NK cells from COVID-19 positive patients exhibit enhanced cytotoxic activity upon NKG2A and KIR2DL1 blockade.

SARS CoV-2 has caused a global pandemic leading to significant morbidity and mortality. There is a need to elucidate and further understand the implications of COVID-19 disease on the immune system to develop improved therapeutic strategies. In particular, Natural Killer (NK) cells play an essential role in mediating the innate immune response against viral infections. To better understand the role of innate immunity in COVID-19, we characterized the phenotype of circulating NK cells from 74 COVID-19 patients and 25 controls. Through evaluating the protein expression of activating and inhibitory NK cell surface molecules using dimension reduction analysis and clustering, we identified 4 specific clusters of NK cells specific to disease state (COVID-19 positive or COVID-19 negative) and characterized COVID-19 positive NK cells as: NGK2A+KIR2DL1+NKG2C-. Utilizing blocking antibodies specific for receptors NKG2A and KIR2DL1, we found that both NKG2A and KIR2DL1 blockade markedly enhances the ability of NK cells from COVID-19 positive patients to lyse SARS-Cov-2 infected cells. Overall, this study reveals new insights into NK cell phenotypes during SARS-CoV-2 infection and suggests a therapeutic approach worthy of further investigation to enhance NK cell-mediated responses against the virus.

An electron microscopic and biochemical study of the potential protective effect of ginger against Cadmium-induced testicular pathology in rats.

Background: Cadmium (Cd) is a toxic heavy metal used in many industries. Since the second half of the 20th century, legislation on Cd use was put to limit the exponential rise in its environmental levels. This study aimed to investigate Cd's functional and ultrastructural changes on rats' reproductive systems and the role of Zingiber officinale (Ginger) in protecting against Cd-induced toxicity. Methods: Thirty adult male albino rats were randomly assigned into three equal groups (n = 10); control, Cd-exposed/untreated, and Cd-exposed/Gin-treated. Rat testes were weighed, and testicular tissue sections were examined under the electron microscope. Semen analysis, morphological examination of spermatozoa, and serum levels of luteinizing hormone (LH), follicle-stimulating hormone (FSH), and testosterone were measured. In addition, testicular tissue homogenates were analyzed for malondialdehyde (MDA), nitric oxide (NO), and reduced glutathione (GSH) levels. Results: Cd-induced significant reduction in the mean testicular weight and GSH levels and plasma testosterone, LH and FSH levels with a concomitant increase in testicular MDA and NO levels. There was also a deterioration in semen analysis parameters and spermatozoa morphology, with testicular structural damage in the form of architecture distortion and necrosis of seminiferous tubules and testicular interstitial cells. Daily administration of ginger for 4 weeks protected against CD-induced toxicity, preserving tissue architecture, improved plasma levels of testosterone, LH and FSH and testicular levels of GSH, and reduced testicular levels of MDA, NO. Conclusion: Ginger has a protective effect on Cd-induced deterioration of testicular tissue's structural and functional integrity by improving testicular tissue antioxidant capacity and steroid production, which ameliorates sex hormone levels in the blood.

Identification of "Double Hit" Lymphomas Using Updated WHO Criteria: Insights From Routine MYC Immunohistochemistry in 272 Consecutive Cases of Aggressive B-Cell Lymphomas.

Aggressive lymphomas with MYC and BCL2 and/or BCL6 translocations ("double hit" lymphomas, DHL) represent a distinct diagnostic category in the updated World Health Organization (WHO) classification. The diagnostic yield of MYC immunohistochemistry (IHC) for the identification of DHL is currently uncertain. MYC IHC was performed in 272 consecutive cases of aggressive B-cell lymphoma, and results correlated with fluorescence in situ hybridization (FISH) for MYC translocations. Among 156 patients with IHC and FISH data, MYC IHC identified MYC translocations with 89% sensitivity, 38% specificity, 92% negative predictive value, and 29% positive predictive value. Three of 15 (20%) of DHL were MYC IHC negative. One case contained a MYC translocation detectable IGH/MYC fusion probes but not MYC break-apart probes. A subset of DHL lack MYC protein expression, and recognition of this subset of cases requires FISH testing. These results provide an appropriate diagnostic algorithm for implementation of 2016 WHO diagnostic criteria.

Erdheim Chester disease in a patient with Burkitt lymphoma: a case report and review of literature.

