Agarose gel keratinocyte outgrowth system as a model of skin re-epithelization: requirement of endogenous acetylcholine for outgrowth initiation.

To better understand the mechanisms of skin re-epithelization, we developed a simple technique that assays the outgrowth of human keratinocytes. Second-passage foreskin keratinocytes were inoculated at high cell density into 3-mm wells cut from agarose gels in standard 6-well tissue culture dishes. The cells settled on the dish bottom and formed a confluent colony. The cells at the periphery of the colony flattened, spread their cytoplasm, and moved away over the dish surface under the agarose gel. The morphology of migrating keratinocytes was observed microscopically through the transparent agarose, and the migration distance was measured after the gels were removed and after cells were fixed and stained. To determine which cell activities were involved in the outgrowth, the effects of cholinergic compounds on keratinocyte outgrowth were compared with their effects on keratinocyte proliferation, cell-plastic attachment, and spreading measured in separate sets of experiments. Outgrowth was inhibited by the specific inhibitor of acetylcholine synthesis bromoacetylcholine (0.05 mM) and restored by 5 mM exogenous acetylcholine. The irreversible muscarinic antagonist propylbenzilylcholine mustard (0.05 mM) abolished the restorative effects of exogenous acetylcholine, and also inhibited outgrowth of intact keratinocytes. In keratinocyte cell cultures, bromoacetylcholine stopped cell division. Propylbenzilylcholine mustard increased cell number, but interfered with cell-plastic attachment and spreading. This suggests that cell-matrix attachment, spreading, and locomotion of human keratinocytes, but not mitosis, mediate the earliest stages of skin re-epithelization, and that endogenous acetylcholine regulates these keratinocyte functions. Specifically, keratinocyte acetylcholine is required to initiate outgrowth.

Keratinocyte muscarinic acetylcholine receptors: immunolocalization and partial characterization.

We have reported previously that human keratinocytes synthesize and secrete acetylcholine and that muscarinic cholinergic drugs have effects on keratinocyte proliferation, adhesion, and migration. This study defines the location of muscarinic acetylcholine receptors in human epidermis and describes some pharmacologic and molecular properties of these receptors. Confocal microscopy employing the anti-muscarinic receptor monoclonal antibody M35 visualized the receptors in the intercellular areas of normal human epidermis. Using immunoelectron microscopy, the receptors appeared to be attached to the keratinocyte plasma membranes. Functional, high-density (Bmax = 8.3 nmol/2 x 10(6) cells) and high-affinity (Kd = 21.5 nM) muscarinic receptors were demonstrated by saturable binding of the reversible radioligand [3H]quinuclidinyl benzilate to the surfaces of freshly isolated epidermal cells at 0 degrees C. Receptor proteins were separated by gel electrophoresis. An apparent isoelectric point of pH 4.3 was determined in immunoblots of sodium-cholate-solubilized receptors separated on isoelectric-focusing gels. Three protein bands, two at approximately 60 kDa and one at 95 kDa, were visualized in immunoblots of membrane-bound or solubilized receptors separated by sodium dodecylsulfate-polyacrylamide gel electrophoresis. The covalent, irreversible ligand [3H]propylbenzilylcholine mustard confirmed these results. Thus, human keratinocytes express a heterogeneous population of muscarinic cholinergic receptors. Because human keratinocytes also express nicotinic cholinergic receptors, endogenously secreted acetylcholine may control different biologic processes in these cells by activating different types of their cholinergic receptors.

Generalized pustular psoriasis. A review of 63 cases.

BACKGROUND: Sixty-three patients with generalized pustular psoriasis were hospitalized during a 29-year period. They were classified into four subgroups on the basis of onset and morphologic pattern of disease: acute (von Zumbusch), subacute annular, chronic (acral), and mixed. This division provides a better understanding of the variability of the disease and helps in choosing treatment. OBSERVATIONS: The average age at onset was 50 years; male and female patients were affected about equally. In 11 patients, flares were precipitated by localized infections. Approximately one fourth of the patients had complications; most were superinfections. The average stay in the hospital was 30 days; factors correlating with a long hospitalization were hypocalcemia, female sex, and a previous history of psoriasis vulgaris or pustular psoriasis. CONCLUSIONS: Whereas topical therapy was helpful, systemic medications were often needed. Coal tar, ultraviolet light, and psoralen-ultraviolet A may be effective; however, they must be used with caution, because they may exacerbate the disease.

Teledermatology in the nursing home.

