Proliferative merkel cells were not detected in human skin

The fetal development of Merkel cells-neuroendocrine cells of the skin—has been a matter of debate for a long time. Recent results have helped to confirm their intraepidermal development in humans. Simple epithelial cytokeratins (CK) nos. 8, 18, 19 and 20 are well established markers at the light microscopic level. These cells could be detected from fetal week 8 within the epidermis with an enormous increase during the following weeks. This gives rise to the question as to whether Merkel cells are undergoing mitoses or whether they are derived from basal keratinocytes. We studied fetal and adult skin using antibodies to simple epithelial CK and to Ki67, a human nuclear cell proliferation-associated antigen in an attempt to answer these questions. In human adult and fetal skin of various stages we could not detect any Merkel cells undergoing cell division. These results suggest that Merkel cells are postmitotic cells to be renewed from undifferentiated keratinocytes with stem cell characteristics.     

Distribution of Merkel cells in acute UVB erythema

Summary Merkel cells (MC) were identified immunohistochemically using antibodies specific for cytokeratin (CK) 20 within human epidermis 12 to 72 h after exposure to UVB (4 MED). 12 h after exposure all MC were normally localized within the epidermal basal layer. However, 24 h after exposure 4% of the MC were detected suprabasally, the remaining 96% still being situated in the basal layer. Surprisingly, at 48 h and 72 h more than 50% had lost contact with the basal membrane. The MC of hair follicles did not show any obvious changes. These results argue, in the context of acute epidermal UV damage, for an abnormal turnover in dermatitis.     

Merkel cells in ontogenesis of human nails

Digital skin of human fetuses is known to contain a particularly high concentration of Merkel cells. Using antibodies against the simple epithelial cytokeratins (CK) 18 and 20, which are sensitive and specific Merkel cell markers, we studied imiminohistochemically the main adnexal structure of digital skin, the nail anlage, in human fetuses (9–22 weeks of gestation) for the presence of Merkel cells. As early as week 9 some clustered Merkel cells were detected in the early matrix primordium. In specimens of week 12–15, abundant Merkel cells were found in the nail anlagen, particularly in the epithelium of the proximal nail-fold and the dorsal and ventral side of the apex region. In contrast, Merkel cells were essentially absent from the epithelium of the ventral matrix (surface-near portion), lunula and nail bed. Correspondingly, in these regions, the adjacent dermis contained hardly any nerve fibres, whereas such fibres, as detected by neurofilament antibodies, were quite numerous adjacent to the proximal nail-fold epithelium. At week 22, the Merkel cell number in the nail anlage had decreased, and in adult nail matrix such cells were very rare. No Merkel cells were found in the dermal tissue surrounding the nail anlage while finger-tip skin of week 15, and particularly of week 22, exhibited single Merkel cells in the upper dermis next to clusters of such cells in the glandular ridges. We also found that Merkel cells were negative for CK 17. These results suggest a possible role for fetal Merkel cells in the proper development of sensory nerve fibres, while they do not seem to be important for the growth of trichocytic tissues, i.e. the hair bulb and the trichocytic portion of the nail anlage.     

Early development of human Merkel cells

Human fetal Merkel cells are now generally considered to be epidermal derivatives. Previous studies using antibodies against the simple epithelial cytokeratins (CKs), 8 and 18, have demonstrated the presence of these cells in the epidermis at as early as fetal week 10 to 12. Using antibodies against CK 20 whose expression within the skin is restricted to Merkel cells, we applied immunofluorescence and immunoperoxidase microscopy to analyze earlier embryonic and fetal human skin (wk 7 to 9). We were able to demonstrate the first Merkel cells at as early as fetal wk 8, i.e., at the same time as the epidermis starts to develop an intermediate, third layer, characterized by the expression of CKs 1, 10, and 11. Most of these early Merkel cells were localized above the basal layer. Their shape was round to oval, dendrites being infrequent and short. At fetal wk 9, Merkel cells were considerably more numerous. These results persuasively argue for a much earlier fetal development of Merkel cells within the epidermis than previously thought. A hypothesis concerning the differentiation of Merkel cells from embryonic basal keratinocytes is discussed.     

Keratin expression in Merkel cells of fetal rat skin.

The cytokeratin expression of Merkel cells in fetal rat skin was studied by light- and electron microscopy. Employing a pre-embedding staining method, 2 monoclonal anti-keratin antibodies (RCK-102, MA-902) were shown to stain Merkel cells specifically. Neighbouring keratinocytes were unstained. The staining reaction seems to be based on the expression of 52.5 kD cytokeratin.     

