Melanin-containing Tumor Research Articles

Depigmentation of Melanin-containing Tissues Using Hypochlorous Acid to Enhance Hematoxylin-eosin and Immunohistochemical Staining.

Pathologists diagnose diseases by observing the histologic and cellular morphology microscopically. However, the high pigmentation in melanin-containing tumors can hide the tumor cell structures, making diagnosing challenging. Previously, hydrogen peroxide and potassium permanganate were utilized for melanin bleaching with several limitations. For instance, hydrogen peroxide has a weak bleaching ability, and the process is time-consuming (12 h). Meanwhile, potassium permanganate affects the antigenicity of antigens and is unsuitable for immunohistochemical (IHC) staining. In this study, the hypochlorous acid (HClO) solution was applied to hematoxylin-eosin and IHC staining of melanin tissue sections. The study discovered that 1% HClO could completely bleach melanin particles in tumor tissues in a short period (19.95 ± 2.53 min) without compromising the hematoxylin-eosin staining. In addition, 2% HClO was utilized for bleaching at room temperature for 61.17 ± 4.32 minutes after the tissue was incubated with 3,3'-diaminobenzidine in IHC staining. This treatment effectively removed melanin without negatively impacting 3,3'-diaminobenzidine signal expression, thus ensuring that the sections met the necessary diagnostic requirements. Therefore, this method could facilitate pathologists in disease diagnosis of melanin-containing tissues.

Melanotic Neuroectodermal Tumor of Infancy: A Clinicopathological and BRAF V600E Mutation Study of 11 Cases.

PurposeTo investigate the clinicopathological features and BRAF V600E mutation of melanotic neuroectodermal tumor of infancy (MNTI).Materials and MethodsEleven cases of MNTI diagnosed at the Department of Oral Pathology were collected. Clinicopathological characteristics were obtained from the medical records. Immunostaining was performed by immunohistochemistry (IHC). Amplification-Refractory Mutation System-qPCR (ARMS-qPCR) and Sanger Sequencing were used to detect BRAF V600E mutation.ResultsOf the 11 cases, 3 cases were female and 8 cases were male. The mean age of the first symptoms was 3.2 months (range: 1 to 6 months). Ten cases (90.9%) located in maxilla but only one (9.1%) in mandible. Most of the cases demonstrated well-defined mass with lytic bone destruction and tooth germ affecting radiologically. Histologically, MNTI was consisted of large polygonal melanin-producing epithelioid cells and small round neuroblast-like cells which arranged in irregular alveolar, tubuloglandular and fissured architecture. The epithelioid cells expressed Vim, Pan-CK, NSE and HMB45, while the smalls cells expressed Syn, NSE and scattered Vim. Most cases showed low Ki-67 index (range: <1% to 50%). None of the MNTI cases showed BRAF V600E mutation. Most cases were treated with enucleation (45.4%) or curettage (36.4%). Among the 11 cases, 6 cases had follow-up information, and 2 cases had recurrence lesions after surgery.ConclusionMNTI, an extremely rare tumor, mainly affects male infants with strong preference for maxilla. Distinct histopathological features and immunohistochemical profile are helpful to distinguish from other melanin-containing tumors and small round cell tumors. No BRAF V600E mutation in MNTI is detected in the present study and needs further investigations. The factors that contribute to the local recurrence of MNTI are controversial, but the close follow-up for the patients is recommended.

Application of methyl green as a counter stain for immunohistochemical staining of melanin-containing tumor

