Melanocytic tumors with spitzoid morphology: correlation of clinicopathologic features and FISH analysis.

BACKGROUND: A subset of melanocytic tumors with spitzoid morphology may lead to potential inaccurate diagnosis and lack of assessment of malignancy potential. In this study, we aimed to evaluate melanocytic tumors with spitzoid morphology using conventional melanoma FISH (RREB-1, CCND1, MYB and CEP6) and 9p21 FISH (CDKN2A) probes and compare the probe results with clinical and histopathological features. METHODS: This study is a multicentric retrospective study including three centers, Istanbul University-Cerrahpasa, Cerrahpasa School of Medicine, Department of Pathology, Acibadem University, School of Medicine, Department of Pathology and ETA Pathology Laboratory. The pathology reports in archives of these three centers between 2015 and 2017 have been reviewed for cases diagnosed as atypical Spitz tumor or melanoma with Spitzoid features. These cases were selected for the study. We analyzed 39 cases of atypical Spitz tumor (AST), 10 cases of melanomas with spitzoid features for clinicopathological data and chromosomal alterations, targeting RREB-1 (6p25), CCND1 (11q13), MYB (6q23), together with 9p21 (CDKN2A), using FISH methodology. RESULTS: Thirty out of total 49 cases showed chromosomal alterations by 4-probe melanoma FISH assay, 22 (56.4%) cases were ASTs, and 8 (80%) cases were melanomas. Eighteen out of 49 cases showed homozygote deletion by 9p21 FISH assay, 12 (30.8%) cases were ASTs, and 6 (60%) cases were melanomas. When histopathological data were compared with FISH results, a statistically significant correlation was found between 9p21 FISH positivity (homozygous deletion) and presence of deep mitosis (p < 0.05). In addition, epidermal consumption (p = 0.07) and increased mitotic activity (p = 0.05) were more frequent in cases with homozygous 9p21 deletion, but these differences did not reach statistical significance. When the clinical features were considered, there was a statistically significant correlation between 9p21 FISH positivity and the diameter (p < 0.05). There was no statistically significant correlation between melanoma FISH assay and any of the histopathological or clinical data. DISCUSSION: These data suggest that 9p21 FISH positivity correlated with more worrisome histopathologic and clinical features, such as deep mitosis, increased mitotic activity, epidermal consumption, and larger lesion size, so these features are precious, pointing out spitzoid lesions with higher risk. However, there is a need for further studies using FISH or similar techniques in order to provide more accurate prognostic information in lesions Blank morphology.

Histologically benign PI-RADS 4 and 5 lesions contain cancer-associated epigenetic alterations.

BACKGROUND: The detection rate of clinically significant prostate cancer has improved with the use of multiparametric magnetic resonance imaging (mpMRI). Yet, even with MRI-guided biopsy 15%-35% of high-risk lesions (Prostate Imaging-Reporting and Data System [PI-RADS] 4 and 5) are histologically benign. It is unclear if these false positives are due to diagnostic/sampling errors or pathophysiological alterations. To better understand this, we tested histologically benign PI-RAD 4 and 5 lesions for common malignant epigenetic alterations. MATERIALS AND METHODS: MRI-guided in-bore biopsy samples were collected from 45 patients with PI-RADS 4 (n = 31) or 5 (n = 14) lesions. Patients had a median clinical follow-up of 3.8 years. High-risk mpMRI patients were grouped based on their histology into biopsy positive for tumor (BPT; n = 28) or biopsy negative for tumor (BNT; n = 17). From these biopsy samples, DNA methylation of well-known tumor suppressor genes (APC, GSTP1, and RARß2) was quantified. RESULTS: Similar to previous work we observed high rates of promoter methylation at GSTP1 (92.7%), RARß2 (57.3%), and APC (37.8%) in malignant BPT samples but no methylation in benign TURP chips. Interestingly, similar to the malignant samples the BNT biopsies also had increased methylation at the promoter of GSTP1 (78.8%) and RARß2 (34.6%). However, despite these epigenetic alterations none of these BNT patients developed prostate cancer, and those who underwent repeat mpMRI (n = 8) demonstrated either radiological regression or stability. CONCLUSIONS: Histologically benign PI-RADS 4 and 5 lesions harbor prostate cancer-associated epigenetic alterations.