Merkel cells and Merkel cell carcinoma express the BCL-2 proto-oncogene

Abstract The bcl-2 proto-oncogene, which is involved in the regulation of apoptosis, is expressed in a wide varity of fetal and adult tissues. We and others have demonstrated recently that in the human skin melanocytes, nevus cells and melanoma cells express bcl-2 constitutively. In the present study, we have analysed the expression of bcl-2 in Merkel cells and in Merkel cell carcinomas. In 2 colour immunofluorescencc staining, normal human Merkel cells as identified by the expression of cytokeratins 8, 18 and 20, were also anti-bcl-2 positive. Staining of paraffin sections of Merkel cell carcinomas with an anti-bcl-2 monoclonal antibody revealed strong bcl-2 protein immunoreactivtiy in all 5 tumors tested. Serial sections of Merkel cell carcinomas stained with the monoclonal antibodies CK 20, CAM 5.2, anti-neuron-specific enolase and anti-bcl-2 showed that the anti-bcl-2 reactive cells were indeed tumor cells. Our data demonstrate for the first time, that normal human Merkel cells and Merkel cell carcinomas express bcl-2 constitutively. Considering the biological function of the bcl-2 proto-oncogene, i.e., its anti-apoptotic effect, it is conceivable that in the near future, modulations of the expression of this protein may offer a new strategy in the therapy of bcl-2 expressing tumors such as Merkel cell carcinoma.     

Merkel cells and Merkel cell carcinoma express the BCL-2 proto-oncogene

Abstract The bcl-2 proto-oncogene, which is involved in the regulation of apoptosis, is expressed in a wide varity of fetal and adult tissues. We and others have demonstrated recently that in the human skin melanocytes, nevus cells and melanoma cells express bcl-2 constitutively. In the present study, we have analysed the expression of bcl-2 in Merkel cells and in Merkel cell carcinomas. In 2 colour immunofluorescence staining, normal human Merkel cells as identified by the expression of cytokeratins 8, 18 and 20, were also anti-bcl-2 positive. Staining of paraffin sections of Merkel cell carcinomas with an anti-bcl-2 monoclonal antibody revealed strong bcl-2 protein immunoreactivity in all 5 tumors tested. Serial sections of Merkel cell carcinomas stained with the monoclonal antibodies CK 20. CAM 5.2, anti-neuron-specific enolase and anti-bcl-2 showed that the anti-bcl-2 reactive cells were indeed tumor cells. Our data demonstrate for the first lime, that normal human Merkel cells and Merkel cell carcinomas express hcl-2 constitutively. Considering the biological function of the bcl-2 proto-oncogene, i.e., its anti-apoptotic effect, it is conceivable that in the near future, modulations of the expression of this protein may offer a new strategy in the therapy of bcl-2 expressing tumors such as Merkel cell carcinoma     

皮肤Merkel细胞癌3例

目的：报告3例皮肤Merkel细胞癌，进一步了解该肿瘤的临床表现、组织病理学特点和电镜观察的特征。方法：采用光镜、免疫组化和电镜技术，对3例患者的皮损进行组织病理学观察及诊断。结果：光镜检查示肿瘤位于真皮内，呈团片状、巢状、岛状或散在分布，细胞大小较一致。免疫组化示NSE、CgA、Sy、EMA、α—ACT在肿瘤细胞呈阳性表达。电镜观察示肿瘤细胞胞质内可见神经分泌颗粒。结论：皮肤Merkel细胞诊断依赖病理学检查，早期诊断、早期治疗是提高生存率的关键。     

Merkel cell polyomavirus DNA detection in lesional and nonlesional skin from patients with Merkel cell carcinoma or other skin diseases

