively traumatized, inner-city children.

Cumulatively traumatized children frequently benefit from theme-centered, trauma-focused psychotherapy groups, which promote recovery from trauma and enhance ego functioning. Unfortunately, even as children's group programs proliferate, therapists are still routinely assigned to supervisors who neither understand nor appreciate the modality. Modality-specific supervision is critical in treating traumatized children in groups, because such children are at significant risk of retraumatization. This article surveys and discusses problematic supervisory practices; advocates modality-specific supervision focusing on peer interaction, multiple transference enactments, and group culture; and describes a process-oriented consultation group that facilitates modality-specific cognitive and affective learning.

In vivo and in vitro effects of androgen on fibroblast growth factor-2 concentrations in the human prostate.

Prostatic growth is primarily regulated by dihydrotestosterone (DHT). Recent studies have demonstrated that a large number of growth factors are present in the human benign prostatic hyperplasia (BPH) prostate, including epidermal growth factor (EGF), transforming growth factor-alpha (TGF-alpha), transforming growth factor beta (TGF-beta), insulin-like growth factor (IGF), and basic fibroblast growth factor (bFGF) (FGF-2). DHT may mediate its mitogenic effects in the prostate by regulating growth factors. To test this hypothesis, we have utilized a histoculture androgen sensitivity assay (HASA) in which 3H-thymidine incorporation is measured in aliquots of BPH tissue in histoculture with either added DHT or hydroxyflutamide (HF). The resulting DHT/HF ratio is an expression of the androgen sensitivity of the tissue. In this study, we have compared the DHT/HF ratio for 3H-thymidine incorporation to the DHT/HF ratio for FGF-2 measured in the histocultured prostates. The DHT/HF ratio for the HASA studies of 3H-thymidine incorporation averaged 2.68 compared to the DHT/HF ratio for FGF-2 in the same specimens of 1.01. These values were significantly different, therefore indicating no relationship between DHT stimulation and FGF-2 levels. In addition, FGF-2 levels were measured in human BPH prostates from patients medically castrated with megesterol acetate and estradiol 17-beta prior to surgery. These values were not significantly different, and therefore do not suggest any effect of DHT on the concentration of prostatic FGF-2. Although these studies did not show any effect of DHT on the regulation of prostatic FGF-2, they do indicate that the HASA assay is feasible and appropriate to use in the study of relationships between DHT and various growth factors.

Sponge-gel-supported histoculture drug-response assay for head and neck cancer. Correlations with clinical response to cisplatin.

OBJECTIVE: Sponge-gel-supported histoculture drug-response assay (SSHDRA) represents a promising method to determine chemosensitivity of solid tumors. To determine whether the assay correlates clinically, we compared the in vivo and in vitro effects of cisplatin in 23 of 26 patients with head and neck cancers. DESIGN: The criterion for in vitro sensitivity to cisplatin was an 84% or greater inhibition by cisplatin of the number of tritiated thymidine-incorporating cells of the histocultured tumors compared with untreated control culture preparations, as measured by means of histologic autoradiography. Comparisons were made with clinical responses, ie, complete response, partial response, or no response. PATIENTS: The study was carried out in patients with head and neck cancers and comprised 21 patients with squamous-cell carcinoma, three patients with other carcinomas, and two patients with sarcoma. RESULTS: Ten of 12 patients with in vitro-sensitive tumors had either complete or partial response clinically. The overall accuracy of the SSHDRA was 74% in this correlative clinical trial; the predictive-positive value was 83%, the sensitivity was 71%, and the specificity was 78%. Seven of 11 patients with in vitro-resistant tumors demonstrated no response for a predictive-negative value of 64%. CONCLUSIONS: We conclude that the SSHDRA shows a high correlation for tumors that demonstrate both in vivo drug resistance and sensitivity. The in vitro-like maintenance of three-dimensional tissue architecture of the tumors in histoculture probably contributes to high clinical predictivity of drug response of the SSHDRA. The data support further comparisons to determine the clinical usefulness of the SSHDRA for identifying complete and partial responders to chemotherapy.

Lack of influence of c-Ha-ras expression on the drug sensitivity of human bladder cancer histocultured in three-dimensions.

The mechanism of drug resistance in human cancers is complex. In addition to overexpression of a series of multiple-drug-resistance genes, there has been the suggestion that the Ha-ras gene may participate in conferring resistance. In this study, a series of three human-bladder carcinoma cell lines were studied, one parental type, one transfected by wild-type Ha-ras and another transfected by mutant Ha-ras. The ras gene was overexpressed in the latter two cell lines which also were more invasive than the parental when injected as individual cells in the nude-mouse bladder. The results described here have indicated that the ras-gene expression level or mutational status did not affect drug resistance when the tumor lines were histocultured as three-dimensional tissue on collagen-sponge-gels. The drug-response spectrum of the histocultured lines qualitatively reflected a clinical experience although all lines were relatively drug resistant, possibly reflecting their three-dimensional configuration in culture.

Correlation of drug response in human tumors histocultured in vitro with an image-analysis MTT end point and in vivo xenografted in nude mice.