BACKGROUND: Erdheim Chester disease (ECD) is a rare, non-Langerhans cell histiocytosis characterized by widespread tissue infiltration by CD68-positive, CD1a-negative foamy histiocytes. ECD can be difficult to identify, and diagnosis relies on the presence of histiocytes with certain histologic and immunophenotypic features in an appropriate clinical and radiologic setting. Clinical signs and symptoms are variable depending on which organ systems are involved. Most patients have at least skeletal involvement with bone pain as well as fatigue. Other common manifestations include diabetes insipidus, cardiac, periaortic, or retro-orbital infiltration/fibrosis, kidney impairment, xanthelasmas, among others. CASE PRESENTATION: Herein, we describe a case of BRAF-mutation positive ECD in a patient with Burkitt lymphoma, and we review recent literature. CONCLUSION: Underlying BRAF and other MAPK pathway mutations are identified in approximately 50% of cases of ECD, which aids in diagnosis as well as enables novel targeted treatments. ECD patients have an increased risk of myeloid neoplasms; however, unlike other histiocytoses, an association with lymphoproliferative disorders has not been recognized.

pSTAT3/pSTAT5 Signaling Patterns in Molecularly Defined Subsets of Myeloproliferative Neoplasms.

BCR/ABL1-negative myeloproliferative neoplasms (MPNs) are characterized by recurrent mutations in JAK2, CALR, and MPL, each of which has been reported to alter JAK/STAT signaling pathways. This report characterizes JAK/STAT signaling patterns in molecularly defined subsets of MPN utilizing immunohistochemistry for pSTAT3 and pSTAT5. Analysis of 30 BCR/ABL1-negative, nonpolycythemia vera MPN identified 15 (50%) with JAK2 V617F, 2 with MPL mutations (7%), and 8 with CALR mutations (27%). All mutations were mutually exclusive, except for 1 case with concurrent JAK2 V617F and CALR mutations. pSTAT3 staining in megakaryocyte nuclei was found in 4 cases (13%) and was not significantly associated with mutation status. pSTAT5 staining in megakaryocyte nuclei was found in 16 cases (53%), as was significantly associated with JAK2 V617F versus CALR mutation (P=0.009). Erythroid staining for pSTAT5 was seen exclusively in "triple-negative (TN)" cases lacking JAK2 V617F, MPL, and CALR mutations (P=0.006, TN vs. other genotypes), and pSTAT5 staining in megakaryocyte nuclei was seen in 2 TN cases. pSTAT5 staining in TN MPN suggests that other unknown abnormalities in this pathway may contribute to the pathogenesis of these cases. Furthermore, the demonstration of distinct STAT staining patterns in molecularly defined MPN suggests that these mutations result in divergent signaling events that may contribute to the biological and prognostic differences in these molecular subsets of MPN.

LEF-1 is a Sensitive Marker of Cribriform Morular Variant of Papillary Thyroid Carcinoma.

Cribriform morular variant of PTC (CMV-PTC) frequently shows activation of the CTNNB1/Wnt pathway with nuclear accumulation of beta catenin. The utility of LEF-1, also in the CTNNB1/WNT pathway, in the diagnosis of CMV-PTC has not been previously studied. LEF-1 immunohistochemistry was performed on seven CMV-PTC, 52 benign cases and 101 malignant thyroid neoplasms. LEF-1 was scored by stain intensity (0 = no nuclear stain, 1 = weak nuclear stain, less than lymphocyte and 2 = strong nuclear stain, intense as lymphocyte) and percentage of positive cells at each intensity, for a maximum total score of 200. Sensitivity and specificity of LEF-1 stain for all cases and to differentiate between regular PTC and CMV-PTC was also calculated. Six of the seven CMV-PTCs showed ≥ 30% strong (2+) nuclear LEF-1 staining and a total score over 100. Beta catenin also showed strong and diffuse nuclear staining in these cases. One CMV-PTC was negative for both LEF-1 and beta catenin and did not have a history of FAP. All control PTC cases uniformly lacked LEF-1 staining at 2+ intensity. LEF-1 had a sensitivity of 86% and specificity of 98% for the diagnosis of CMV-PTC. LEF-1 is highly sensitive and specific marker for CMV-PTC, especially when used in the setting of a PTC neoplasm. The pattern of staining is important with ≥ 30% of cells showing strong 2+ nuclear staining having the highest combined sensitivity and specificity.

cMYC expression in thyroid follicular cell-derived carcinomas: a role in thyroid tumorigenesis.

