We have previously shown expression of CRH and POMC genes and peptides
in the human skin. To ascertain the identity of those peptides, we used
methods of peptide extraction and purification combined ...with the highly
specific technique of liquid chromatography-mass spectrometry.
Testing extracts of human skin, we identified endogenous peptides with
masses and retention times corresponding to CRH, ACTH 1–39, ACTH
1–13, and α-MSH standards. Thus, conclusive evidence is provided for
the presence of CRH and the POMC-derived ACTH 1–39, ACTH 1–13, andα
-MSH peptides in human skin. Direct identification of these peptides
is consistent with translation of the corresponding genes, and it also
suggests intermediate pituitary lobe-like POMC peptide processing.
Since measurement of lysophosphatidate phosphatase activity is important in studies of tumorigenesis, we attempted to develop a simpler alternative to the more complex methods currently available. ...Measuring the phosphate released would permit use of the same method for a variety of phosphatases with physiological substrates, many of which are nonchromogenic. The Malachite green method of K. Itaya and M. Ui (1966, Clin. Chim. Acta 14, 361) has adequate sensitivity for quantitating phosphatase activity in biological samples. In samples with high endogenous phosphate concentrations pretreatment with 50 mg Dowex 1 × 10 (100–200 mesh, OH− form) usually permitted reliable determination of phosphatase activity. For 34 consecutive runs the mean relative difference (phosphorus activity − vitamer activity)/phosphorus activity obtained from the simultaneous measurement of both the phosphate released and the corresponding organic product (pyridoxal and pyridoxine) was −0.03 ± 0.09. The within run and between run coefficients of variation (three runs of four to five replicates) were 0.05 and 0.04, respectively. Pyridoxine 5′-phosphate hydrolase activity (pH 10) in cultured skin cells (normal and cancerous) ranged from 2 to 12 nmol phosphorus/min · mg protein. Lysophosphatidate phosphatase activity (pH 7.4) ranged from 3 to 14 nmol phosphorus/min · mg protein. The current approach permits the measurement of phosphatase activity with a single method using a variety of substrates and incubation conditions.
Here we show that cyclophosphamide induces disruption of follicular melanogenesis, which is characterized by abnormal transfer of pigment granules to ectopic hair bulb locations, extrafollicular ...melanin incontinence, disordered formation of melanosomes, and inhibition of melanosome transfer into precortical keratinocytes. This is in contrast to dexamethasone-induced termination of follicle melanogenesis, which activates premature but predominantly normal catagen development. Cyclophosphamide-induced pigmentation disruption was accompanied by significant alterations of biochemical and biophysical markers of melanogenesis, compared to control mice treated either with vehicle or with topical dexamethasone. Electron paramagnetic resonance spectroscopy shows a decline in the melanin signal and predominant eumelanin production. Tyrosine hydroxylase activity of tyrosinase and dihydroxyphenylalanine oxidation drop rapidly, while DOPAchrome tautomerase activity increases and dihydroxyindole carboxylic acid conversion factor activity remains unchanged in cyclophosphamide-treated mice compared to controls. These observations emphasize the key role of tyrosinase as opposed to post-dihydroxyphenylalanine oxidase steps in normal and pathological termination of melanogenesis and shows that tyrosinase is the most sensitive target of the melanogenic apparatus for pharmacological regulation. Follicle pigmentation recovers only during the subsequent hair cycle, i.e., after a new anagen hair bulb has been constructed, which points to the existence of a relatively chemoresistant melanoblast-like cell population residing in the noncycling part of the hair follicle.
We postulate that in mammalian systems neurotransmitter and hormone-like functions of L-tyrosine (LT) and L-DOPA (LD) are mediated via specific membrane-bound and/or nuclear receptors. The structure ...and function of these receptors may represent an evolutionary continuum of regulatory proteins binding LT or LD in unicellular and lower multicellular organisms.
