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Background: Arterial punctures associated with blood donation are uncommon, and characterized by bright red color and rapid blood flow. We wanted to establish the normal range of whole blood unit collection times (CT), with a specific interest in determining the criteria for units with unusually rapid blood flow, thus assisting phlebotomy staff in the recognition of a possible arterial puncture. Methods: Nine blood service regions participated in the study. Each region selected a day during the week of 061200 for determination of the CT for all allogeneic units on that day. Units considered to be not sufficient in quantity, based on a weight <475 grams, were excluded. CT were calculated from the start and end times of phlebotomy, as documented on the blood donation record, and recorded in whole minute increments (min). Results: CT of 4771 units were evaluated. The distribution curve was skewed towards longer collection times, and ranged from 3 to 34 min. Statistical analysis showed that the mode was 6 min, the median was 7 min, and the mean was 7.5 min. CT longer than 11 min are outliers, and CT longer than 14 min are far outliers, as they are at least 2.5 standard deviations longer than the mean. A CT of 3 min is on the border for an outlier on the lower end. None of the phlebotomies were noted to be arterial punctures. Conclusions: The majority (84%) of whole blood unit collections were completed within a 5 to 10 min time interval. Four min collections occurred 3.6% of the time, and are still consistent with venous draws. Three min collections were very unusual, occurring in less than 1/2000 donations. Donors of units collected in less than 4 min have unusually rapid flow, and should be evaluated for other signs of a possible arterial puncture. [ABSTRACT FROM AUTHOR]

Epigenetic regulation of gene expression by DNA methylation is a central mechanism governing the silencing of tumor suppressor genes in many forms of cancer. Current methods have not proven optimal for the quantitative analysis of DNA methylation and corresponding in situ protein expression within cells in small specimens like skin biopsies. We have overcome this limitation by combining and modifying several techniques: target cell enrichment, DNA micro-isolation, one-step denaturation/bisulphite conversion/in-column desulphonation, specially designed PCR amplification, pyrosequencing and multispectral image analysis. Using this approach optimized for small samples, we can quantify minor alterations in gene methylation and protein expression using minimal amounts of tissue. Comparative studies of fresh and processed cells showed that our method is valid for DNA in both fresh and formalin-fixed, paraffin-embedded specimens. We can measure the effects of DNA methylation inhibitors, administered in vitro or in vivo, on the promoter methylation and protein expression of selected genes in specific cells. This novel approach should prove useful for a wide variety of investigative and clinical applications in dermatology and other specialties where the collection of small, routinely processed biopsy specimens is common. We refer to this method as Q-GAME (quantitative gene analysis of methylation and expression)., (© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Low proliferation rates, sparse apoptotic cells, and resistance to chemotherapeutic agents suggest that defective apoptotic mechanisms may be important in the pathogenesis and progression of cutaneous T-cell lymphomas (CTCLs). Functional studies of CTCL cell lines and leukemic cells further support abnormal expression of Fas apoptotic pathway proteins as a mechanism for resistance to apoptosis., Objective: We sought to compare the Fas apoptotic pathway protein expression of mycosis fungoides (MF) with CD30(+) lymphoproliferative disorders (CD30(+)d) in the context of insights gained from functional studies of CTCL cells., Methods: We conducted immunohistochemical analysis of 36 MF and 36 CD30(+)d skin biopsy specimens with antibodies against Fas, Fas ligand, FLICE-like inhibitory protein, Fas-associated death domain, and total and cleaved caspases 8 and 3., Results: Almost all MF lesions (94%) were Fas ligand-negative. In all, 64% of MF lesions were Fas low. An additional 25% of MF lesions were both Fas high and FLICE-like inhibitory protein high (Fas pathway inhibitor). Altogether, this equated to a phenotype predictive of apoptotic resistance in 89% of MF samples. In 9 of 10 cases of CD30(+)d, the apoptotic phenotype predictive of sensitivity/resistance correlated with signs of regression/progression, respectively., Limitations: The study is limited by its retrospective design., Discussion: Our in situ analysis of MF and CD30(+)d tissue samples correlates well with previous functional studies of CTCL cell lines and leukemic blood. Our data strengthen the hypothesis that abnormal expression of upstream Fas pathway factors (Fas, Fas ligand) and the inhibitor FLICE-like inhibitory protein contributes to defective apoptosis in CTCLs., (Copyright © 2012 American Academy of Dermatology, Inc. Published by Mosby, Inc. All rights reserved.)

