An Optimized Melanin Depigmentation Method for Histopathological and Immunohistochemical Staining of Formalin-Fixed Paraffin-Embedded Tissues.

Introduction. The difficulty in diagnosis of severe melanocytic lesions is a problem to be overcome in pathological practice. Melanin bleaching is an effective approach to ameliorate melanin disturbances in severely pigmented lesions. Although various methods for improving melanin pigmentation in immunohistochemical staining have been reported, these depigmentation methods still need to be optimized and standardized. In this study, the coloring efficiency of 3,3'-diaminobenzidine (DAB) and alkaline phosphatase (AP) after melanin depigmentation was compared under the automatic immunohistochemical staining platform. Methods. The applicability of the optimized depigmentation method was validated in 10 formalin-fixed paraffin-embedded (FFPE) blocks of ocular melanoma tissues. Specimens were demelaninized with 10% hydrogen peroxide at 60°C for immunohistochemical staining (Melan-A and SOX10), and tissue chromogenic staining was performed with DAB and AP detection systems, respectively. Results. The optimized depigmentation method including immunohistochemistry (IHC) could be completed in 3 h, effectively preserving cell morphology and immunoreactivity. Among these, the color-rendering effect and contrast of AP are better than DAB. Conclusion. This optimized method can effectively remove melanin and improve the accuracy of IHC staining interpretation. AP staining has better visibility and readability without the interference of residual melanin. The comparison results showed that after melanin depigmentation, the immunohistochemical staining agent was replaced with red AP, which avoided the misjudgment caused by brown DAB when melanin depigmentation was incomplete. This improved method can be applied to future histopathological and immunohistochemical staining of melanin-deposited tissues.

Concurrence of Marjolin's Ulcer in the Lower Limb in a Patient with Idiopathic Multicentric Castleman Disease: A Case Report.

Idiopathic multicentric Castleman disease (iMCD) is characterized by the benign proliferation of lymphoid cells in multiple regions. However, the co-occurrence of epithelial malignancy and idiopathic multicentric Castleman disease (iMCD) is rarely reported. Herein, we present a case of iMCD mimicking lymph nodal metastasis of Marjolin's ulcer in the lower extremity. A 53-year-old male presented with an unhealed chronic ulcer on the left lower leg and foot accompanied by an enlarged mass in the left inguinal region. Intralesional biopsy was performed, and pathological examination showed squamous cell carcinoma (SCC). Imaged studies revealed left calcaneus bone invasion, and lymph nodal metastasis was suspected by the cancer TNM staging of T4N2M0 pre-operatively. The patient received below-knee amputation and lymph node dissection; intraoperative histological examination showed no lymphatic nodal malignancy and diagnosed the patient as having iMCD with lymphadenopathy. The patient recovered uneventfully and was referred to a hematologist for further treatment.

IgG4-related disease mimicking renal pelvis tumor with peritoneal carcinomatosis.

IgG4-RD may rarely present as a retroperitoneal fibrosis, mimicking carcinomatosis. Clinicians should consider this disease when encountering an idiopathic peritoneal and retroperitoneal fibrosis with renal involvement and hydronephrosis.

Successful management of type IV hypersensitivity reactions to human insulin analogue with injecting mixtures of biphasic insulin aspart and dexamethasone.

Although hypersensitivity reaction to insulin was supposed to be less-frequent with current insulin analogue, case reports with different types of allergic reactions to insulin analogue were still reported. The most common form is type I hypersensitivity reaction with IgE-mediated. Besides, type III (IgG and IgM-mediated) and type IV (T-cell mediated delayed reaction) hypersensitivity reactions were also reported. Here we presented a long-standing type 2 diabetes with insulin requirements with hypersensitivity reactions to insulin actrapid, insulin aspart, insulin glargine, insulin detemir, and biphasic insulin aspart 30. Insulin desensitization was performed as initial management but failed as skin biopsy with immunohistochemical staining proved type IV hypersensitivity reaction. We continued with the next treatment approach using subcutaneous injection with the mixture of biphasic insulin aspart 30 and dexamethasone to alleviate allergy, and the result was successful with steroid-free biphasic insulin aspart 30 injection eight months later. Besides, the treatment effect had lasted after ten years even with switched type of insulin analogue from biphasic insulin aspart 30 to insulin glargine and insulin aspart. The case report demonstrated a good example of how clinicians deal with the rare but important questions of hypersensitivity reactions to insulin analogue.

Melanin Bleaching With Warm Hydrogen Peroxide and Integrated Immunohistochemical Analysis: An Automated Platform.