OBJECTIVE: To examine a still-image store-and-forward teledermatology system for use in the care of nursing home residents. DESIGN: Diagnosis and treatment plans made from a teledermatology system were compared with those made from an on-site dermatology consultation. SETTING: This study involved the dermatologic care of nursing home residents. PATIENTS: Dermatologic consultations sent to the senior author's office from the participating nursing home were eligible for the study. In a consecutive manner, 29 residents with a total of 30 skin conditions were enrolled. INTERVENTION: A nurse collected and sent the histories and images using the teledermatology system. A diagnosis and treatment plan was determined by examining a transmitted still image and patient history alone and in combination by 2 to 3 dermatologists independently. An independent dermatologist made an on-site dermatologic consultation within 2 days after the images had been collected. MAIN OUTCOME MEASUREMENT: The diagnosis and treatment plans made from the teledermatology system were compared with those made by the on-site dermatologist. RESULTS: Twenty-nine patients with 30 skin conditions were enrolled in the study. Correct diagnoses were made for 60 (67%) of 90, 51 (85%) of 60, and 53 (88%) of 60 patients given the history alone, image alone, and both, respectively. The correct treatment plan was seen in 63 (70%) of 90, 52 (87%) of 60, and 54 (90%) of 60 patients given the history alone, image alone, and both, respectively. No incorrect diagnoses or treatment plans would have given rise to substantial morbidity. The dermatologists felt comfortable in making a diagnosis and treatment plan in all cases in which they had access to both the image and patient history. CONCLUSION: This study provides evidence that nursing home teledermatology consults may replace some on-site consultations by offering quality care in a cost-effective manner.

Ultrastructure of collagen thermally denatured by microsecond domain pulsed carbon dioxide laser.

BACKGROUND: Clinical improvement in photodamaged skin after carbon dioxide (CO2) laser resurfacing is thought to result in part from thermal collagen shrinkage. The presence of such collagen has not been unequivocally demonstrated. To identify and characterize the morphological features of collagen after CO2 laser exposure, we irradiated ex vivo human facial skin and bovine calcaneus tendon with microsecond domain pulsed CO2 laser energy and examined specimens for histopathological and ultrastructural changes in collagen. OBSERVATIONS: In dermis and tendon, 3 zones of collagen structure were apparent on electron microscopy. The first, most superficial zone demonstrated loss of collagen structure. The second zone consisted of admixed normal collagen fibers and thickened collagen fibers. Zone 3 consisted of normal-appearing collagen fibers. CONCLUSIONS: Ultrastructural examination of irradiated collagen revealed distinct morphological zones of denatured collagen fibers. Partially denatured fibers had an increased diameter consistent with lineal shrinkage. Zonal distinction was undetectable by light microscopy. Ultrastructurally, the zones of denatured collagen located above the normal fibers correlated with the zone of altered material seen on light microscopy. These findings suggest that collagen fiber shrinkage does occur after pulsed CO2 laser irradiation and that this phenomenon contributed, at least in part, to the immediate tissue contraction observed clinically.

Nodular basal cell carcinoma in vivo vs in vitro. Establishment of pure cell cultures, cytomorphologic characteristics, ultrastructure, immunophenotype, biosynthetic activities, and generation of antisera.

BACKGROUND AND DESIGN: In this study we developed an in vitro model of nodular basal cell carcinoma (BCC). We obtained pure cultures of BCC cells and compared the morphologic characteristics, ultrastructure, immunophenotype, and behavior of cultured tumor cells with those of their in vivo counterparts. Tumors were excised from patients undergoing Mohs micrographic surgery. We established 69 primary cell cultures from 32 patients with nodular BCC. RESULTS: Three cell types grew in primary cultures: fibroblasts, normal-appearing keratinocytes, and cells with dual (spindle and epithelioid) morphologic characteristics. Contaminating fibroblasts were removed using 0.125% trypsin-0.02% edetic acid, and normal-appearing keratinocytes were cornified and eliminated by temporarily increasing the concentration of calcium in the growth medium. The cells with dual morphologic characteristics remained intact and exhibited relentless growth in pure cultures. That these seemingly immortal cell strains represent true nodular BCC was demonstrated by (1) their biphasic morphologic characteristics and very slow cell growth rate, (2) their capability for anchorage-independent growth in soft agar, (3) their ultrastructural similarities to freshly excised nodular BCC, (4) their ability to generate antibodies selectively labeling nodular BCC tumor nests in vivo, and (5) their immunophenotypic similarities to BCC in vivo on more than 20 different cell markers. CONCLUSIONS: This study provides a simple technique for establishing pure cell cultures of nodular BCC and describes extensively the in vitro parameters of tumor cell growth. The striking differences in behavior of cultured tumor cells in the presence or absence of normal-appearing keratinocytes suggest that normal human epidermal keratinocytes can suppress the growth of BCC cells.