A case of primary cutaneous anaplastic large-cell lymphoma with sarcomatoid histologic features is described. Sarcomatoid anaplastic large-cell lymphoma has previously been reported only ten times in the literature. It is a diagnostic challenge because of sarcomatoid features that include atypical spindle-shaped cells, a storiform architecture, and a mucinous stroma. Clues pointing toward the diagnosis of lymphoma may be very subtle. Awareness of this entity can lead to more accurate recognition of a potentially fatal disease.

Polo-like kinases are serine/threonine kinases crucial for mitosis and DNA integrity. Plk1, the most well studied member of this family, is upregulated in several cancers, as well as in dividing cells with peak expression during G2/M phase. Recently, employing lesional skin from patients with cutaneous T-cell lymphoma (CTCL), we showed that Plk1 was increased mainly in advanced lesions. In this study, employing western blot and quantitative RT-PCR analyses, we demonstrated that Plk1 was overexpressed in multiple CTCL cell lines (HH, Hut78, MyLa, SeAx and SZ4). Further, a genetic knockdown (by short hairpin RNA) or enzyme activity inhibition (via a small molecule inhibitor, GW843682X) was found to result in a decrease in cell growth, viability and proliferation. Plk1 inhibition in CTCL cells also resulted in: (1) increased G(2)/M phase cell cycle arrest, (2) alteration in key mitotic proteins, (3) apoptosis and (4) multiple mitotic errors. Given our findings, clinical trials of Plk1 inhibitors in CTCL may be a promising area for further translational investigation. We speculate that overexpression of Plk1 may prove to be relevant to the progression and prognosis of CTCL through its direct impact on the regulation of tumor cell proliferation and indirect influence on the acquisition of somatic mutations by proliferating tumor cells.

Cutaneous angiosarcoma is probably the most malignant neoplasm involving the skin. Three clinical variants of cutaneous angiosarcoma are recognized, including angiosarcoma of the scalp and face of elderly patients, angiosarcoma associated with chronic lymphedema, and postirradiation angiosarcoma. Histopathologically, these three variants of angiosarcoma show similar features, which consist of poorly circumscribed, irregularly dilated, and anastomosing vascular channels lined by prominent endothelial cells that dissect through the dermis. Focally, neoplastic endothelial cells show large, hyperchromatic, and pleomorphic nuclei, protruding within vascular lumina and creating small papillations. Usually, inflammatory infiltrate is sparse and consists of a patchy, perivascular lymphoid infiltrate around the neoformed vessels. In rare instances, cutaneous angiosarcomas may exhibit prominent inflammatory infiltrate, and the neoplasm may be mistaken for an inflammatory process, both from clinical and histopathologic points of view. We describe four examples of cutaneous angiosarcomas with dense lymphocytic infiltrates involving the neoplasm. Immunohistochemically, lymphocytes expressed immunoreactivity for CD3, CD5, and CD45 markers, whereas the germinal centers were positive for CD20, CD79a, and Bcl-6. The neoplastic endothelial cells expressed immunoreactivity for the CD31, CD34, podoplanin, Prox-1, Lyve-1, and D2-40. We discuss the possible relationship between neoplastic endothelial lymphatic cells and reactive lymphocytes. Cutaneous angiosarcoma with prominent lymphocytic infiltrate may be readily mistaken for cutaneous follicle center cell lymphoma or cutaneous pseudolymphoma.