目的： 探讨甲基绿对比染色法在含黑色素肿瘤免疫组织化学染色中的应用。 方法： 含黑色素肿瘤30例，其中蓝痣10例，黑色素瘤20例，构建组织芯片。采用高锰酸钾氧化草酸漂白法脱黑色素后行免疫组织化学检测和免疫组织化学染色后用1%甲基绿对比染色进行比较，应用EliVision免疫组织化学法检测HMB45、Melan A、S-100蛋白、SOX10的表达，分析两种方法对含黑色素肿瘤不同免疫组织化学指标判读的影响。 结果： 经过高锰酸钾氧化草酸漂白法脱黑色素处理后，部分病例出现破片、脱片现象；HMB45的染色效果与1%甲基绿对比染色后HMB45染色效果基本一致；SOX10出现假阴性；Melan A大部分病例出现表达减弱或阴性表达；S-100蛋白表达良好。经1%甲基绿对比染色后黑色素被染成绿色，与免疫组织化学染色结果的棕黄色形成对比，免疫组织化学染色结果定位准确，易于判读。2例含重度黑色素的病例组织由于色素覆盖厚重，影响不同指标的显色结果判读。 结论： 含黑色素的肿瘤组织免疫组织化学染色后采用1%甲基绿对比染色法，在解决轻-中度含黑色素肿瘤免疫组织化学判读上相对比传统的高锰酸钾氧化草酸漂白法脱黑色素方法好。.

Photothermal effect by 808-nm laser irradiation of melanin: a proof-of-concept study of photothermal therapy using B16-F10 melanotic melanoma growing in BALB/c mice.

The photothermal effect is undergoing great interest due to advances in new photosensitizing materials and better-suited light sources, but studies are frequently hampered by the need to employ exogenous photothermal agents and expensive irradiation devices. Here we present a simple strategy based on direct NIR irradiation of the melanin pigment with a commercial 808-nm laser pointer. Proof-of-concept studies showed efficient photothermal effects on melanin in vitro and in vivo. After NIR irradiation, BALB/c mice bearing B16-F10 melanotic melanoma tumors revealed severe histopathological damage and massive necrosis in melanin-containing tumor tissue, while surrounding healthy tissues showed no damage. Therefore, the feasibility of this approach may allow implementing direct procedures for photothermal therapy of pigmented tumors.

Melanotic Neuroectodermal Tumor of Infancy: A Case Report

Melanotic neuroectodermal tumor of infancy (MNTI) is a rare, benign, fast-growing, melanin-containing tumor originating from the neural crest. MNTI can present locally aggressive behavior including bone destruction. A 2-month-old female presented a bluish swelling, with well-defined limits, in the premaxilla. Computed tomography revealed a homogeneous and hypodense image associated with the upper left central primary incisor. The diagnostic hypothesis was dentigerous cyst. An incisional biopsy was performed, and microscopic examination revealed a benign neoplasm exhibiting proliferation of a dual population of cells arranged in solid nests or cords. The first cell type consisted of small rounded cells with scarce cytoplasm, and the second consisted of epithelioid cells containing brown cytoplasmic granules consistent with melanin. Based on these findings, the definitive diagnosis was MNTI. Surgical excision was performed, and there was no tumor recurrence during 1 year of follow-up.

Primary extramedullary spinal melanoma mimicking spinal meningioma: A case report and literature review.

Primary spinal melanoma is a rare lesion, which occurs throughout the cranial and spinal regions, however, is primarily observed in the middle or lower thoracic spine. The clinical features of primary spinal melanoma are complex and unspecific, resulting in a high misdiagnosis rate. In the present case report, a rare case of spinal melanoma exhibiting the dural tail sign and mimicking spinal meningioma is reported. The initial diagnosis, using magnetic resonance imaging (MRI), was unclear. Thus, melanin-containing tumors and spinal meningioma should have been considered in the differential diagnosis. The tumor was completely resected using a standard posterior midline approach, which was followed by chemotherapy. Subsequent to the surgery, the patient was discharged with improved motor capacity and a follow-up MRI scan showed no recurrence after six months. The present study demonstrates that it is critical for neurosurgeons to focus on increasing the accuracy of initial diagnoses in order to make informed decisions regarding the requirement for surgical resection. The present case report presents the clinical, radiological and pathological features of primary extramedullary spinal melanoma mimicking spinal meningioma to emphasize the importance of early identification and diagnosis.

Primary tumor versus metastasis: new experimental models for studies on cancer cell homing and metastasis in melanoma

Primary tumor versus metastasis: new experimental models for studies on cancer cell homing and metastasis in melanoma

Malignant melanoma rather than malignant cutaneous melanoma?

