Measurement of chemopreventive efficacy in skin biopsies.

OBJECTIVE: To explore methods suitable for quantitative assessment of the efficacy of chemopreventive intervention. STUDY DESIGN: High-resolution imagery of nuclei from the suprabasal and basal cell layers of sun-damaged skin were recorded. There were 10 cases. A shave biopsy was taken from an area of clearly evident solar keratosis before and after treatment with 2-difluoromethyl-dlornithine (DFMO) and from the colateral forearm, treated with a placebo. A number of karyometric variables were computed and combined to derive marker features that provided a numeric measure of the degree of nuclear deviation from normal. RESULTS: DFMO treatment was effective overall in reducing the degree of nuclear abnormality seen in the biopsies; in 8 of the 10 cases there was a significant improvement. The placebo-treated arm did not show a statistically different abnormality from the untreated arm. CONCLUSION: Karyometric analysis can provide numeric measures that allow documentation of statistically significant regression of actinic keratotic lesions following treatment with DFMO.

Relationship of p53 mutations to epidermal cell proliferation and apoptosis in human UV-induced skin carcinogenesis.

Human skin is continually subjected to UV-irradiation with the p53 gene playing a pivotal role in repair of UV-induced DNA damage and apoptosis. Consequently, p53 alterations are early events in human UV-induced skin carcinogenesis. We studied 13 squamous cell carcinomas (SCC), 16 actinic keratoses (AK), 13 samples adjacent to an AK (chronically sun-damaged), and 14 normal-appearing skin samples for p53 mutation, p53 immunostaining (IHC), apoptosis (in situ TUNEL and morphology), and proliferation (PCNA). The frequency of p53 mutation increased from 14% in normal skin, to 38.5% in sun-damaged skin, 63% in AK, and 54% in SCC. p53 IHC increased similarly. Apoptosis (TUNEL) increased from 0.06 +/- 0.02%, to 0.1 +/- 0.2, 0.3 +/- 0.3, and 0.4 +/- 0.3 in normal skin, sun-damaged skin, AK, and SCC, respectively. Apoptosis was strongly correlated with proliferation (i.e., TUNEL and PCNA, r = 0.7, P < 0.0001), and proliferation was significantly increased in the progression from normal skin to SCC. Bax was significantly increased in SCC compared to AK. These data imply that apoptosis in samples with a high frequency of p53 mutation may not necessarily be p53-dependent. We suggest that there is a mechanism for apoptosis in response to increased cellular proliferation that is p53-independent.

Nuclear morphometry in solar keratosis.

OBJECTIVE: To carry out a feasibility study for the development of procedures for the objective characterization and grading of solar keratotic skin lesions. STUDY DESIGN: Imagery from sections of skin shave biopsies from 12 light-skinned individuals were digitized. A minimum of 25 nuclei from a solar keratotic lesion and 25 nuclei from a location in histologically normal appearing skin adjacent to the lesion were recorded for each case. Values of karyometric features were computed, and a discriminant function distinguishing normal nuclei from nuclei exhibiting solar irradiation damage was derived. RESULTS: Approximately 50% of nuclei in solar keratotic lesions were markedly affected by solar irradiation, but even in biopsies from histologically normal appearing skin, 3-30% of nuclei showed signs of such damage. Nuclei from solar keratotic lesions exhibiting such damage had numerous morphometric and karyometric features commonly found in malignant cells. The state of progression of a solar keratotic lesion can be graded by a plot of proportion of nuclei exhibiting solar damage versus the average discriminant function score of the most affected nuclei. This plot provides a monotonically rising progression curve and a numeric grading score. CONCLUSION: Karyometry of nuclei from skin biopsies allows objective assessment of the progression of solar keratotic lesions. Similarity of feature values in nuclei from solar keratotic lesions to those in malignant lesions was noted. The progression curve derived in this study could serve to measure the efficacy of chemopreventive or therapeutic intervention.

Expression of p53 protein in actinic keratosis, adjacent, normal-appearing, and non-sun-exposed human skin.

