Lineage determination of CD20- B-cell neoplasms: An immunohistochemical study

We studied 61 CD20- B-cell lymphomas, including 29 cases of precursor B-cell lymphoblastic leukemia/lymphoblastic lymphoma (B-ALL/B-LBL), 25 cases of CD20- recurrent mature B-cell lymphoma after rituximab therapy, and 7 cases of CD20- diffuse large B cell lymphoma (DLBCL). We used markers specific for B lineage: CD79a, Pax-5, OCT.2, and BOB.1. All B-ALL/B-LBLs expressed Pax-5 (29/29 [100%]), 25 (93%) of 27 expressed BOB.1, 23 (79%) of 29 expressed CD79a, and 6 (22%) of 27 expressed OCT.2. The percentages of cases expressing Pax-5, CD79a, OCT.2, and BOB.1 in CD20- recurrent mature B-cell lymphomas after rituximab treatment were 88% (21/24), 84% (21/25), 81% (17/21), and 73% (16/22), respectively. CD20- DLBCLs rarely express routine B-lineage markers, such as CD79a and Pax-5, but they expressed OCT.2 or BOB.1. Pax-5, BOB.1, and CD79a antigens are the most reliable B-lineage markers for paraffin immunophenotyping B-ALL/B-LBL. CD79a and Pax-5 should be used as the first-line B lineage-specific markers for rituximab-treated CD20-mature B-cell lymphomas. If negative, OCT.2 or BOB.1 may be useful. The newly identified B-lineage markers, OCT.2 and BOB.1, may be the most useful for the B-lineage determination of CD20-plasmablastic or primary effusion subtypes of DLBCL. © American Society for Clinical Pathology

Prevention of early renal disease, dyslipidaemia and lipid peroxidation in STZ-diabetic rats by LR-9 and LR-74, novel AGE inhibitors

Background: Increased formation of advanced glycation/lipoxidation end-products (AGEs/ALEs) has been implicated in the pathogenesis of various diabetic complications. Several compounds have been developed as inhibitors of AGE/ALE formation. We examined the effects of two new AGE/ALE inhibitors, LR-9 and LR-74, on the development of early renal disease and lipid metabolism in streptozotocin (STZ)-induced diabetic rats. Methods: Diabetic Sprague-Dawley rats were treated with either of the LR compounds for 32 weeks. Progression of renal disease was evaluated by measurements of urinary albumin and plasma creatinine concentrations. AGE/ALE and nitrotyrosine levels in kidneys were determined by immunohistochemistry. AGE-induced chemical modification of the tail tendon collagen and levels of Nε-(carboxymethyl) and (carboxyethyl)- lysines (CML and CEL) in skin collagen were measured. Plasma lipids and their lipid hydroperoxide concentrations were also determined. In vitro, both compounds were tested for inhibiting lipid peroxidation reactions. Results: Treatment of either LR compounds significantly inhibited the increase in albuminuria, creatinaemia, hyperlipidaemia and lipid peroxidation in diabetic rats without any effect on hyperglycaemia. Both compounds also reduced CML-AGE and nitrotyrosine accumulation in kidney glomeruli and tubules, AGE-linked fluorescence and cross-linking of tail collagen, and levels of CML and CEL in skin collagen. In vitro, LR compounds inhibited the oxidation of human low-density lipoprotein (LDL). Conclusion: Both compounds can inhibit the progression of renal disease and also prevent dyslipidaemia in type-1 diabetic animals. These compounds may have an additional beneficial effect as an antioxidant against lipid peroxidation, and thus may provide alternative therapeutic options for the treatment of various diabetic macrovascular complications. Copyright © 2005 John Wiley & Sons, Ltd

Stromal response to prostate cancer: nanotechnology-based detection of thioredoxin-interacting protein partners distinguishes prostate cancer associated stroma from that of benign prostatic hyperplasia.