Summary Background A novel polyomavirus, the Merkel cell polyomavirus (MCPyV), has recently been identified in Merkel cell carcinoma (MCC). Objectives To investigate the specificity of this association through the detection, quantification and analysis of MCPyV DNA in lesional and nonlesional skin biopsies from patients with MCC or with other cutaneous diseases, as well as in normal skin from clinically healthy individuals. Methods DNA was extracted from lesional and nonlesional skin samples of patients with MCC or with other cutaneous diseases and from normal‐appearing skin of clinically healthy subjects. MCPyV DNA was detected by polymerase chain reaction (PCR) and quantified by real‐time PCR. Additionally, the T antigen coding region was sequenced in eight samples from seven patients. Results MCPyV DNA was detected in 14 of 18 (78%) patients with MCC, five of 18 (28%) patients with other skin diseases (P = 0·007) and one of six (17%) clinically healthy subjects. In patients with MCC, viral DNA was detected in nine of 11 (82%) tumours and in 10 of 14 (71%) nontumoral skin samples (P = 0·66). MCPyV DNA levels were higher in MCC tumours than in nontumoral skin from patients with MCC, and than in lesional or nonlesional skin from patients with other cutaneous disorders. Signature mutations in the T antigen gene were not identified in the two MCC tumour specimens analysed. Conclusions High prevalence and higher levels of MCPyV DNA in MCC supports a role for MCPyV in tumorigenesis. However, the high prevalence of MCPyV in the nontumoral skin and in subjects without MCC suggests that MCPyV is a ubiquitous virus.     

Neurochemical markers in human cutaneous Merkel cells

Abstract Merkel cells (MCs) are specialized sensory cells widely distributed in the epithclia of vertebrates. A variable immunohistochemical pattern of neuronal and neurotransmitter markers has been demonstrated in MCs of several species including man. In the present study, we investigated the expression of neurochemical markers in a selected population of human cutaneous MCs by immunofluorescence. The structural neural proteins protein gene product 9.5 and neuron-specific enolase were found to be the most reliable markers for MC identification. Moreover, neurofil-ament immunoreactivity was shown in a small subset of epidermal MCs. Among the neurotransmitter markers, evidence for expression of calcitonin gene-related peptide, vasoactive intestinal polypeptide, peptide histidine isoleucine amide, neuropeptide Y, neurokinin A, galanin, substance P, som-atostatin and phenylethanolamine N-methyltransferase was found. These immunoreactivities were highly variable as far as number of positive cells and staining intensity were concerned. The results indicate that a complex and heterogenous immunophenotype can be expressed even within a homogeneous population of human MCs.     

Merkel cells in the vellus hair follicles of human facial skin: a study using confocal laser microscopy

Many cases of Merkel cell carcinoma have recently been reported, and most of them have been localized on the facial skin. In this study, we investigated Merkel cells in the vellus hair follicles of facial region to characterize these cells in human subjects. Skin specimens doubly stained with cytokeratin (CK) 20 and either protein gene product (PGP) 9.5 or vasoreactive intestinal polypeptide (VIP) were examined by confocal laser microscopy. Many of the Merkel cells in the vellus hair follicles of the facial skin were localized in the bulge area. Some of these cells were attached to nerve terminals, although most of them were not associated with them. Our results suggest that there are two types of Merkel cells in the bulge area of the vellus hair follicles of facial skin: cells wholly unassociated with the nerve terminals and cells associated with thin nerve fibers. We postulate that the former cells may be undifferentiated (immature) and the latter differentiated (mature). If this is so, there is a chance that Merkel cell carcinoma originates from the undifferentiated Merkel cells in the bulge of the vellus hair with the formation of tumor masses in the dermis and no involvement of the epidermis. The Merkel cells connected with nerve fibers may secrete endocrine substances via a regulation of autonomic nerves.     

Primary cutaneous ganglioneuroma: anatomico‐clinical study of 4 cases with focus on Merkel cells

Cutaneous ganglioneuromas (CGNs) are exceptional. We aim to describe the anatomico‐clinical profile of primary CGN and report 4 cases. Patients were 2 men and 2 women aged 53 to 76 years, who had flesh‐colored nodules on the back, associated with adjacent keratotic changes, that is, epidermal nevus (1 case) or seborrheic keratosis (3 cases). Histopathology showed ganglion cells within a proliferation of Schwann cells. The epidermis was acanthotic, associated with sebaceous induction in 2 cases, with follicular hyperplasia as in fibroepithelial tumors (1 case) or with tricholemmoma (2 cases). Cytokeratin‐20 immunostaining showed Merkel cells in the epidermis. A higher density of Merkel cells was observed in BerEP4+ follicular structures. Along with 16 published cases, our study indicates that a nodule associated with seborrheic keratosis on the back may represent a CGN, a complex mesenchymal and epidermal/follicular lesion of neuroectodermal lineage, associating neuronal proliferation and Merkel cell hyperplasia with follicular induction.     