We report a case of an 84-year-old woman with a history of resected cutaneous melanoma who presented with asthenia, weight loss, and severe anemia. An esophagogastroduodenoscopy showed five black-pigmented submucosal tumors with large ulcerations in the antrum of the stomach and in the duodenum, suspected metastases from melanoma. Histopathological examination indicated the presence of melanin-containing tumor cells and confirmed an ulcerated pigmented melanoma. To examine the entire gastrointestinal tract, capsule endoscopy was performed and it showed a simultaneous massive involvement of the jejunum and ileum, with more than 40 lesions. This case highlights the importance of a complete endoscopic gastrointestinal examination in patients with melanoma metastatic to the bowel and suggests that capsule endoscopy is an easy, noninvasive, and effective diagnostic procedure to investigate small-bowel involvement.

Preclinical Evaluation of an 131I-Labeled Benzamide for Targeted Radiotherapy of Metastatic Melanoma

Radiolabeled benzamides are attractive candidates for targeted radiotherapy of metastatic melanoma as they bind melanin and exhibit high tumor uptake and retention. One such benzamide, N-(2-diethylamino-ethyl)-4-(4-fluoro-benzamido)-5-iodo-2-methoxy-benzamide (MIP-1145), was evaluated for its ability to distinguish melanin-expressing from amelanotic human melanoma cells, and to specifically localize to melanin-containing tumor xenografts. The binding of [(131)I]MIP-1145 to melanoma cells in vitro was melanin dependent, increased over time, and insensitive to mild acid treatment, indicating that it was retained within cells. Cold carrier MIP-1145 did not reduce the binding, consistent with the high capacity of melanin binding of benzamides. In human melanoma xenografts, [(131)I]MIP-1145 exhibited diffuse tissue distribution and washout from all tissues except melanin-expressing tumors. Tumor uptake of 8.82% injected dose per gram (ID/g) was seen at 4 hours postinjection and remained at 5.91% ID/g at 24 hours, with tumor/blood ratios of 25.2 and 197, respectively. Single photon emission computed tomography imaging was consistent with tissue distribution results. The administration of [(131)I]MIP-1145 at 25 MBq or 2.5 GBq/m(2) in single or multiple doses significantly reduced SK-MEL-3 tumor growth, with multiple doses resulting in tumor regression and a durable response for over 125 days. To estimate human dosimetry, gamma camera imaging and pharmacokinetic analysis was performed in cynomolgus monkeys. The melanin-specific binding of [(131)I]MIP-1145 combined with prolonged tumor retention, the ability to significantly inhibit tumor growth, and acceptable projected human dosimetry suggest that it may be effective as a radiotherapeutic pharmaceutical for treating patients with metastatic malignant melanoma.

Induction of Primary Cutaneous Melanomas in C3H Mice by Combined Treatment with Ultraviolet Radiation, Ethanol and Aloe Emodin ¶

The role of ultraviolet (UV) radiation in the induction of nonmelanoma skin cancer is widely accepted, although its precise contribution to the development of primary cutaneous melanoma skin cancer requires further definition. We found that painting aloe emodin, a trihydroxyanthraquinone from Aloe barbadensis, in ethyl alcohol vehicle on the skin of mice in conjunction with exposure to UVB (280–320 nm) radiation results in the development of melanin-containing skin tumors. C3H/HeN mice were treated thrice weekly with aloe emodin in a 25% ethanol in water vehicle and exposed to 15 kJ/m2 UV radiation. Neither ethanol vehicle nor aloe emodin alone induced skin tumors in the absence of UV radiation. In two separate experiments, 20–30% of the mice treated with a combination of UV radiation and ethanol vehicle and 50–67% of the UV-irradiated animals given aloe emodin in ethanol vehicle developed primary cutaneous melanin-containing tumors. The diagnosis of melanoma was established using Fontana silver stain for melanin; these tumors were negative for vimentin and keratin. Melanin-containing melanosomes were observed by transmission electron microscopy in tumors diagnosed as melanomas. Although the mechanism of carcinogenesis in these mice is currently unknown, our findings have led to the development of the first facile murine model for the induction of primary melanoma. This model has the potential to clarify the role of UV radiation in the etiology of malignant melanoma.