Favorable locoregional control in clinically node-negative hormone-receptor positive breast cancer with low 21-gene recurrence scores: a single-institution study with 10-year follow-up.

BACKGROUND: Recent studies have shown a lower likelihood of locoregional recurrences in patients with a low 21-gene recurrence score (RS). In this single-institution study, we investigated whether there are any associations between different cutoff values of 21-gene RS, histopathological factors, and outcome in patients with long-term follow-up. METHODS: The study included 61 patients who had early-stage (I-II) clinically node-negative hormone receptor-positive and HER2-negative breast cancer and were tested with the 21-gene RS assay between February 2010 and February 2013. Demographic, clinicopathological, treatment, and outcome characteristics were analyzed. RESULTS: The median age was 48 years (range, 29-72 years). Patients with high histologic grade (HG), Ki-67 ≥ 25%, or Ki-67 ≥ 30% were more likely to have intermediate/high RS (≥ 18). Based on the 21-gene RS assay, only 19 patients (31%) received adjuvant chemotherapy. At a median follow-up of 112 months, 3 patients developed locoregional recurrences (4.9%), which were treated with endocrine therapy alone. Among patients treated with endocrine treatment alone (n = 42), the following clinicopathological characteristics were not found to be significantly associated with 10-year locoregional recurrence free survival (LRRFS): age < 40 years, age < 50 years, high histological or nuclear grade, high Ki-67-scores (≥ 15%, ≥ 20%, ≥ 25%, ≥ 30%), presence of lymphovascular invasion, luminal-A type, multifocality, lymph node positivity, tumor size more than 2 cm, RS ≥ 18, and RS > 11. However, patients with RS ≥ 16 had significantly poorer 10-year LRRFS compared to those with RS < 16 (75% vs. 100%, respectively; p = 0.039). CONCLUSIONS: The results suggest that patients with clinically node-negative disease and RS ≥ 16 are more likely to benefit from adjuvant chemotherapies. However, those with RS < 16 have an excellent outcome and local control in long-term follow-up with endocrine treatment alone.

Histopathological characteristics and CD163 immunostaining pattern in fibrous papule of the face.

BACKGROUND: Signs of inflammation including epidermal interface changes, spongiosis, and dermal inflammation as well as pagetoid dyskeratosis are rarely described in fibrous papule (FP). We aimed to describe the inflammatory parameters, the rate of pagetoid dyskeratosis, along with CD163 immunohistochemical staining as an adjunctive diagnostic tool in FP. METHODS: Histopathology samples of all biopsy-proven FP cases were retrieved from archives and investigated for inflammatory parameters, presence of pagetoid dyskeratosis, as well as CD163, CD10, and CD34 immunostaining pattern of dermal spindle/stellate or multinucleate cells (graded from 0 to 4). RESULTS: Thirty-two cases of FP were identified. A high rate of inflammatory parameters including interface changes (20/32), spongiosis (31/32), and dermal lymphocytic inflammation (31/32) were detected. Pagetoid dyskeratosis was identified in eight out of 32 cases (25%). A grade 4 staining revealing a strong dendritic pattern was confirmed in all FP cases with CD163 immunohistochemistry including atypical variants such as granular FP, compared with CD10 (11/32) and CD34 (3/32). CONCLUSION: The dendritic cellular proliferation in FP may represent an inflammatory response to various stimuli; pagetoid dyskeratosis is a relatively common and underrecognized epidermal feature and CD163 immunostaining may be used as an adjunctive diagnostic tool in unusual histopathological subtypes.

Does it take three to tango? An unsuspected multimorbidity of CD8+ T cell lymphoproliferative disorder, malaria, and EBV infection.