Nonmelanoma skin cancer, including both squamous cell carcinoma and basal cell carcinoma, is a significant and increasing health problem in the United States. The precursor lesion of cutaneous squamous cell carcinoma, actinic keratosis (AK), is a major risk factor for nonmelanoma skin cancer, and it is also a marker of long-term sun exposure. AKs themselves can serve as biomarkers in chemopreventive studies, but in addition, they may contain phenotypic and genetic alterations that are related to the process of UV-induced skin carcinogenesis. One of these alterations, the tumor suppressor gene p53, is altered early in UV-induced skin carcinogenesis in humans. p53 protein expression was measured by immunohistochemistry in biopsies from AKs, tissue immediately adjacent to AKs (AK-adjacent), normal-appearing upper medial arm skin, and non-sun-exposed skin from 19 subjects. There was a significant difference and a progressively increasing mean p53 labeling index in total epidermis (basal and suprabasal layers) between upper medial arm skin (0.9 +/- 1.8%) and AK-adjacent (12.1 +/- 14.4%; P = 0.0004) and between AK (27.7 +/- 21.3%) and AK-adjacent skin (P = 0.04), whereas upper medial arm skin was marginally different from non-sun-exposed skin (0.1 +/- 0.2; P = 0.05). This pattern of p53 expression was also seen when epidermis was separated into basal and suprabasal layers. We conclude that epidermal p53 protein expression is associated with histological evidence of chronic sun damage.

Treatment of acute respiratory failure secondary to pulmonary oedema with bi-level positive airway pressure by nasal mask.

We report the successful outcome of first-line intervention of noninvasive positive pressure ventilation (NPPV) in four patients, three of whom had hypercapnic acute respiratory failure (ARF) and one hypoxaemic ARF, secondary to pulmonary oedema. The clinical condition showed rapid improvement and the NPPV, performed together with aggressive medical treatment, was effective in decreasing the respiratory frequency, and in correcting gas exchange abnormalities within the first 3 h. The average duration of nasal mask ventilation was 11 h (range 6-15 h). The patients were weaned, following ARF, by removing the ventilator whenever inspiratory positive airway pressure (IPAP) was 5 cmH2O. NPPV was applied, by nasal mask, using a bi-level positive airway pressure (BiPAP) delivering pressure support ventilation (PSV). We conclude that application of noninvasive positive pressure ventilation may be effective in correcting gas exchange abnormalities, in relieving respiratory distress and, perhaps, in avoiding endotracheal intubation in selected patients with acute respiratory failure secondary to reversible medical condition such as pulmonary oedema.

Evaluation of proliferating cell nuclear antigen as a surrogate end point biomarker in actinic keratosis and adjacent, normal-appearing, and non-sun-exposed human skin samples.

The incidence of nonmelanoma skin cancer, including both squamous cell carcinoma and basal cell carcinoma, is a significant health problem in the United States. Actinic keratosis (AK), the precursor of cutaneous squamous cell carcinoma, is a major risk factor for nonmelanoma skin cancer. In addition, AKs are tissue targets for the identification of biomarkers for use in chemopreventive studies. The biomarker addressed in this study is epidermal cell proliferation, as quantitated by proliferating cell nuclear antigen (PCNA). Shave biopsies were obtained from AKs, tissue immediately adjacent to AKs, normal-appearing, upper-medial arm skin, and non-sun-exposed skin from 19 subjects. When any degree of PCNA staining was considered positive (semiquantitative 1-4 scale), there was a significant difference and a progressively increasing mean PCNA labeling index (LI) in the total epidermis (basal and suprabasal layers), beginning with non-sun-exposed buttock skin, with the lowest LI (2.5 + or - 1.6%), followed by upper-medial arm skin (12.3 + or - 7.4%; P = 0.0015), skin adjacent to AKs (19.2 + or - 12.2%; P = 0.0218), and finally, AKs with the highest LI (34.6 + or - 20.1%; P = 0.0017). This same pattern was observed when the epidermis was separated into basal and suprabasal layers, with the exception of a nonsignificant result for upper-medial arm skin compared with adjacent skin in the basal layer (P = 0.3981). PCNA LIs were also analyzed separately by staining intensity (i.e., scores of 1-4). The PCNA LI in skin with varying degrees of sun damage and/or histological atypia is a candidate surrogate end point biomarker for skin cancer chemoprevention studies.

