Sergij Goerdt

Other Functions, Other Genes: Alternative Activation of Antigen-Presenting Cells

We would like to thank Drs. C. C. Geilen, S. Runkel, and N. Runkel for providing comments on this review.

Alternative versus Classical Activation of Macrophages

In parallel to the Th1/Th2 paradigm, antigen-presenting cells (APC) are divided into classically activated APC (dendritic cells/effector macrophages) and alternatively activated APC (IL-4-induced, alternatively activated macrophages/IL-10-induced, immature dendritic cells). Alternatively activated APC share a special molecular repertoire including receptors of innate immunity with broad specificity for foreign antigen and anti-inflammatory cytokines such as IL-1Ra and alternative macrophage activation-associated CC-chemokine-1. Alternatively activated APC mediated tolerance and downregulated inflammation. Abuse of alternatively activated APC in support of infectious susceptibility or tumor immune escape is counteracted by the classical pathway. Thus, classically and alternatively activated APC secure the balance between proinflammatory and anti-inflammatory immune reactions.

Alternatively activated macrophages differentially express fibronectin and its splice variants and the extracellular matrix protein betaIG-H3.

Alternative activation of macrophages, induced by Th2 cytokines and glucocorticoids, is essential for the proper functioning of anti‐inflammatory immune reactions. To this end, alternatively activated macrophages (aaMΦ) express a not yet fully unravelled set of genes including cytokines such as alternative macrophage activation‐associated CC‐chemokine (AMAC)‐1 and pattern recognition molecules such as the scavenger receptor CD163. In order to further characterize the molecular repertoire of aaMΦ, differential gene expression was analyzed by combining subtractive suppression cloning and differential hybridization. We show here that aaMΦ induced by interleukin (IL)‐4 overexpress the prototype extracellular matrix (ECM) protein fibronectin on the mRNA and protein level. This overall increase is accompanied by a shift in fibronectin splice variants from an embryonic to a mature pattern. In addition, the expression of another ECM protein, βIG‐H3, is also upregulated by IL‐4 in aaMΦ. In contrast to IL‐4 and in line with its inhibitory effect on wound healing, dexamethasone exerts a strongly suppressive effect on fibronectin and βIG‐H3 expression. In conclusion, overexpression of ECM proteins induced by IL‐4 in macrophages suggests that aaMΦ may be involved in ECM deposition and tissue remodelling during the healing phase of acute inflammatory reactions and in chronic inflammatory diseases.

DIFFERENCES IN ANGIOGENIC POTENTIAL OF CLASSICALLY VS ALTERNATIVELY ACTIVATED MACROPHAGES

Macrophages (M phi) are important for angiogenesis during inflammation, wound repair, and tumor growth. However, well-characterized M phi subsets such as IFN-gamma-induced, classically activated (ca) M phi or IL-4/glucocorticoid-induced, alternatively activated (aa) M phi have not been thoroughly examined for a positive or negative association with angiogenesis. While caM phi populate early inflammatory reactions and high-turnover granulomas, aaM phi occur in healing wounds and chronic inflammation. In contrast to caM phi-dominated lesions, aaM phi-rich lesions are highly vascularized. In order to determine their angiogenic potential in vitro, these M phi subsets as well as unstimulated control macrophages (coM phi) were analyzed by RT-PCR for mRNA expression of 10 angiogenic factors after 3 and 6 days of culture. Early during activation, caM phi and coM phi expressed equal levels of 8 of 10 angiogenic factors (PDGF-A, MK, TNF-alpha, TGF-beta 1, PDGF-B, HGF, TGF-alpha, IGF-1), while aaM phi showed expression of only 4 of these factors (TGF-beta 1, PDGF-B, HGF, GF-1). After maturation, TGF-alpha and IGF-1 showed a shift in mRNA expression from caM phi to aaM phi resulting in a considerably enhanced expression of these factors in day-6 aaM phi as compared to day-6 caM phi and coM phi while PDGF-A, MK, and TNF-alpha remained suppressed in day 6 aaM phi. In all M phi subsets including controls, mRNA expression of aFGF and bFGF was minimal or absent while TGFG-beta 1, HGF, and ODGF-B were constitutively expressed. In order to functionally integrate angiogenic factor mRNA expression profiles, mitogenic activity of M phi subsets towards microvascular endothelium was assessed by cocultivation. Coculture experiments revealed that endothelial proliferation induced by aaM phi was 3.0-3.5x higher than induced by caM phi. In conclusion, mature aaM phi are well equipped to play an important role in protracted M phi-associated angiogenic processes. Presumably due to expression of predominantly angio-inhibitory cytokines such as TNF-alpha by caM phi but much less by aaM phi, caM phi exhibit only a low angiogenic potential in vitro and in vivo despite considerable expression of angiogenic factor mRNA.