Merkel cell carcinoma in situ: New insights into the cells of origin

We report a case of a 71-year-old male who presented with a small skin-coloured plaque on his cheek. Histopathology demonstrated an intraepidermal carcinoma with follicular involvement. No evidence of dermal invasion was seen. Immunohistochemical studies showed areas of positive staining for CK20, EMA and synaptophysin. Histopathology findings were found to be most consistent with a diagnosis of intraepidermal carcinoma with features of Merkel cell carcinoma in situ, in combination with a squamous cell carcinoma in situ, with follicular involvement. Recent advances and findings suggest Merkel cell polyomavirus MCPyV-positive Merkel cell carcinoma and MCPyV-negative Merkel cell carcinoma have different cells of origin from different germ layers.     

Usefulness of ulceration and hyperkeratosis as clinical predictors of Merkel cell polyomavirus‐negative and combined Merkel cell carcinoma: A retrospective study

Merkel cell carcinoma is a rare neuroendocrine carcinoma of the skin that is associated with Merkel cell polyomavirus (MCPyV). The clinical appearance and demographic characteristics of this tumor have been described using the mnemonic AEIOU: asymptomatic, expanding rapidly, immune suppression, older than 50 years, and ultraviolet‐exposed fair skin. In addition, MCC can be categorized based on morphology as pure MCC or combined MCC that exhibits neuroendocrine and other phenotypic elements. There is limited information regarding the clinical characteristics and prognosis of combined MCC. This retrospective study aimed to identify factors, such as ulceration or hyperkeratosis, that could predict MCPyV status and morphological variants. Twenty patients with MCC were divided into groups based on MCPyV status and morphology: MCPyV‐positive or MCPyV‐negative MCC and pure or combined MCC. The patients’ MCPyV status was immunohistochemically determined using the CM2B4 antibody to the MCPyV large T‐antigen. The patients’ clinicopathological characteristics were evaluated to identify predictors of MCPyV‐negative MCC and combined MCC. The presence of ulceration/hyperkeratosis predicted the presence of MCPyV‐negative MCC (80% of cases) and combined MCC (50% of cases). None of the 10 patients with MCPyV‐positive MCC had ulceration/hyperkeratosis. The clinical presence of ulceration/hyperkeratosis may help guide the diagnosis of MCPyV‐negative MCC and combined MCC.     

Merkel cell carcinoma: a clinicopathological study of 11 cases

OBJECTIVE: To report our 12-year experience with Merkel cell carcinomas (MCCs) from a clinical and pathological point of view. SUBJECTS AND SETTING: Eleven MCCs were diagnosed at our institution between 1991 and 2002. METHODS: A retrospective clinical, histopathological and immunohistochemical study was performed. Age, gender, location, size, stage, treatment and follow-up data were collected. Histopathological pattern and immunohistochemical study with CAM 5.2, cytokeratin 20 (CK20), CK7, Ber EP4, neurofilaments, synaptophysin, chromogranin, S100 protein, p53 protein, CD117, leucocyte common antigen (LCA) and Ki-67 were accomplished. RESULTS: Six females and five males with a mean age of 82 years were identified. Tumours were located on the face (n = 6), extremities (n = 3) and trunk (n = 1). At diagnosis, one patient was in stage Ia, six in stage Ib, three in stage II and one in stage III. All but one patient experienced wide surgical excision of the tumour. Additional treatment consisted of lymph node dissection in two patients, radiotherapy in four patients and systemic chemotherapy in one patient. Local recurrence developed in five patients. Three patients died because of MCC after 14 months of follow-up. Intermediate-size round cell proliferation was found in all cases. Additional small-size cell pattern and trabecular pattern were observed in seven and six cases, respectively. Eccrine and squamous cell differentiation were found in three cases. A dot-like paranuclear pattern was observed in all cases with CAM 5.2 and neurofilaments, and in 89% of cases with CK20. Seventy-five per cent of cases reacted with Ber EP4, chromogranin and synaptophysin, 70% with p53, 22% with S100 protein, 55% with CD117 and none with LCA. Ki-67 was found in 75% of tumoral cells on average. Fifty per cent of MCCs reacted with CK7 and showed eccrine differentiation areas. CONCLUSIONS: MCC is an aggressive neuroendocrine tumour of the elderly. Wide surgical excision is the recommended treatment. Lymph node dissection, adjuvant radiotherapy and chemotherapy decrease regional recurrences but have not been demonstrated to increase survival. Immunohistochemically, MCC is an epithelial tumour with neuroendocrine features.