BACKGROUND: Malaria is known to cause acute and deadly complications. However, malaria can cause unforeseen pathologies due to its chronicity. It increases the risk of endemic Burkitt Lymphoma development by inducing DNA damage in germinal centre (GC) B cells, and leading higher frequency of Epstein-Barr virus (EBV)-infected cells in GCs. EBV is well known for its tropism for B cells. However, less is known about EBV's interaction with T cells and its association with T cell lymphoma. CASE PRESENTATION: A 43-year-old Sudanese male admitted to hospital in Istanbul, Turkey, a non-endemic country, with hyperpigmented painful skin rashes on his whole body. A complete blood count and a peripheral blood smear during admission revealed large granular lymphocytes (LGLs) with abnormally higher CD8 T cell numbers. Additional skin biopsy and pathology results were compatible with CD8+ T cell lymphoproliferative disorder with skin involvement. Patient was treated and discharged. However, a pathologist noticed unusual structures in skin tissue samples. Careful evaluation of skin biopsy samples by polarized microscopy revealed birefringent crystalloid structures resembling malarial haemozoin mainly loaded in macrophages and giant histiocytes. After purification of DNA from the skin biopsy samples, nested PCR was performed for the detection of Plasmodium parasites and Plasmodium falciparum DNA was amplified. Because, the co-presence of EBV infection with malaria is a well-known aetiology of lymphoma, EBV-early RNA (EBER) transcripts were investigated in paraffin-embedded tissue samples and found to be positive in macrophage-like histiocytes. CONCLUSIONS: This is a unique case of malaria and EBV infection in a T-LGL lymphoma patient who presented in a non-endemic country. This case emphasizes the clinical importance of EBV monitoring in T-LGL patients with skin involvement. Notably, Plasmodium infection should be examined in patients from malaria endemic regions by pathological and molecular investigations.

Intra-surgical total and re-constructible pathological prostate examination for safer margins and nerve preservation (Istanbul preserve).

PURPOSE: To demonstrate a novel frozen section analysis technique during robot assisted radical prostatectomy with 2 distinct advantages: evaluation of the entire circumference and easier reconstruction for whole mount evaluation. MATERIAL AND METHODS: Istanbul Preserve was performed on patients who underwent robotic prostatectomy with nerve sparing between 10/2014 and 7/2016. Gland was sectioned at 3-4mm intervals from apex to bladder neck. Entire tissue representing margins (except for the most anterior portion) was circumferentially excised and microscopically analyzed. In margin positivity, approach was individualized based on extent of positive margin and Gleason pattern. A matched cohort was established for comparison. Retrospective analysis of a prospectively maintained database was performed. Impact of FSA on PSM rate was primarily assessed. RESULTS: Data on 170 patients was analyzed. Positive surgical margin was reported in 56(33%) on frozen section. Neurovascular bundle was partially or totally resected in 79% and 18%. Conversion of positive margin to negative was achieved in 85%. Overall positive margin rate decreased from 22.5% to 7.5%. Nerve sparing increased from 87% to 93%. Location of positive margin at frozen was at the neurovascular bundle area in 39%; thus Istanbul Preserve detected 61% additional margin positivity compared to other techniques. Reconstruction for whole mount was easy. CONCLUSION: Istanbul Preserve is a novel technique for intraoperative FSA during RARP allowing for microscopic examination of the entire prostate for margin status and easy re-construction for whole mount examination. It guarantees safer margins together with increased rate of nerve sparing.

Cutaneous foreign body granulomas associated with lipolytic cocktail: Who is the enemy, mesotherapy or drugs injected?

Mesotherapy is widely used for its lipolytic effect as an alternative procedure to surgical methods. Although many benefits of lipolytic mesotherapy have been observed, numerous side effects have also been reported. Here, we report a case of cutaneous foreign body granulomas that occurred after lipolytic mesotherapy.

GnRH agonist leuprolide acetate does not confer any protection against ovarian damage induced by chemotherapy and radiation in vitro.