BACKGROUND: cMYC regulates approximately 15% of human genes and is involved in up to 20% of all human cancers. Reports discussing cMYC protein expression in thyroid carcinomas are limited, with controversies pertaining to cMYC expression patterns noted in the literature. The aims of the current study were to clarify patterns and intensities of cMYC expression in follicular cell-derived thyroid carcinomas across a spectrum of cancer morphologies and disease aggressivities, to correlate cMYC with BRAFV600E expression, and to evaluate the potential role of cMYC in progression of well-differentiated thyroid carcinomas into less well-differentiated carcinomas. METHODS: Immunohistochemical studies using specific monoclonal antibodies for cMYC and BRAFV600E were performed on tissue microarrays built from follicular cell-derived thyroid carcinomas (25 papillary, 24 follicular, 24 oncocytic variant of follicular, and 21 undifferentiated). In addition, cMYC IHC testing was also performed on whole tissue tumor sections from a subset of patients. Nodular hyperplasia cases were used as non-neoplastic controls. Appropriate positive and negative controls were included. RESULTS: cMYC was expressed almost exclusively in a nuclear fashion in both thyroid carcinomas and nodular hyperplasias. cMYC expression was weakly positive in both nodular hyperplasias and well-differentiated carcinomas. The majority of undifferentiated carcinomas (UDCs) showed strong nuclear cMYC positivity. PTC cases that were positive for cMYC (6/25) harbored the BRAF V600E mutation. A correlation was confirmed between cMYC intensity and tumor size in UDCs. UDC cases that developed out of well-differentiated thyroid carcinomas showed frank overexpression of cMYC in the undifferentiated tumor components. CONCLUSIONS: Our study suggests that nuclear overexpression of cMYC correlates with tumorigenesis / dedifferentiation in follicular cell derived thyroid carcinomas, a concept that has not been shown before on whole tissue sections.

Cytologic perspectives on neoteric B-cell lymphoproliferative disorders.

The 2008 World Health Organization (WHO) classification of hematopoietic and lymphoid tissues has been recently revised, and publication of the updated 2016 version is expected soon. Given that cytopathologists are often involved in the diagnosis of primary, recurrent, and transformed lymphoproliferative disorders, knowledge of updates to the WHO lymphoma classification, including terminology, pathogenesis, ancillary techniques, and targeted therapies is necessary. Herein, we reference the last decade of cytology specific literature for seven newer B-cell disorders and provide illustrative examples of each entity from our files.

Anaplastic lymphoma kinase positive large B-cell lymphoma: Literature review and report of an endoscopic fine needle aspiration case with tigroid backgrounds mimicking seminoma.

Anaplastic lymphoma kinase-positive large B-cell lymphoma (ALK+ LBCL) is a rare distinct type of non-Hodgkin's lymphoma that arises in association with alterations of the ALK gene. This distinct disease entity is typically associated with an aggressive clinical course and appears in light microscopic preparations as a monomorphic population of large, immunoblast-like cells. In this report, we describe a case of ALK+ LBCL diagnosed by transgastric endoscopic ultrasound-guided fine needle aspiration (EUS FNA) of splenic hilar lymph nodes. Modified Giemsa stained direct smears from the FNA sample demonstrated large lesional cells with foamy cytoplasm and macronucleoli admixed with small lymphocytes in tigroid backgrounds, mimicking the cytologic appearance of seminoma. Ancillary immunohistochemical studies subsequently confirmed the diagnosis of ALK+ LBCL with the lesional cells being immunoreactive for CD138, VS38c, MUM1, ALK1, and lambda light chain. The cohesiveness of the cells, the cellular morphology, and the tigroid backgrounds were all pitfalls for accurate diagnosis of this rare specific type of lymphoid malignancy by cytology. To our knowledge this is the first case report detailing the diagnosis of ALK+ LBCL by EUS FNA and the first report describing a glycogen-rich tigroid background in direct FNA smears. Establishing a refined diagnosis in cases of this rare form of LBCL is necessary, as therapies targeting ALK may be of value in clinical management. Diagn. Cytopathol. 2017;45:148-155. © 2016 Wiley Periodicals, Inc.

Characterization of an Oncolytic Adenovirus Vector Constructed to Target the cMet Receptor.

The cMet receptor is a homodimer with tyrosine kinase activity. Upon stimulation with its ligand, hepatocyte growth factor (HGF), the receptor mediates wide physiologic actions. The HGF-cMet signaling pathway is dysregulated in many cancers, which makes cMet an important target for novel therapeutic interventions. Oncolytic adenoviruses (Ads) have been used for the past three decades as a promising therapeutic approach for a wide array of neoplastic diseases. To date, achieving cancer-specific replication of oncolytic Ads has been accomplished by either viral genome deletions or by incorporating tumor selective promoters. To achieve novel specificity of oncolytic Ad infection of cancer cells that overexpress cMet, we inserted the HGF NK2 sequence, corresponding to a competitive antagonist of HGF binding to the cMet receptor, into the Ad serotype 5 (Ad5) fiber gene. The resulting vector, Ad5-pIX-RFP-FF/NK2, was rescued, amplified in HEK293 cells, and characterized. Binding specificity and viral infectivity were tested in various cancer cell lines that express varying levels of cMet and hCAR (the Ad5 receptor). We found that Ad5-pIX-RFP-FF/NK2 demonstrated binding specificity to the cMet receptor. In addition, there was enhanced viral infectivity and virus replication compared with a non-targeted Ad vector. Although NK2 weakly induces cMet receptor activation, our results showed no receptor phosphorylation in the context of an oncolytic Ad virus. In summary, these results suggest that an oncolytic Ad retargeted to the cMet receptor is a promising vector for developing a novel cancer therapeutic agent.