OBJECTIVE: Diagnosing melanocytic lesions is among the most challenging problems in the practice of pathology. The difficulty of physically masking melanin pigment and the similarity of its color to commonly used chromogens often complicate examination of the cytomorphology and immunohistochemical staining results for tumor cells. Melanin bleach can be very helpful for histopathological diagnosis of heavily pigmented melanocytic lesions. Although various depigmentation methods have been reported, no standardized methods have been developed. This study developed a fully automated platform that incorporates hydrogen peroxide-based melanin depigmentation in an automated immunohistochemical analysis. METHODS AND MATERIALS: The utility of the method was tested in 1 cell block of malignant melanoma cells in pleural effusion, 10 ocular melanoma tissue samples, and 10 cutaneous melanoma tissue samples. Our results demonstrated that the proposed method, which can be performed in only 3 hours, effectively preserves cell cytomorphology and immunoreactivity. RESULTS: The method is particularly effective for removing melanin pigment to facilitate histopathological examination of cytomorphology and for obtaining an unmasked tissue section for immunohistochemical analysis.

Fatal invasive aspergillosis: a rare co-infection with an unexpected image presentation in a patient with dengue shock syndrome.

INTRODUCTION: Pulmonary infiltration and pleural effusion caused by permeability syndrome are the hallmark of pulmonary manifestation of dengue cases. OBJECTIVE AND METHODS: We report a 95-year-old chronic obstructive pulmonary disease case having dengue shock syndrome. Chest X-ray examination revealed diffuse lung infiltration. However, bilateral pneumotoceles were unexpectedly found in computed tomography (CT) images. Dengue virus type 2 infection was confirmed by virus culture, serology and reverse transcriptase-polymerase chain reaction. Profound shock with bilateral lung infiltration developed rapidly in 2 days with supportive care and empirical ampicillin/ sulbactam. Bronchoscopy revealed a whitish plaque over bilateral upper bronchi. Biopsy via bronchoscopy revealed moulds with vascular invasion. Culture of bronchial alveolar lavage yielded Aspergillus flavus. The patient died despite amphotericin B treatment, which was started since finding the whitish plaque with bronchoscopy examination. RESULTS AND CONCLUSION: Besides to considering capillary leakage syndrome, our case report and literature review alert clinicians that CT and bronchoscopy may help to identify the true pathogen though all cases with concurrent dengue and Aspergillus infections had fatal outcomes.

A cohort study on 10-year survival of sporadic medullary thyroid carcinoma with somatic RET mutation.

Somatic rearranged during transfection (RET) mutations are reported in 40-50% of sporadic medullary thyroid carcinoma (sMTC) patients with prognostic significance. As there is a lack of somatic RET mutations reported previously for the Taiwanese population, we tried to assess the presence of somatic RET mutations and evaluate the potential outcome predictors for our sMTC patients. We collected data from seven sMTC patients from the years 1997 to 2005 and analyzed their clinic-pathological features up to 2015. All patients were still alive to follow up for 11∼18 years. Tumor DNAs were extracted to assess exons 10-11 and 13-16, and the intron-exon boundaries of the RET gene. Six cases (86%) were screened positive of somatic RET gene mutations in hotspot regions, one at M918T, one at C620R, and three at C634S, with another two rare mutations at L629Q and V642I. Comparing the current tumor, node, metastases staging system, the 10-year survival outcomes for our sMTC patients was not predicted by serum calcitonin and/or carcinoembryonic antigen, surgical extent, and presence of the somatic RET gene mutations. The small cohort demonstrated a relatively good outcome of sMTC patients to survive >10 years. In addition, intensive treatment with total thyroidectomy with extensive neck lymph node dissection seemed to be the critical determinant of better survival outcome for sMTC patients.

Liraglutide prevents and reverses monocrotaline-induced pulmonary arterial hypertension by suppressing ET-1 and enhancing eNOS/sGC/PKG pathways.

Liraglutide, a glucagon-like peptide-1 receptor (GLP-1R) agonist, is widely used to treat diabetes. However, its effect on pulmonary arterial hypertension (PAH) is unknown. In this study, we investigated its effects on rats with monocrotaline (MCT)-induced PAH and mechanisms on rat pulmonary artery smooth muscle cells (PASMCs). Liraglutide was investigated for both prevention and treatment of MCT-induced PAH. The hemodynamic and body weight changes, right heart hypertrophy, lung morphology, immune-reactivity of endothelial nitric oxide synthase (eNOS), endothelin-1 and cyclic guanosine monophosphate (cGMP) levels, protein expressions of eNOS, soluble guanylyl cyclase (sGCα), protein kinase G (PKG) and Rho kinase (ROCK) II pathway were measured in both in vivo and in vitro. Cell migration and cell cycle were also determined. Liraglutide both prevented and reversed MCT-induced PAH, right ventricle hypertrophy and pulmonary vascular wall remodeling. Protein expression of ROCK II was increased while eNOS, sGC and PKG were decreased. Pretreatment with liraglutide inhibited platelet-derived growth factor (PDGF)-BB stimulated PASMCs migration, which were associated with cell-cycle arrest at G0/G1 phase. Liraglutide may have both preventive and therapeutic effects on MCT-induced PAH, through the eNOS/sGC/PKG and Rho kinase pathways. Thus, liraglutide may have a therapeutic role in pulmonary vascular remodelling.