Histological staining of reactive stroma has been shown to be a predictor of biochemical recurrence in prostate cancer, however, molecular markers of the stromal response to prostate cancer have not yet been fully delineated. The objective of this study was to determine whether or not the stromal biomarkers detected with a thioredoxin-targeted nanodevice could be used to distinguish the stroma associated with benign prostatic hyperplasia from that associated with PCA. In this regard, we recently demonstrated that a thioredoxin-targeted nanodevice selectively binds to reactive stroma in frozen prostate tumor tissue sections. To accomplish this, random frozen prostate tissue sections from each of 35 patients who underwent resection were incubated with the nanodevice and graded for fluorescent intensity. An adjacent section from each case was stained with Hematoxylin & Eosin to confirm the diagnosis. Select cases were stained with Masson's Trichrome or immunohistochemically using antibodies to thioredoxin reductase 1, thioredoxin reductase 2 or peroxiredoxin 1. Our results demonstrate that the graded intensity of nanodevice binding to the stroma associated with PCA was significantly higher (p = 0.0127) than that of benign prostatic hyperplasia using the t-test. Immunohistochemical staining of adjacent sections in representative cases showed that none of the two commonly studied thioredoxin interacting protein partners mirrored the fluorescence pattern seen with the nanodevice. However, thioredoxin reductase 2 protein was clearly shown to be a biomarker of prostate cancer-associated reactive stroma whose presence distinguishes the stroma associated with benign prostatic hyperplasia from that associated with prostate cancer. We conclude that the signal detected by the nanodevice, in contrast to individual targets detected with antibodies used in this study, originates from multiple thioredoxin interacting protein partners that distinguish the M2 neutrophil and macrophage associated inflammatory response in prostate cancer-associated stroma from the CD4+ T-Lymphocyte linked inflammation in benign prostatic hyperplasia

Overlapping features between dedifferentiated liposarcoma and undifferentiated high-grade pleomorphic Sarcoma

Dedifferentiated liposarcoma (DDL), occurring in up to 10% of well differentiated liposarcoma cases, has similar histologic features to that of undifferentiated high-grade pleomorphic sarcoma (UHGPS); the former develops in a background of atypical lipomatous tumors/well differentiated liposarcoma, whereas the latter shows no specific line of differentiation. The retroperitoneum and thigh represent the most common anatomic locations for both the sarcomas. Despite their morphologic similarity, the issue of whether these 2 sarcomas share overlapping immunohistochemical and molecular features has not been well studied. We examined the expression of the lipogenic tumor-related markers peroxisome proliferator-activated receptor γ (PPAR-γ), CDK4, and MDM2 in 15 cases of DDL and 45 cases of retroperitoneal/thigh UHGPS. Patients with DDL ranged from 31 to 82 years (mean 63y) with a male:female ratio of 5:3. Patients with UHGPS ranged from 14 to 80 years (mean 52y) with a male:female ratio of 3:2. All 15 DDLs expressed CDK4 and MDM2 (100%), and 8 of 15 cases expressed PPAR-γ (53%). Twenty-three of 45 (51%) UHGPS expressed at least 1 of these 3 markers. We also studied MDM2 and CDK4 gene amplification by fluorescence in situ hybridization in 28 immunohistochemically positive cases, including 5 DDLs and 23 UHGPSs. All 5 cases of DDL showed MDM2 and/or CDK4 amplification (100%), whereas 6 of 45 UHGPSs showed MDM2 and/or CDK4 amplification (13%). Our results demonstrate that (1) the lipogenic tumor markers CDK4 and MDM2 can be used as surrogate immunohistochemical markers for the diagnosis of malignant lipomatous tumors with high sensitivity; (2) approximately 26% of retroperitoneal/thigh UHGPS cases that were positive for PPAR-γ, CDK4, or MDM2 by immunohistochemistry showed characteristic CDK4 and MDM2 gene amplification, suggesting that a subset of UHGPS cases represent DDL despite lacking histologic evidence of lipoblasts. Copyright © 2009 by Lippincott Williams & Wilkins