STUDY QUESTION: Is there any in vitro evidence for or against ovarian protection by co-administration of a GnRH agonist with chemotherapy in human? SUMMARY ANSWER: The co-administration of GnRH agonist leuprolide acetate with cytotoxic chemotherapy agents does not preserve ovarian reserve in vitro. WHAT IS KNOWN ALREADY: Randomized controlled trials of the co-administration of gonadotrophin-releasing hormone (GnRH) agonists with adjuvant chemotherapy to preserve ovarian function have shown contradictory results. This fact, together with the lack of a proven molecular mechanism of action for ovarian protection with GnRH agonist (GnRHa) places this approach as a fertility preservation strategy under scrutiny. We therefore aimed in this study to provide in vitro evidence for or against the role of GnRHa in the prevention of chemotherapy-induced damage in human ovary. STUDY DESIGN, SETTINGS, SIZE AND DURATION: This translational research study of ex vivo and in vitro models of human ovary and granulosa cells was conducted in a university hospital between 2013 and 2015. PARTICIPANTS/MATERIALS, SETTING, METHODS: Ovarian cortical pieces (n = 15, age 14-37) and mitotic non-luteinized (COV434 and HGrC1) and non-mitotic luteinized human granulosa cells (HLGC) expressing GnRH receptor were used for the experiments. The samples were treated with cyclophosphamide, cisplatin, paclitaxel, 5-FU, or TAC combination regimen (docetaxel, adriamycin and cyclophosphamide) with and without GnRHa leuprolide acetate for 24 h. DNA damage, apoptosis, follicle reserve, hormone markers of ovarian function and reserve (estradiol (E2), progesterone (P) and anti-mullerian hormone (AMH)) and the expression of anti-apoptotic genes (bcl-2, bcl-xL, bcl-2L2, Mcl-1, BIRC-2 and XIAP) were compared among control, chemotherapy and chemotherapy + GnRHa groups. MAIN RESULTS AND THE ROLE OF CHANCE: The greatest magnitude of cytotoxicity was observed in the samples treated with cyclophosphamide, cisplatin and TAC regimen. Exposure to these drugs resulted in DNA damage, apoptosis and massive follicle loss along with a concurrent decline in the steroidogenic activity of the samples. GnRHa co-administered with chemotherapy agents stimulated its receptors and raised intracellular cAMP levels. But it neither activated anti-apoptotic pathways nor prevented follicle loss, DNA damage and apoptosis induced by these drugs. LIMITATIONS, REASONS FOR CAUTION: Our findings do not conclusively rule out the possibility that GnRHa may offer protection, if any, through some other mechanisms in vivo. WIDER IMPLICATIONS OF THE FINDINGS: GnRH agonist treatment with chemotherapy does not prevent or ameliorate ovarian damage and follicle loss in vitro. These data can be useful when consulting a young patient who may wish to receive GnRH treatment with chemotherapy to protect her ovaries from chemotherapy-induced damage.

The magnitude of gonadotoxicity of chemotherapy drugs on ovarian follicles and granulosa cells varies depending upon the category of the drugs and the type of granulosa cells.