Alopecia areata (AA) is a T cell-mediated autoimmune disease that can be treated with the contact sensitizer diphenylcyclopropenone (DCP). Peripheral blood leukocytes from AA patients are relatively resistant to apoptosis which might be due to decreased Fas Ligand (FasL) expression, or to an increase in CD44v7 expression. Moreover it has been suggested in a murine model of AA that contact allergen treatment might interfere with the emigration of Langerhans cells into the draining lymph node, thus hampering autoreactive T-cell activation. To assess whether and which of these mechanisms is of clinical relevance, immunohistochemistry was performed in scalp biopsies of successfully DCP-treated AA patients in the early phase of hair regrowth. In line with recent studies in a murine model of AA, there was no evidence that DCP treatment would interfere with extravasation and skin homing of activated leukocytes. Perifollicular infiltrates of DCP-treated as compared to untreated AA patients actually showed an increased number of perifollicular CD8(+) and CD1a(+) cells. Furthermore, the expression of CD44 and CD49d, which are of major importance in leukocyte extravasation, was even increased in DCP-treated as compared to AA patient infiltrates. The same accounted for the skin homing receptor CD44v10. When we evaluated the leukocyte subpopulations in DCP-treated as compared to untreated AA patients' skin biopsies, there was an undue increase in CD1a(+) cells, that could well be indicative of hampering of the emigration of antigen presenting cells (APC) by allergen treatment. Most importantly, the number of perifollicular TUNEL- and FasL-positive cells was strikingly increased, whereas the number of CD44v7(+) cells remained unaltered. Taken together, this study provides strong evidence that long term treatment with a contact sensitizer allows for the recovery of hair follicle by driving autoreactive T cells into activation-induced cell death. In addition the replacement with newly activated autoreactive T-cells might be impaired due to a DCP-mediated hindrance of APC emigration.

Although midcarpal wrist arthrodesis is recognized as a standard procedure to treat scapholuate advanced collapse (SLAC) and scaphoid nonunion advanced collapse (SNAC) of the wrist, little has been reported about patients with bilateral involvement and the number, cause, and results of failed cases requiring conversion to total wrist arthrodesis. This study investigated the results of 20 patients with bilateral procedures and of 22 patients who underwent total wrist fusion after failed midcarpal arthrodesis out of an overall group of 907 patients treated by this method during a 12-year period. Of these, 16 bilateral and 20 converted cases were reexamined after an average of 48 months and 42 months, respectively. Patients after bilateral midcarpal arthrodesis experienced a pain reduction by an average of 54% of the preoperative pain values at rest and by 56% at stress on the visual analog scale (scale range: 0 to 100) and from intolerable (3.7) to pain only during stress (1.9) on the verbal scale (scale range: I to 4). A mean arc of wrist extension and flexion of 53 degrees on the right and 49 of the left wrist was preserved. The mean DASH score was 45 points and 70% of the patients felt impaired only during certain activities. Total arthrodesis reduced pain in 18 of 20 reexamined wrists by 67% of the previous values after the failed partial arthrodesis at rest and by 46% at stress on the visual analog scale andfrom intolerable pain (3.7) to pain only during stress (2.1) on the verbal scale. Seven of the 20 reexamined patients noted complete pain relief at rest and two also under stress conditions. The DASH score averaged 39 points. A mean Krimmer score of 46 points and a mean Buck-Gramcko and Lohman evaluation of 6 points represented a satisfactory result. Grip strength of the operated hand averaged 53% of the opposite side. Subjectively, 30% felt impaired only during certain activities, 55%felt considerably and 15% strongly limited in daily life. However, all but two patients were satisfied with the secondary total wrist fusion as pain was considerably reduced. Midcarpal arthrodesis reliably reduced pain and preserved valuable wrist mobility thus improving daily activity and quality of life also in bilateral carpal collapse. In the rare cases when midcarpal arthrodesis failed, total wrist arthrodesis markedly improved the complaints in most patients, but in contrast to other studies complete pain was seldom.