We have in this study used the 3-(4,5-dimethyl-2- thiazoyl) -2,5-diphenyl 2-H-tetrazolium bromide (MTT) end point in our histoculture drug-response assay. We have previously demonstrated that the formazan crystals formed by MTT reduction by mitochondrial succinate dehydrogenase reflect polarized light and can be measured by pixel analysis in intact tissue. The results described here indicate a total specificity of 93.8% and a total accuracy of 74.6% of the MTT end point for drug response in histoculture correlating with nine different human xenograft tumors grown in nude mice with respect to the in vivo drug response data. This in vitro system allows prediction of positive and negative responses to drugs, with a rate of 70% and 71.8%, respectively. The system described here has potential for clinical use because of the possibility of simultaneous description of the MTT values and heterogeneous response to drugs within individual tumors.

Noncolorimetric measurement of cell activity in three-dimensional histoculture using the tetrazolium dye 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide: the pixel image analysis of formazan crystals.

We describe a novel system for measuring the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction in three-dimensional histoculture which is no longer dependent on colorimetric determination of extracted formazan, but rather is based on a pixel image analysis of formazan crystals, and which allows intratumor heterogeneity to be taken into account. The MTT test is based on the enzymatic reduction of the tetrazolium salt 3-(4,5-dimethylthiazol-2-yl)-2,5-dipheniltetrazolium bromide to formazan crystals by living, metabolically active cells, but not in dead cells. The reaction was carried out in situ in six-well plates on gel-supported histocultured human tumors. After a 24-h incubation with different drugs the tumors were incubated with a solution of MTT. Frozen sections of the tumor pieces were made and the slides were then stained with a propidium iodide solution, whose fluorescence is proportional to the number of cells present. We demonstrate here that the formazan crystals, formed by MTT reduction, reflect polarized light and that this can be quantified by using an image analysis system based on bright-pixel quantitation directly on a frozen section of the original tissue. Combined with the use of the fluorescent dye propidium iodide, also measured by pixel analysis, we can express a ratio between the total amount of MTT reduction and the total number of cells present in the specimen that expresses the effect of drugs on the histocultured tumors. Since histology is well maintained in histoculture it is possible to take into account the heterogeneity present in the tumor with regard to drug response.(ABSTRACT TRUNCATED AT 250 WORDS)

In vitro histoculture of human tumors with fluorescent dye end-points measured by confocal microscopy: high correlation of in vitro and in vivo chemosensitivity.

Tumors from 40 patients and 7 established human xenograft tumor lines were grown in three-dimensional histoculture. A Viable-Cell-Index (VCI) based on fluorescent dyes and Growth Fraction Index (GFI) based on [3H]thymidine incorporation were measured by confocal microscopy and histological autoradiography, respectively, after treatment with cytotoxic agents. Chemosensitivity in vitro with the two methods was correlated with chemosensitivity of the same set of human xenografted tumor lines grown in nude mice. The percent accuracy of in vitro to in vivo correlation with VCI (73%) was higher than GFI (63%). The number of false positives with VCI was 12.1% (4/33), and with GFI was 31.3% (10/32). The results thus indicated that in vitro histoculture with fluorescent vital-dye end-points to measure cell viability is of potential use to determine tumor chemosensitivity.

Correlation of histology and drug response of human tumors grown in native-state three-dimensional histoculture and in nude mice.

An in vitro histoculture system in which a native-state collagen-sponge gel supports the three-dimensional growth of tumor tissue has been recently developed that allows the culture and drug response assay for most every tumor type. Important features of the histoculture system include the maintenance of three-dimensional tissue architecture and the use of histological autoradiography to measure nuclear incorporation of [3H]thymidine as an endpoint. We describe in this report in vitro-in vivo correlations for drug response and tumor histology by using human tumor xenografts grown in the native-state three-dimensional histoculture system and as xenografts in nude mice. This comparison eliminates many of the confounding variables seen in most correlative clinical trials. Results demonstrate (i) a very high preservation of in vivo tissue architecture in vitro, (ii) an 86% accuracy in vitro of predicting drug resistance in vivo using suprapharmacologic doses of drugs in vitro, and (iii) an overall predictive frequency of drug resistance and sensitivity ranging from 53% for 5-fluorouracil to 78% for doxorubicin.

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.

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.

A general native-state method for determination of proliferation capacity of human normal and tumor tissues in vitro.

An important need in cancer research and treatment is a physiological means in vitro by which to assess the proliferation capacity of human tumors and corresponding normal tissue for comparison. We have recently developed a native-state, three-dimensional, gel-supported primary culture system that allows every type of human cancer to grow in vitro at more than 90% frequency, with maintenance of tissue architecture, tumor-stromal interaction, and differentiated functions. Here we demonstrate that the native-state culture system allows proliferation indices to be determined for all solid cancer types explanted directly from surgery into long-term culture. Normal tissues also proliferate readily in this system. The degree of resolution of measurement of cell proliferation by histological autoradiography within the cultured tissues is greatly enhanced with the use of epi-illumination polarization microscopy. The histological status of the cultured tissues can be assessed simultaneously with the proliferation status. Carcinomas generally have areas of high epithelial proliferation with quiescent stromal cells. Sarcomas have high proliferation of cells of mesenchymal organ. Normal tissues can also proliferate at high rates. An image analysis system has been developed to automate proliferation determination. The high-resolution physiological means described here to measure the proliferation capacity of tissues will be important in further understanding of the deregulation of cell proliferation in cancer as well as in cancer prognosis and treatment.