Clonal T-cell receptor γ-chain gene rearrangement by PCR-based GeneScan analysis in advanced cutaneous T-cell lymphoma: a critical evaluation

Detection of clonal T‐cell receptor γ (TCRγ)‐chain gene rearrangement is a promising approach to distinguish between cutaneous T‐cell lymphomas (CTCLs) and reactive T‐cell infiltrates. Despite the improved sensitivity by using the polymerase chain reaction (PCR) rather than Southern blot analysis, monoclonality could be demonstrated in only 53–90 per cent of CTCL biopsies in recent studies. In the present study, formalin‐fixed, paraffin‐embedded specimens of 21 selected patients with clear‐cut advanced‐stage CTCL were analysed using a semi‐nested TCRγ PCR with newly developed consensus primer pairs. Detection of PCR products was done by GeneScan analysis (GSA); this technique is advantageous due to its sensitivity and accuracy in the detection and size determination of PCR products and it is easier to interpret than direct read‐outs from TGGE or DGGE gels. In serial dilution experiments, TCRγ‐PCR‐GSA allowed the detection of clonal, rearranged T‐cells with a high in vitro sensitivity against a polyclonal background (1–6 per cent). Despite the selection of clear‐cut, advanced‐stage CTCL cases, however, dominant clonal TCRγ‐chain gene rearrangement was found in only 16 of the 21 patients analysed, indicating an overall clinical sensitivity of 76 per cent. Specificity was evaluated using biopsy specimens from 21 control patients suffering from long‐standing psoriasis (nu2005=u200513) and eczema (nu2005=u20058). Surprisingly, GeneScan profiles showing apparently single dominant peaks were detected in 14 per cent of these skin lesions, but these profiles turned out to be pseudo‐monoclonal by repeated determinations. In conclusion, TCRγ‐PCR‐GSA does not suffice reliably to exclude malignancy, due to its limited clinical sensitivity, but with precautions taken to detect pseudo‐monoclonality and to secure specificity, TCRγ‐PCR‐GSA is a valuable instrument in the diagnosis of CTCL. Copyright

Expression of vascular endothelial growth factor (VEGF) in various compartments of the human hair follicle

Abstract Hair follicle vascularization appears to be closely related to the processes involved in hair cycle regulation, in which growth factors, cytokines and other bioactive molecules are involved. In particular, vascular endothelial growth factor (VEGF), essential for angiogenesis and vascular permeability, may be responsible for maintaining proper vasculature around the hair follicle during the anagen growth phase. The aim of our study was to compare the in vitro angiogenic capacity, i.e. the steady-state expression of the VEGF gene, of different cultured cell types derived from normal human hair follicles, corresponding to different follicular compartments. Human dermal papilla cells (DPC), fibrous sheath fibroblasts, dermal fibroblasts, and follicular and interfollicular keratinocytes were cultured and studied in vitro for VEGF expression at the mRNA level using RT-PCR, and for VEGF protein synthesis by radioimmunoprecipitation and immunocytochemistry. In vivo examination for VEGF expression in human terminal hair follicles was performed using immunohistochemical methods. In the present report the expression of four different VEGF molecular isoforms, differing in their angiogenic capacity, are described in different cultured follicular cell types for the first time. Cultured follicular cells strongly expressed mRNA of four VEGF molecular species identified as the 121-, 145-, 165- and 189-amino acid splice variants, the most prominent being the 121-amino acid molecule. DPC, and also other mesenchymal cells such as fibrous sheath fibroblasts and dermal fibroblasts, in vivo and in vitro strongly expressed VEGF mRNA and synthesized a 46-kDa VEGF protein, whereas follicular and interfollicular keratinocytes in vitro expressed lower levels of VEGF mRNA and proteins than mesenchymal cells. As the highest expression of VEGF was found in DPC, we suggest that DPC are mainly responsible for angiogenic processes possibly related to the human hair cycle.