Analysis of the p53 gene in human precancerous actinic keratosis lesions and squamous cell cancers.

A biomarker of skin cancer would be beneficial in evaluating the efficacy of potential cancer chemoprevention agents. To this end, we investigated the tumor suppressor gene p53 in precancerous actinic keratosis lesions (AK) and malignant squamous cell carcinomas (SCCs) using polymerase chain reaction and single-strand conformation polymorphism analysis (PCR-SSCP) techniques. In addition, p53 protein expression was evaluated using immunohistochemistical analysis with the PAB 1801 monoclonal antibody. Nine out of 13 (69%) SCCs and 8 of 15 (53%) AKs were positive for p53 mutations. In contrast, normal skin samples were negative for p53 mutations. Sequence analysis of AKs and SCCs showed primarily C to T transition mutations. Nuclear immunochemical staining for p53 was observed in 12/15 (80%) AK and 12/13 (92%) SSCs. These results suggest that p53 mutations may be involved in the malignant conversion of AKs to SCCs and that p53 may be useful as a biomarker to study the potential modulatory effects of cancer chemopreventive agents against skin cancer.

Chronic solar ultraviolet damage associated with malignant melanoma of the skin.

A retrospective study of 226 cases of cutaneous malignant melanoma (CMM) was done to determine if patients with CMM showed evidence of chronic solar ultraviolet radiation damage, i.e., a past history of actinic keratosis (AK), basal cell carcinoma (BCC), squamous cell carcinoma (SCC) anywhere on the patient's skin, and solar elastosis (SE) at the site of a CMM. This statistical analysis consisted of 119 clinical records and 158 pathology slides after all cases of lentigo CMM were deleted. CMM showed no statistical correlation with AK, BCC, SCC, or SE. This supports the idea that it is not the chronic solar-ultraviolet radiation exposure that causes CMM.

Malignant melanoma in southern Arizona. Increasing incidence and sunlight as an etiologic factor.

This study demonstrated an increasing incidence of malignant melanoma during the past ten years and a very high incidence of this tumor in southern Arizona. During the ten-year period, 533 melanomas were removed from white patients. Of these, 52% were male and 48% were female. The number of melanomas increased yearly, from 20 in 1969 to 120 in 1978, a crude rate incidence of 6.49 to 28.57 (27.20 standardized) per 100,000, respectively. This reflects an average annual increase of 34% to 37% and a 340% increase for the period. The highest incidence of tumor was in the 50 to 59 year and 60 to 69 year age groups. The most common site of occurrence was the back, with twice as many tumors arising there in males. The legs were involved in 13% of patients, with an occurrence rate eight times higher in females. The extremely high incidence of melanomas in southern Arizona is probably due to meteorologic and geographic factors allowing large amounts of ultraviolet light to reach the earth's surface.

Metastasizing keratoacanthomas? The difficulties in differentiating keratoacanthomas from squamous cell carcinomas.

The problem in diagnosis of keratoacanthoma versus squamous cell carcinoma has been reviewed, and 13 patients are presented to illustrate the difficulties in differentiating between these two lesions. If the pathologist is in doubt, the lesion should be called "probable keratoacanthoma, but squamous cell carcinoma cannot be ruled out." We should all be aware that even the most careful pathologist, using all the information and material available, may still erroneously diagnose a lesion as a benign keratoacanthoma--one which, if inadequately treated, may metastasize or recur as a squamous cell carcinoma. Therefore, it is important for the clinician to treat most keratoacanthomas by adequate removal and close follow-up.

Formalizing physician input in equipment decisions.

The Tucson Medical Center has established a special equipment committee, consisting of representatives from the medical staff, administration, purchasing, and biomedical engineering, to handle its many special equipment purchase requests. This joint responsibility in decision-making has widespread acceptance among physicians and administrators. The institution's rigorous procedure and its subsequent benefits are outlined.