STUDY QUESTION: Do different chemotherapy drugs exert the same magnitude of cytotoxicity on dormant primordial follicles and the growing follicle fraction in the ovary in vivo and on mitotic non-luteinized and non-mitotic luteinized granulosa cells in vitro? SUMMARY ANSWER: Cyclophosphamide (alkylating agent) and cisplatin (alkylating like) impacted both primordial and pre-antral/antral follicles and both mitotic and non-mitotic granulosa cells, whereas the anti-metabolite cancer drug gemcitabine was detrimental only to pre-antral/antral follicles and mitotic non-luteinized granulosa cells. WHAT IS KNOWN ALREADY: It is already known that anti-metabolite cancer drugs are less detrimental to the ovary than alkylating and alkylating like agents, such as cyclophosphamide and cisplatin. This assumption is largely based on the results of clinical reports showing lower rates of amenorrhea in women receiving anti-metabolite agent-based regimens compared with those treated with the protocols containing an alkylating drug or a platinum compound. But a quantitative comparison of gonadotoxicity with a histomorphometric proof of evidence has not been available for many chemotherapy drugs. Therefore, we combined in this study in vivo and in vitro models of human and rat origin that allows a comparative analysis of the impact of different chemotherapy agents on the ovary and granulosa cells using real-time quantitative cell indices, histomorphometry, steroidogenesis assays, and DNA damage and cell death/viability markers. We also aimed to investigate if there is a difference between mitotic and non-mitotic granulosa cells in terms of their sensitivity to the cytotoxic actions of chemotherapy drugs with different mechanisms of action. This issue has not been addressed previously. STUDY DESIGN, SIZE, DURATION: This translational research study involved in vivo analyses of ovaries in rats and in vitro analyses of granulosa cells of human and rat origin. PARTICIPANTS/MATERIALS, SETTING, METHODS: For the in vivo assays, 54 4- to 6-week old Sprague-Dawley young female rats were randomly allocated into four groups of 13 to receive a single IP injection of: saline (control), gemcitabine (200 mg/kg), cisplatin (50 mg/kg) or cyclophosphamide (200 mg/kg). The animals were euthanized 72 h later. Follicle counts and serum AMH levels were compared between the groups. In vitro cytotoxicity studies were performed using mitotic non-luteinized rat (SIGC) and human (COV434, HGrC1) granulosa cells, and non-mitotic luteinized human (HLGC) granulosa cells. The cells were plated at a density of 5000 cells/well using DMEM-F12 culture media supplemented with 10% FBS. Chemotherapy agents were used at their therapeutic blood concentrations. The growth of mitotic granulosa cells was monitored real-time using xCelligence system. Live/dead cell and apoptosis assays were also carried out using intravital Yo-Pro-1 staining and cleaved caspase-3 expression, respectively. Estradiol (E2), progesterone (P) and anti-Mullerian hormone (AMH) levels were assayed with ELISA. MAIN RESULTS AND THE ROLE OF CHANCE: Cyclophosphamide and cisplatin caused massive atresia of both primordials and growing follicles in the rat ovary whereas gemcitabine impacted pre-antral/antral follicles only. Cyclophosphamide and cisplatin induced apoptosis of both mitotic non-luteinized and non-mitotic luteinized granulosa cells in vitro. By contrast, cytotoxicity of gemcitabine was confined to mitotic non-luteinized granulosa cells. LIMITATIONS, REASONS FOR CAUTION: This study tested only three chemotherapeutic agents. The experimental methodology described here could be applied to other drugs for detailed analysis of their ovarian cytotoxicity. WIDER IMPLICATIONS OF THE FINDINGS: These findings indicate that in vivo and in vitro cytotoxic actions of chemotherapy drugs on the ovarian follicles and granulosa cells vary depending upon the their mechanism of action and the nature of the granulosa cells.

Stem cell and extracellular matrix-related molecules increase following melatonin treatment in the skin of postmenopausal rats.

The menopause has a negative effect in the skin. Melatonin affects skin functions and structures through actions mediated by cell-surface and putative-nuclear receptors expressed in skin cell. We have therefore determined the effects of melatonin treatment on stem cell in the epidermis and extracellular matrix related molecules in the dermis the skin of postmenopausal rats. A total of 45 female rats were divided into 5 groups: control group, group A [ovariectomy (OVX)], group B (OVX +10 mg/kg/day melatonin), group C (OVX +30 mg/kg/day melatonin), group S (sham operated + 10 mg/kg/day melatonin). Ventral skin samples were excised at 12th week after ovariectomy. Hematoxylin-eosin, periodic acid- methylamine silver, elastic van Gieson staining techniques were used to measure histomorphometrically the thickness of elastic fibers and basement membrane, depths of the epidermis, dermis, and subcutaneous fat layer. Immunohistochemical staining methods were used for fibroblast growth factor ß (FGF ß), collagen type I, fibronectin, ß-catenin, c-kit, c-Myc evaluation. Epidermal thickness, subcutaneous fat layer, and elastic fibers were significantly decreased in group C, and there was a significant increase after melatonin treatment. Although there was no difference in dermal thickness of group C, melatonin also significantly increased the dermal thickness. High FGF ß, type I collagen, fibronectin, ß-catenin, c-Myc immunoreactivity developed following melatonin in all groups. Thus melatonin treatment of postmenopausal rats was mostly due to the decrease of stem cell and extracellular matrix-related molecules in the skin.