Melkersson-Rosenthal syndrome in childhood : a challenge in differential diagnosis and treatment

Facial palsy and orofacial swelling in childhood represent a challenge in differential diagnosis for paediatricians and dermatologists. One possible entity, Melkersson–Rosenthal syndrome (MRS), is a rarity in childhood. We describe a 9‐year‐old girl with the diagnosis of MRS who had episodic swelling of the upper lip and complete peripheral facial palsy, associated with herpes and recurrent bacterial infections. Therapeutic options for MRS in childhood are limited. Our patient benefited from a 2‐month course of prednisolone 1u2003mgu2003kg−1 daily. We review previously published cases of MRS in childhood, and discuss the differential diagnosis of orofacial swelling and facial palsy as well as treatment options in children.

Dissection of macrophage differentiation pathways in cutaneous macrophage disorders and in vitro

Abstract Macrophages play important roles in immunity and inflammation, and in allergic, granulomatous and neoplastic diseases. Here, we present the indepth results of an ongoing study of macrophage differentiation pathways in cutaneous macrophage disorders and in vitro. Up to now, a total of 40 cases of cutaneous macrophage disorders (histiocytoses and granulomas) and related diseases were examined using a panel of monoclonal and polyclonal antibodies to macrophage differentiation antigens (mAb MS‐1, mAb αCDla, mAb αCD34, mAb RM 3/1, mAb αCD11c, mAb αCD36, mAb MAC 387, mAb 27E10, polyclonal antibodies ccMRP‐8 and ‐14, mAb aCD68. mAb 25F9, mAb DRC1‐R4/23, and mAb 1F10). Of these, MS‐1 high molecular weight protein, synthesized by non‐continuous sinusoidal endothelial cells and highly dendritic perivascular macrophages in normal human organs, is the most specific macrophage differentiation marker. MS‐1 high molecular weight protein is selectively expressed by cutaneous non‐Langerhans cell histiocytoses, and proves to be a valuable diagnostic tool for these diseases. MS‐1 high molecular weight protein is not found in Langerhans cell histiocytosis cells, epithelioid cells in sarcoidosis, and palisading histio‐cytes in granuloma annulare. MS‐1+ macrophages may be found intermingled in cellular type dermatofibroma and in foreign body granulomas; they differ from MS‐1+ non‐Langerhans cell histiocytosis cells by their highly dendritic morphology, and thus rather resemble the MS‐1+ macrophages in normal skin. RM 3/1 antigen shows a similar, but broader expression pattern including non‐Langerhans cell histiocytoses, xanthel‐asmata palpebrarum, foreign body granulomas, granuloma annulare, and cellular type dermatofibroma. Moreover, xanthelasmata palpebrarum para‐digmatically represent a class of macrophage lesions with strong RM 3/1, but little MS‐1 antigen expression. In sarcoidosis, RM 3/1 + macrophages are only found at the very periphery of epithelioid cell granulomas. In contrast, 25F9 antigen is strongly and consistently expressed in epithelioid cells of sarcoidosis, and in foreign body granulomas. In cultured human monocytes/macrophages, RM 3/1 antigen is expressed early on, while MS‐1 high molecular weight protein and 25F9 antigen are late and very late macrophage differentiation antigens, respectively. Expression of RM 3/1 antigen and MS‐1 high molecular weight protein is inducible by glucocorticoid and interleukin‐4, and less so by interleukin‐13 and interlcukin‐10, and combinations thereof, while 25F9 antigen seems to be less influenced by these agents. Interferon‐y (and less so tumor necrosis factor‐ex) inhibit expression of all three antigens in cultured human monocytes/macrophages. In monocytic leukemia cell line THP‐1. RM 3/1 antigen is only induced by a combined treatment of phorbolester and glucocorticoid and peaks at 3‐7 days. In different subclones of the cell line, 25F9 antigen is either inducible by phorbolester alone or by combined treatment with glucocorticoid or it is constitutively expressed and hardly modulated. In contrast, MS‐1 high molecular weight protein cannot be induced in THP‐1 cells by the agents tested. Inhibition of MS‐1 high molecular weight protein, and RM 3/1 and 25F9 antigens by interferon‐y suggests that macrophages characterized by these phenotypic traits must by counted among the alternatively activated macrophage populations. As a result of our study, alternative macrophage activation should be viewed as a multifacetted process resulting in various, partially overlapping, partially complementary macrophage phenolypes both in vivo and in vitro.

