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.

Quantitative Proteomic Analysis of Differentially Expressed Protein Profiles Involved in Pancreatic Ductal Adenocarcinoma.

OBJECTIVES: The aim of this study was to identify differentially expressed proteins among various stages of pancreatic ductal adenocarcinoma (PDAC) by shotgun proteomics using nano-liquid chromatography coupled tandem mass spectrometry and stable isotope dimethyl labeling. METHODS: Differentially expressed proteins were identified and compared based on the mass spectral differences of their isotope-labeled peptide fragments generated from protease digestion. RESULTS: Our quantitative proteomic analysis of the differentially expressed proteins with stable isotope (deuterium/hydrogen ratio, ≥ 2) identified a total of 353 proteins, with at least 5 protein biomarker proteins that were significantly differentially expressed between cancer and normal mice by at least a 2-fold alteration. These 5 protein biomarker candidates include α-enolase, α-catenin, 14-3-3 ß, VDAC1, and calmodulin with high confidence levels. The expression levels were also found to be in agreement with those examined by Western blot and histochemical staining. CONCLUSIONS: The systematic decrease or increase of these identified marker proteins may potentially reflect the morphological aberrations and diseased stages of pancreas carcinoma throughout progressive developments leading to PDAC. The results would form a firm foundation for future work concerning validation and clinical translation of some identified biomarkers into targeted diagnosis and therapy for various stages of PDAC.

Primary small cell carcinoma of kidney after renal transplantation: a case report and literature review.

Extrapulmonary small cell carcinoma (EPSCC) is a rare neoplasm comprising 2.5% to 5% of small cell carcinomas (SCCs). Bladder SCC is the most common site of genitourinary tract. Primary renal SCC is extremely rare. We report a case of primary SCC of the kidney which is rarely reported in the urinary tract and presents an aggressive clinical picture. A 59-year-old female visited a urologic clinic with complaint of persistent left flank soreness 10 years after undergoing renal transplantation. Abdominal computed tomography showed a left renal pelvis tumor. After the patient received left nephroureterectomy with bladder cuff resection, her pathology results showed SCC. After surgery, she received adjuvant systemic chemotherapy, and her recovery has been uneventful as of 8 months. Primary renal SCC presents with an advanced tumor stage and a short median survival period, therefore early intervention and close follow-up are recommended.

Lingual leiomyomatous hamartoma with bifid tip and ankyloglossia in a patient without oral-facial-digital syndrome: a case report and literature review.

Here is a rare case of lingual leiomyomatous hamartoma (LLH) with bifid tongue tip and tongue-tie in a patient with non-oral-facial-digital syndrome (OFDS). A 29-year-old male consulted for a painless tumor over the midline of the tongue dorsum measuring 2 × 1.5 cm. The tumor was excised and the tongue-tie was corrected. Diagnosis of LLH was based on histo-pathologic and immuno-histochemical studies. The epidemiologic data and differential diagnosis of LLH, as well as related literature, are discussed. To date, only 14 cases of LLH have been reported in English literature. This may be the first reported case of LLH with bifid tip and ankyloglossia in a non-OFDS patient.

Melanin bleaching with dilute hydrogen peroxide: a simple and rapid method.

Melanins are naturally occurring pigments in both normal and pathologic tissues. Two common bleaching processes are potassium permanganate followed by oxalic acid treatment and dilute hydrogen peroxide (H2O2) process. The potassium permanganate/oxalic acid method is faster and more easily incorporated in conventional daily immunostaining protocols, whereas the dilute H2O2 method requires 24 hours. This study aimed to reduce melanin bleaching time by using a 10% H2O2 dilution. First, reaction time was reduced to 30 minutes by raising the temperature to 65°C. Second, containers with high thermal conductivity were used to improve bleaching effectiveness. Experimental comparisons of melanin treatments with H2O2 contained in an iron jar, a glass coplin jar, and a plastic steel jar obtained bleaching time of 20, 30, and 40 minutes, respectively. These modifications of the conventional bleaching method significantly improve the speed and efficiency of the procedure and are recommended when performing immunohistochemical studies.

Lymphoepithelioma-like carcinoma of ureter-a rare case report and review of the literature.

Lymphoepithelioma-like carcinoma (LELC) is best known to occur in the nasopharynx. When LELC occurs in the urinary tract, this extremely rare neoplasm most commonly affects the bladder but has also been reported in the renal pelvis, ureter, prostate [1], and urethra [2]. We present a case of LELC arising in the right proximal ureter of a 64-year-old male patient with hydronephrosis and nausea. Computed tomography demonstrated right ureter tumor. On biopsy, the patient was diagnosed with infiltrating urothelial carcinoma. An operation consisting of right nephroureterectomy and bladder cuff removal was carried out. The pathologic examination showed pure subtype of LELC, pT3N0. Unlike lymphoepithelioma in the nasopharynx, immunohistochemical analysis of this urinary LELC was negative for the Epstein-Barr virus. No disease progression was noted at 6 months' follow-up. Only eight previous cases of LELC involving the ureter have been reported, and a review of the available literature and a summary of ureter cases are presented here. This is the first report of a ureteral LELC case and third urothelial LELC cases [3] in Taiwan.

A mucormycosis case in a cirrhotic patient successfully treated with posaconazole and review of published literature.

Mucormycosis is an invasive fungal infection associated with a high mortality rate, especially in immunocompromised hosts. Mucormycosis rarely occurs in cirrhotic patients. Here, we report a case of mucormycosis with underlying liver cirrhosis and diabetes mellitus. The patient suffered from maxillary sinusitis and osteomyelitis, and the infection was successfully treated with antifungal agents, surgical debridement, and hyperbaric oxygen therapy. The antifungal treatments used were liposomal amphotericin B, itraconazole, and posaconazole. Although our patient had liver cirrhosis (Child-Pugh classification B), no hepatic decompensation was developed during the treatment course of posaconazole. This is the first report of the safe and effective use of posaconazole for the treatment of mucormycosis in a cirrhotic patient.

Comparison of Gd-Bz-TTDA, Gd-EOB-DTPA, and Gd-BOPTA for dynamic MR imaging of the liver in rat models.

To evaluate the competitive potential of a new lipophilic paramagnetic complex, Gd-Bz-TTDA [4-benzyl-3,6,10-tri (carboxymethyl)-3,6,10-triazado-decanedioic acid] compared with two other commercially available MR hepatobiliary contrast agents, gadobenate dimeglumine (Gd-BOPTA) and gadoxetic acid (Gd-EOB-DTPA), dynamic MR imaging studies were performed on normal and hepatocellular carcinoma (HCC) rat models using a 1.5-Tesla MR scanner. The results indicate that normal rats that were injected with 0.1 mmol/kg Gd-Bz-TTDA showed significantly more intense and persistent liver enhancement than those that were injected with the same dose of Gd-EOB-DTPA or Gd-BOPTA. All of these agents showed similar enhancement patterns in the implanted HCC. The liver-lesion contrast-to-noise ratios were higher and more persistent in rats that were injected with Gd-Bz-TTDA. These results indicate that Gd-Bz-TTDA is comparable with the commercially available hepatobiliary agents, Gd-EOB-DTPA and Gd-BOPTA, and can result in more intense and prolonged liver enhancement while still providing better liver-lesion discrimination. These results warrant further large-scale studies.