Lichen sclerosus (LS) shares with vitiligo a milky-white appearance. By biopsy, pathognomonic dermal sclerosis readily distinguishes LS from vitiligo and other causes of leukoderma. To determine what ...the mechanism of hypopigmentation is in LS, we examined samples from LS cases for alterations in melanin content (Fontana-Masson stain) and melanocyte number (HMB-45 PMEL-17/gp100, Mel-5 TRP-1, Mart-1 Melan A) and compared these findings with those in controls of normal skin, acute scars, vitiligo, and lichen planus (LP; a common inflammatory cause of hyperpigmentation). The degree and extent of melanization found in LS overlapped with that in acute scars showing predominantly hypomelanized keratinocytes, with that in LP containing regions with numerous melanophages, and with that in vitiligo exhibiting focal regions of keratinocytes devoid of melanin pigment. By hematoxylin-eosin staining and immunocytochemistry for Mel-5 and Mart-1, LS had a lower mean count of melanocytes than acute scars, LP, and normal skin per 200 basal keratinocytes. In addition, a few LS cases had a significant loss of melanocytes comparable to that of vitiligo. Surprisingly, Mart-1 identified rare melanocytes in 67% of vitiligo cases and a significantly larger pool of melanocytes in LS and controls other than those labeled by Mel-5. Furthermore, LP and evolving lesions of LS contained the highest Mart-1 counts. HMB-45-immunoreactive melanocytes were found in the majority of acute scars and in LP and late-stage LS lesions at significantly lower levels than Mel-5- and Mart-1- labeled melanocytes, but they were not found in vitiligo or normal skin. We propose that several mechanisms may play a role in the production of leucoderma in LS: 1) decreased melanin production; 2) block in transfer of melanosomes to keratinocytes; and 3) melanocyte loss. The latter finding may be the pathogenic connection (lichenoid dermatitis of LS triggering an autoimmune reaction to melanocytes) that underlies the documented association of LS with vitiligo.
The incidence and mortality of melanoma has seemed to level off for certain groups after a steady increase during the last 50 years. This trend is suspected to be secondary to education efforts aimed ...at prevention and better detection and removal of thin, biologically benign melanomas. Since no effective systemic therapies exist for metastatic melanoma, early detection and removal of thin melanomas offer the best chance of cure. For thicker melanomas, sentinel lymph node biopsy has improved the accuracy of staging and prognostic evaluation. However, approximately one third of patients diagnosed with metastatic melanomas present without previous regional lymph node metastases. As the genomic understanding of melanoma's pathogenesis grows, new methods likely will be developed to more accurately identify the people at risk for melanoma, those who have high-risk melanomas, and those who have disseminated disease. We review current and potential biomarkers useful for the screening for and prevention, diagnosis, staging, and prognosis of melanoma.
20-Hydroxyvitamin D(3) (20(OH)D(3)), the major product of CYP11A1 action on vitamin D(3), is biologically active and is produced in vivo. As well as potentially having important physiological ...actions, it is of interest as a therapeutic agent due to its lack of calcemic activity. In the current study we have examined the ability of CYP24A1, the enzyme that inactivates 1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)), to metabolize 20(OH)D(3). Rat CYP24A1 was expressed in Escherichia coli, purified by Ni-affinity chromatography and assayed with substrates incorporated into phospholipid vesicles which served as a model of the inner mitochondrial membrane. In this system CYP24A1 metabolized 1,25(OH)(2)D(3) with a catalytic efficiency 1.4-fold higher than that seen for 25-hydroxyvitamin D(3) (25(OH)D(3)). CYP24A1 hydroxylated 20(OH)D(3) to several dihydroxy-derivatives with the major two identified by NMR as 20,24-dihydroxyvitamin D(3) (20,24(OH)(2)D(3)) and 20,25-dihydroxyvitamin D(3) (20,25(OH)(2)D(3)). The catalytic efficiency of CYP24A1 for 20(OH)D(3) metabolism was more than 10-fold lower than for either 25(OH)D(3) or 1,25(OH)(2)D(3) and no secondary metabolites were produced. The two major products, 20,24(OH)(2)D(3) and 20,25(OH)(2)D(3), caused significantly greater inhibition of colony formation by SKMEL-188 melanoma cells than either 1,25(OH)(2)D(3) or the parent 20(OH)D(3), showing that CYP24A1 plays an activating, rather than an inactivating role on 20(OH)D(3).
Many reports exist of pigmented adnexal tumors containing dendritic melanocytes such as pigmented basal cell carcinomas and pigmented pilomatricomas. Correspondingly, melanocytes are a known ...component of the bulbs of anagen follicles. The phenomenon of melanization of adnexal tumors highlights the interrelationship between melanocytes and adnexal epithelium and may represent normal melanocytes colonizing a neoplastic proliferation. We report on two cases of a unique tumor composed of neoplastic matrical cells with a significant component of melanocytes. Both cases presented as pigmented papules in older men (66 and 80 years, forearm and pectoral region, respectively). Histologically, these were well-defined nodular proliferations composed of variably melanized, pleomorphic, and mitotically active matrical and supramatrical cells forming clusters of "shadow cells." Admixed with the epithelial cells were numerous melanized dendritic melanocytes. Shadow cells expressed keratin 13, and a subpopulation of S-100 protein-positive dendritic cells were evident. No recurrence of any type was found after reexcisions 4 months and 2 years later. We propose the name of melanocytic matricoma for these two heretofore unreported cases of a unique neoplasm composed of matrical cells and melanocytes recapitulating epithelial-melanocyte interaction in the follicular anagen bulb. Although their small size, circumscription and clinical course suggest a benign nature, melanocytic matricomas' cytologic atypia disclose the potential for malignant behavior.
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