An improved method for RNA isolation and removal of melanin contamination from melanoma tissue: Implications for tumor antigen detection and amplification

Several kits are available to isolate RNA 1 from tissues. However, melanoma tissue is often rich in melanin that co-purifies with DNA/RNA and inhibits subsequent PCR reactions, hampering tumor antigen detection and amplification. This problem has not yet been addressed systematically. Here we generated a photometric protocol to determine both the melanin and RNA concentration by correcting the latter for the absorption coefficient of melanin at 260 nm. Subsequently, different combinations of silica-based RNA-binding, size-exclusion, and ion-exchange columns were used in 8 protocols for isolation of RNA from melanoma tissue to compare efficacy of melanin removal and yield of RNA. Furthermore, the capability of the different RNA preparations to function as template in RT-PCRs with products of different length, i.e. GAP-DH, tyrosinase, and gp100, was tested. We found that the combination of silica-based RNA-binding and size-exclusion columns was not sufficient to remove melanin from highly contaminated tumor samples, and subsequent RT-PCR failed to give larger products. However, protocols including ion-exchange columns resulted in efficient removal of melanin, while retaining reasonable RNA yields in samples from highly pigmented melanomas. Efficient RT-PCR of larger products turned out to be inversely correlated to the melanin contamination. This RNA-purification method will help scientists to isolate polynucleotides from melanin-containing tumor samples, which subsequently can be used in antigen detection assays and vaccination strategies using amplified total tumor RNA.

Induction of Primary Cutaneous Melanomas in C3H Mice by Combined Treatment with Ultraviolet Radiation, Ethanol and Aloe Emodin¶

The role of ultraviolet (UV) radiation in the induction of nonmelanoma skin cancer is widely accepted, although its precise contribution to the development of primary cutaneous melanoma skin cancer requires further definition. We found that painting aloe emodin, a trihydroxyanthraquinone from Aloe barbadensis, in ethyl alcohol vehicle on the skin of mice in conjunction with exposure to UVB (280-320 nm) radiation results in the development of melanin-containing skin tumors. C3H/HeN mice were treated thrice weekly with aloe emodin in a 25% ethanol in water vehicle and exposed to 15 kJ/m2 UV radiation. Neither ethanol vehicle nor aloe emodin alone induced skin tumors in the absence of UV radiation. In two separate experiments, 20-30% of the mice treated with a combination of UV radiation and ethanol vehicle and 50-67% of the UV-irradiated animals given aloe emodin in ethanol vehicle developed primary cutaneous melanin-containing tumors. The diagnosis of melanoma was established using Fontana silver stain for melanin; these tumors were negative for vimentin and keratin. Melanin-containing melanosomes were observed by transmission electron microscopy in tumors diagnosed as melanomas. Although the mechanism of carcinogenesis in these mice is currently unknown, our findings have led to the development of the first facile murine model for the induction of primary melanoma. This model has the potential to clarify the role of UV radiation in the etiology of malignant melanoma.

Spotaneous Melanocytosis in Transgenic Mice

Clone B is a 2-kb fragment of cloned genomic DNA involved in adipocyte differentiation in vitro. Insertion of this DNA fragment into the genome of a variety of cell lines results in committing the recipient cells to undergo adipocyte differentiation. Construction of transgenic mice with Clone B DNA resulted in an unexpected phenotype--spontaneous melanocytosis. The present study describes the distribution and morphology of melanin-containing lesions in these transgenic mice. Spontaneous melanin-containing dermal lesions appeared on the ears, snout, and perianal regions of transgenic mice by the age of 3-4 months. Multifocal dermal masses rapidly developed into raised lesions, which appeared to spread to adjacent skin. Ultrastructural examination of lymph nodes, spleen, and dermal lesions of these mice revealed membrane-bound melanin with effacement f the organelle structure of severely affected cells. Protein gel electrophoresis revealed elevated activity of tyrosinase in the pinnae, skin, perianal mass, and lymph nodes. This line of transgenic mice may provide a useful model for investigation of the etiology and progression of benign and malignant melanin-containing tumors.

Immunohistochemical detection of melanoma-associated antigens on formalin-fixed, paraffin-embedded canine tumors.