Erworbene reaktiv perforierende Dermatose Erfolgreiche Behandlung mit Allopurinol in 2 Fällen

ZusammenfassungDie perforierenden Dermatosen bilden eine heterogene Gruppe von Erkrankungen, die durch transepidermale Eliminationsphänomene gekennzeichnet sind. Abzugrenzen sind die primär perforierenden Dermatosen von solchen perforierenden Dermatosen, bei denen Perforation oder Elimination sekundär und als seltenes Teilsymptom bei bestehenden Dermatosen anderer Provenienz auftreten können. Zu den primär perforierenden Dermatosen zählen dabei als Prototypen die Hyperkeratosis follicularis et parafollicularis in cutem penetrans (M. Kyrle), die Elastosis perforans serpiginosa und die perforierende Follikulitis. Die erworbene reaktiv perforierende Dermatose (syn. erworbene reaktiv perforierende Kollagenose) stellt zusammen mit der hereditären, reaktiv perforierenden Kollagenose eine weitere Variante der primär perforierenden Dermatosen dar. Wir stellen hier eine 84jährige und eine 96jährige Patientin mit einer erworbenen reaktiv perforierenden Dermatose vor. Bei beiden Patientinnen lagen eine diabetogene und eine hyperurikämische Stoffwechsellage vor. Eine Therapie mit Allopurinol (100 mg/d) führte bereits nach kurzer Dauer (1 bis 2 Wochen) zu einer weitgehenden Abheilung der disseminierten Läsionen. Über einen Nachbeobachtungszeitraum von jeweils 6 Monaten befanden sich beide Patientinnen in Vollremission. Diese Beobachtungen belegen erneut die Existenz und Massivität der Erkrankung und zeigen, daß Allopurinol neben der bekannten urikostatischen Wirkung möglicherweise in der Haut immunologische oder differenzierungsmodulierende Effekte vermittelt.SummaryPerforating disorders represent a heterogenous group of dermatoses characterized by transepithelial elimination of dermal structures. Primary perforating disorders should be distinguished from secondary perforating disorders in which perforation with transepithelial elimination is a rare component of a variety of dermatoses. The primary perforating disorders are hyperkeratosis follicularis et parafollicularis in cutem penetrans (Kyrle’s disease), elastosis perforans serpiginosa and perforating folliculitis. Acquired reactive perforating dermatosis (also known as acquired reactive perforating collagenosis) together with the hereditary variant of the reactive perforating collagenosis represent further examples of the primary perforating disorders. We report on 84 year old and 96 year old female patients with an acquired perforating dermatosis. Both of the patients additionally showed diabetes and hyperuricemia. Oral administration of allopurinol (100 mg daily) led to a healing of the disseminated skin lesions in 1–2 weeks. After a follow-up period of 6 months, both patients were in complete remission. On one hand, these results prove again the existence and the severity of this disease, and on the other hand suggest an immunomodulating or differentiation-promoting action in addition to the uricostatic effect of allopurinol.

Nodular and bullous cutaneous mastocytosis of the xanthelasmoid type: case report.

Severe generalized nodular and bullous mastocytosis of the xanthelasmoid type is described in a 7‐month‐old boy. Reddish to yellowish‐brown xanthelasmoid papules and nodules first developed in the inguinal region a few weeks after birth and then progressively spread to cover nearly the entire body surface. There was severe pruritus and recurrent episodes of blistering. The diagnosis of cutaneous mastocytosis of the xanthelasmoid type with subepidermal bullae was confirmed by skin biopsies showing solid and deeply penetrating infiltrates of metachromatic mast cells under light and electron microscopy. Systemic involvement of other organs, however, was excluded by bone scintigraphy, abdominal ultrasound, bone marrow aspiration and echocardiography. The extensive skin involvement was reflected in highly elevated urinary levels of histamine (263·4u2003μgu2003L−1) and its metabolite N‐methylimidazole acetic acid (20·8u2003mgu2003L−1). The patient was systematically well and received only symptomatic treatment. Over a period of 1 year, the condition gradually improved, and the skin lesions began to flatten and regress.