Gene rearrangement analysis in lymphoid neoplasia.

Current uses for gene rearrangement analysis in clinical dermatology are listed in Table 3. This technique is useful for determining the existence of clonal populations within a background of polyclonal lymphoid cells; therefore, it is helpful in the diagnosis and staging of patients with CTCL and PTCL. Although dual genotypes do occur, this technique is usually capable of determining lineage in a clonal lymphoid infiltrate and elucidating and characterizing the etiopathogenesis of certain neoplasms. On the basis of this review of the literature and our own experience, we conclude that gene rearrangement analysis shows great promise for monitoring response to therapy and detecting progression or relapse in patients with CTCL and PTCL. With the recent technology of PCR, it is possible to amplify specific sequences of DNA, detect molecular alterations in individual malignant T cells, and even identify exogenous retroviral gene sequences in tissues of patients with CTCL. Although gene rearrangement analysis has supported or established the clonal nature of lymphomatoid papulosis, pre-Sézary syndrome, granulomatous slack skin syndrome, and follicular mucinosis, the clinical significance of these findings is not yet clear. In the case of primary cutaneous B-cell lymphoma and its benign counterpart, B-cell pseudolymphoma, further investigation will be needed to determine the clinical significance of clonal rearrangements.

Oral drug reactions.

Oral drug reactions have many clinical manifestations and are produced by numerous medications. These reactions may be the result of an allergic reaction to systemically administered drugs or as an indirect effect of the action of the drug on other tissues. Other oral drug reactions may be the result of local or topical medications. These reactions are either a result of an allergic, delayed-type hypersensitivity, or a local primary irritation. The appearance may be nonspecific or it may resemble several distinct clinical entities. The diagnosis of these oral drug reactions is made with a good clinical history and examination, along with a high index of suspicion. Often there are multiple factors involved that complicate the clinical picture. The clinician who is familiar with the types of oral drug reactions caused by medications, the mechanisms by which these reactions occur, and which medications are most likely to cause the reaction will be prepared to make the correct diagnosis and treatment recommendations.

T-cell receptor gene rearrangement analysis in the early diagnosis of cutaneous T-cell lymphoma.

The authors describe the techniques of T-cell gene rearrangement in the diagnosis and early detection of cutaneous T-cell lymphoma. Current and future applications of these techniques are discussed.

Laser resurfacing for dermal photoaging.

The skin is a highly organized system composed of resident cells, extracellular matrix, blood vessels, and circulating cells that all work together to maintain cutaneous integrity. Environmental insults, particularly sunlight, act to alter the skin permanently, producing visibly undesirable effects. By wounding the skin or inducing a healing response with minimal wounding, the repair process can be activated to return the skin to a more normal condition. Owing to the complexity of the healing response, even the most well-studied and precise laser system can result in unpredicted results when used to treat photo-damaged skin. Through continued research into the normal functioning of skin, the alterations brought about by chronic photodamage, and the repair process, an integrated approach to treatment of photoaging will evolve. Agents such as alpha-hydroxy acids, retinoids, and growth factors that impact the healing response can be combined with various lasers to optimize improvement of photo-damaged skin, while minimizing the adverse consequences of treatment.

The highly effective use of topical zinc pyrithione in the treatment of psoriasis: a case report.

Psoriasis is a common condition which, at times, can be difficult to treat. Patients with psoriasis face many social challenges and can suffer a great deal with their disease. As a result, any new topical agent is a welcomed addition to our therapeutic armamentarium. We present here a report of a case using a novel topical preparation of zinc pyrithione for the treatment of psoriasis. Topical zinc pyrithione appears to be a safe and effective treatment for psoriasis.

Actinic cheilitis. Treatment with the carbon dioxide laser.

Actinic cheilitis is a premalignant condition that can be treated in several ways. A total of 43 patients with biopsy-proven actinic cheilitis were treated with the carbon dioxide (CO2) laser. After follow-up of at least 10 months, 26 patients thought that the lip was cosmetically improved, and 40 thought that the function of the lip was improved or had not changed. Complications were few and included only mild hypertrophic scarring, which resulted most often from the diagnostic biopsy and was corrected with topical or intralesional steroids or no therapy except simple massage. The CO2 laser is a simple, inexpensive, effective therapy for actinic cheilitis.

Lipophagic panniculitis in re-excision specimens.