Seven monoclonal antibodies (Moabs), recognizing melanoma-associated antigens in human tissues, were evaluated for their ability to immunohistochemically stain formalin-fixed, paraffin-embedded canine melanomas. Only 2 Moabs, designated human melanosome-specific antigen (HMSA)-1 and HMSA-5, stained routinely processed canine melanomas, staining 21/35 (60%) and 24/35 (69%), respectively. Twenty-nine of 35 (83%) melanomas tested were stained if results of the 2 Moabs were combined. Monoclonal antibody HMSA-1 also stained neoplastic cells of 10/35 (29%) tumors of nonmelanocytic origin and some neurons and salivary gland epithelial cells in normal canine tissues. However, Moab HMSA-1 staining in the nonmelanocytic tumors, consisting of small, discrete periodic acid-Schiff-positive cytoplasmic droplets, was readily distinguishable from the diffusely granular, cytoplasmic staining of melanocytic tumors. In addition to melanomas, Moab HMSA-5 stained melanocytes and some melanin-containing tumor cells of a pigmented basal cell tumor and melanocytes in normal canine skin. Monoclonal antibodies HMSA-1 and HMSA-5 immunohistochemically identified the majority of canine melanomas, had limited and distinguishable staining in normal tissues and nonmelanocytic tumors, and therefore may be a useful adjunct for the diagnosis of canine melanoma in formalin-fixed, paraffin-embedded tissues.

Peripheral carcinoid tumor of the lung with focal melanin production

A case of carcinoid tumor of the lung with focal melanin production was encountered in a 56 year old Japanese woman. The tumor was found 16 years previously by mass survey chest X-ray and had enlarged two-fold in the intervening period. The tumor consisted of a variety of tumor cells showing a spindle, polygonal and pleomorphic appearance with abundant vasculature in the stroma. All tumor cells showed argyrophilia, together with a few showing argentaffinity. Melanin-containing tumor cells were also present in parts. Ultrastructurally, most tumor cells possessed various numbers of neurosecretory granules and a few of them contained granular type melanosomes. Tumor cells were connected with desmosomes and a few of them contained tonofilament-like microfilaments. Only a few contained both neurosecretory granules and melanin. By immunohistochemistry, serotonin, met-enkephalin and beta-endorphin positive cells were observed scattered throughout the tumor. A few tumor cells positive for tyrosine hydroxylase were also detected. Additionally, most tumor cells were positive for keratin. On the basis of these findings, the tumor of the current case is a pulmonary carcinoid tumor with focal melanin production.

Melanosis Renum Bovis

The peculiar type of pigmentation of the kidneys in cattle commonly known as “black kidneys“ has been studied. 1) The pigmentation is localized to the neck, the proximal convoluted tubule and its straight terminal portion. Deposits may occasionally be observed in the parietal layer of the capsule of Bowman. Outside the kidneys very small amounts of pigment may be found in the lymph nodes, particularly those of the kidneys, in the liver, and in the spleen. 2) Focal, productive inflammatory changes in the renal interstitial tissue were found by statistical analysis to occur with greater frequency in black pigmented kidneys than in the controls. 3) Histochemical study and analysis of the isolated pigment suggest that the pigment is a true melanin. The term melanosis renum bovis is therefore suggested for the condition. 4) Chromatoraphy of acid and alkali hydrolysates prepared under pressure (l10°C) demonstrated the following amino acids in the purified product: Lysine, asparaginic acid, glutamic acid, glysine, alanine, valine, isoleucine, phenylalanine, cystine-cysteine, "fast arginine", proline, methionine sulfone, treonine, serine and possibly histidine and tyrosine. The pigment contained 9,5 per cent nitrogen, 44,9 per cent carbon and 6,4 per cent hydrogen. The ash content was 4,2 per cent, and the content of iron 0,9 per cent. 5) The pigment is excreted in the urine but in very small amounts. Urine containing the pigment does not give the same reactions as urine from individuals suffering from melanin-containing tumors, Addison’s disease, ochronosis or alkaptonuria. 6) Certain circumstances indicating the renal parenchyma as the site of formation of the pigment are discussed. 7) The far more frequent occurrence of the anomaly in a certain breed (Redt Dansk Malkekvreg) than in others suggests a hereditary factor. Some kind of local allergic re action may possibly also be involved.