Biomarker identification with ligand-targeted nucleoprotein assemblies

Aims: Since many biomarkers of both the tumor and its microenvironment are expected to involve differential expression of divalent proteins capable of protein or peptide ligand interaction, we are developing multivalent nanodevices for the identification of biomarkers in prostate cancer. Patients & Methods: We compared a multivalent thioredoxin-targeted nanodevice with monovalent thioredoxin in binding to human prostate cell line(s) and freshly frozen tissue specimens obtained after resection from patients with biopsy-proven prostate cancer. Conclusion: The nanodevice binds specifically with enhanced avidity to tumor microenvironment-associated stromal cells in prostate cancer tissue specimens. Cells that bind the nanodevice also reacted with antibodies to dimeric thioredoxin reductases 1 and 2, suggesting the utility of the nanodevice as a potentially specific and functional marker of tumor stromal cells. © 2011 Future Medicine Ltd

Distinction of hepatocellular carcinoma from benign hepatic mimickers using Glypican-3 and CD34 immunohistochemistry

Distinguishing a well-differentiated hepatocellular carcinoma (HCC) from normal and cirrhotic liver tissue or benign liver nodules, such as hepatic adenoma (HA) and focal nodular hyperplasia (FNH), may be very difficult in some cases, particularly in small needle core biopsies. We studied the expression of Glypican-3 (GPC3) and CD34 in 107 cases of HCC, 19 cases of HA, and 16 cases of focal nodular hyperplasia (FNH). In addition, we studied GPC3 expression in 225 cases of nonhepatic human tumors with epithelial differentiation. Ninety-four of 107 cases (88%) of HCC showed focal or diffuse cytoplasmic GPC3 staining, whereas all HA and FNH cases were GPC3-negative, and only 7 of 225 cases (3%) of nonhepatic tumors with epithelial differentiation expressed GPC3. The sensitivity and specificity of GPC3 for HCC was 88% and 97%, respectively. There were three CD34 staining patterns observed in hepatic tissue: negative, incomplete positive, and complete positive. In negative staining pattern, only blood vessels in portal triads or rare sinusoidal spaces immediately adjacent to portal tracts were positive. The negative staining pattern was seen in normal or cirrhotic liver tissue only. The complete CD34 staining pattern showed virtually all sinusoidal spaces with CD34-positive staining throughout the lesion. The complete CD34 staining pattern was seen in virtually all cases of HCC and in only some cases of HA and FNH. The incomplete CD34 staining pattern was characterized by either CD34 positivity in virtually all sinusoidal spaces in some but not all nodules or CD34 positivity in the peripheral sinusoidal spaces adjacent to portal triads. The incomplete CD34 staining pattern was seen in rare cases of HCC and in most cases of HA and FNH. We conclude that GPC3 is a very specific marker not only for differentiating HCC from nonhepatic tumors with epithelial differentiation, but also for differentiating HCC from HA and FNH. GPC3 immunoreactivity, in combination with a complete CD34 immunostaining pattern, greatly facilitates the accuracy of distinguishing between malignant hepatic lesions and benign mimickers. © 2008 Lippincott Williams & Wilkins, Inc

Distinguishing PCA from BPH Based on Nanodevice Binding to Thioredoxin Interacting Proteins.

<p>Surgically resected tissue specimens were obtained from 35 patients. Serial sections of the tissue specimens (5 µm thick) were incubated with 20 nM of the nanodevice in PBS and 1% BSA. Fluorescence binding was observed and a numerical grading system was given as the level of nanodevice fluorescence in the stroma by two independent investigators (the author EMS and GB from acknowledgements). <b>0-</b> No visible signal, <b>1-</b> weakly visible signal, <b>2-</b> a moderately intense visible signal and <b>3-</b> a bright and intense signal within the reactive stroma. The binding level for PCA-associated stroma was significantly greater than that of BPH (p = 0.0127) based on the t-test.</p

Schematic Diagram of the Self-Assembly Sequence for the Nanodevice (NP-Trx₃).