Lipophagic panniculitis consists of a macrophage infiltrate in the subcutaneous tissue. The macrophages transform into foam cells within the panniculus; they replace lipocytes and may form giant cells. Although those pathologic features have been described as diagnostic of Weber-Christian disease, we report the occurrence of lipophagic panniculitis in re-excision specimens. Among 252 re-excision specimens from previously biopsied skin tumors, 5 cases in which masses of lipophages were infiltrating and replacing the subcutaneous tissue were found. The infiltrate was localized to the deep dermis and superficial subcutaneous tissue below and beside the initial biopsy site. In 3 cases, suture or hair was detected within the tissue, and granulation tissue with foreign body giant cells was observed along the dermal suture line. In 4 cases there was evidence of phlebitis within or close to areas of infiltration. None of these patients developed symptomatic panniculitis. Lipophagia can be a normal response of wound healing in some patients.

Generalized pustular psoriasis in childhood. Report of thirteen cases.

Generalized pustular psoriasis is rare in children. Less than 100 cases have been reported. We describe 13 children with this type of psoriasis. Seven had acute onset of widespread sterile pustules coalescing into lakes of pus with subsequent exfoliation (the Zumbusch pattern). This usually occurred in infancy and was difficult to control; recurrences developed several times per year. Three had the subacute benign annular pattern. They tended to be older and often had resolution within several years. Three had a mixed pattern with Zumbusch flares preceded by an annular or acral pattern. Most patients had an eruption preceding the generalized pustular psoriasis and often had precipitating factors. Generally, generalized pustular psoriasis has little serious chronic morbidity. The condition in most patients was well controlled with topical therapy. Systemic steroids were not helpful.

Patch testing in prurigo nodularis.

32 patients with prurigo nodularis evaluated at the Mayo Clinic from 1975 to 1987 have been patch tested for sensitivity to appropriate allergen series; 25 of these had relevant positive reactions and subsequent follow-up to 5 to 14 years was available for 11. 6 patients had persistent disease and 5 had resolution or marked improvement. 3 of these latter patients noted a strong positive correlation between improvement and avoidance of contact allergens. Screening for contact sensitivity may be helpful in the management of this refractory dermatosis, particularly if there is a coexistent dermatitis.

Hidroacanthoma simplex: an ultrastructural and immunohistochemical study.

A representative case of hidroacanthoma simplex was studied with routine light microscopy, immunohistochemistry, and electron microscopy. Staining with the periodic acid-Schiff reagent and immunostaining with anti-keratin antibodies were useful in demarcating the tumor cells from adjacent normal epithelium. However, antibodies to carcinoembryonic antigen and epithelial membrane antigen did not help us to segregate or identify the neoplastic cells. Electron microscopy revealed tumor cells markedly different in appearance from luminal cells of the acrosyringium. Hidroacanthoma simplex does not appear to be derived from luminal cells of the acrosyringium. We propose criteria for the histologic diagnosis of this benign neoplasm.

Intermittent leukapheresis: an adjunct to low-dose chemotherapy for Sézary syndrome.

Eleven patients with Sézary syndrome were treated with intermittent leukapheresis in addition to low-dose chlorambucil and prednisone. The results were as good as or better than those with chemotherapy alone. We believe the combined program with continuous leukapheresis to be optimal therapy but note that intermittent treatment offers some benefit for patients.

Transient acantholytic dermatosis associated with lymphomatous angioimmunoblastic lymphadenopathy.

Transient acantholytic dermatosis is a papulovesicular cutaneous eruption first described in 1970. There have been subsequent reports of similar disorders occurring in patients with malignancy. Angioimmunoblastic lymphadenopathy with dysproteinemia is a disorder characterized by an acute onset of generalized lymphadenopathy associated with fever, malaise, pruritus, night sweats, and hepatosplenomegaly. The patient described had a papular acantholytic dermatosis associated with the development of angioimmunoblastic lymphadenopathy with dysproteinemia-like T-cell lymphoma. The cutaneous manifestations of angioimmunoblastic lymphadenopathy with dysproteinemia are discussed.

Primary B-cell lymphoma with histologic features of a T-cell neoplasm.

A 58-year-old white man had dermatomyositis and primary cutaneous B-cell lymphoma. The cutaneous lymphoma was evidenced by a noduloulcerative disease of the lower extremities. Histologic results resembled a T-cell process with a diffuse, superficial infiltrate composed of small- and medium-sized lymphocytes with angioinvasion and epidermotropism. The infiltrate extended into the deep dermis and panniculus with scattered large lymphocytes and necrosis. With the help of gene-rearrangement analysis and immunophenotyping, the true B-cell lineage was discovered. The importance of gene-rearrangement analysis and immunophenotyping in the diagnosis of cutaneous lymphoma is emphasized.