<p>The fluorescein-labeled nanodevice (ND-Trx₃) displays three copies of the bacterial thioredoxin (Trx) as a cellular targeting ligand. It is assembled by annealing three synthetic oligodeoxynucleotides containing fluoresein (*) at a centrally-located site and 5-Fluorocytosine (F) at each of the three methyltransferase recognition sites, followed by covalent linkage of methyltransferase fusion proteins as previously described <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060562#pone.0060562-Huber1" target="_blank">[19]</a>. Fluorescence labeling permits visualization of the bound device with fluorescence microscopy. Multivalency improves the avidity of the device since many of the known thioredoxin interacting partners (<i>e.g.</i> the human thioredoxin reductases) are dimeric <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060562#pone.0060562-Singer1" target="_blank">[9]</a>.</p

ND-Trx₃ Binding to PCA and BPH in Frozen Tissue Sections.

<p>Representative frozen tissue sections were incubated for 1 minute with 20nM of the fluorescence-labeled ND-Trx₃ in 200 µl of ice cold PBS or with 200 µl of PBS alone as control. Adjacent sections were also stained with H&E and Masson's Trichrome. The slides were then analyzed by fluorescent microscopy and a tiled image of the entire tissue slice was obtained to identify regions of ND-Trx₃ binding. Tiled images photographed at 100X magnification allowed visualization of the entire tumor specimen after size reduction. Tumor (T) and Stromal (S) regions are indicated in each panel. PBS: <u>P</u>hosphate <u>B</u>uffered <u>S</u>aline, H&E: Hematoxylin and Eosin. Trichrome-stained sections were counter stained with hematoxylin (blue nuclear stain). The pattern of fluorescence observed with the nanodevice in the cancer specimen (<b>A</b>) was confined to the stromal region identified as the violet region in the adjacent section stained with Masson's Trichrome (<b>C</b>). Control BPH sections (<b>B</b>) did not bind the nanodevice significantly and yielded very low levels of fluorescence, and only weak staining with Masson's Trichrome (<b>D</b>). H&E stained sections containing tumor gave blue color in the tumor regions and light brown staining in regions of reactive stroma (<b>E</b>) and with BPH (<b>F</b>).</p

Porous Polymer Structures with Tunable Mechanical Properties Using a Water Emulsion Ink

Recently, the manufacturing of porous polydimethylsiloxane (PDMS) with engineered porosity has gained considerable interest due to its tunable material properties and diverse applications. An innovative approach to control the porosity of PDMS is to use transient liquid phase water to improve its mechanical properties, which has been explored in this work. Adjusting the ratios of deionized water to the PDMS precursor during blending and subsequent curing processes allows for controlled porosity, yielding water emulsion foam with tailored properties. The PDMS-to-water weight ratios were engineered ranging from 100:0 to 10:90, with the 65:35 specimen exhibiting the best mechanical properties with a Young’s Modulus of 1.17 MPa, energy absorption of 0.33 MPa, and compressive strength of 3.50 MPa. This led to a porous sample exhibiting a 31.46% increase in the modulus of elasticity over a bulk PDMS sample. Dowsil SE 1700 was then added, improving the storage capabilities of the precursor. The optimal storage temperature was probed, with −60 °C resulting in great pore stability throughout a three-week duration. The possibility of using these water emulsion foams for paste extrusion additive manufacturing (AM) was also analyzed by implementing a rheological modifier, fumed silica. Fumed silica’s impact on viscosity was examined, revealing that 9 wt% of silica demonstrates optimal rheological behaviors for AM, bearing a viscosity of 10,290 Pa·s while demonstrating shear-thinning and thixotropic behavior. This study suggests that water can be used as pore-formers for PDMS in conjunction with AM to produce engineered materials and structures for aerospace, medical, and defense industries as sensors, microfluidic devices, and lightweight structures