Clinicopathologic features of lingual canine T-zone lymphoma.

Canine T-zone lymphoma (TZL) is a subtype of T-cell lymphoma characterized by unique histologic pattern and cytomorphology, immunophenotypic loss of CD45 expression, and an indolent clinical behaviour. Dogs with TZL typically present with 1 or more enlarged lymph nodes and/or lymphocytosis. We describe a novel extranodal presentation of TZL involving the tongue. Twelve dogs with tongue masses were diagnosed with lingual TZL based on a variable combination of immunophenotyping via flow cytometry, cytology, histopathology, immunohistochemistry and/or PCR for antigen receptor rearrangement (PARR) assay. Eleven dogs exhibited concurrent lymphocytosis and/or lymph node enlargement. Three cases were initially diagnosed as plasma cell tumours based on histology alone, thereby revealing a potential diagnostic challenge. Seven dogs achieved clinical remission and 4 achieved stable disease following variable treatment, consistent with the indolent nature of typical TZL involving the lymph nodes and peripheral blood. In 1 case the TZL resulted in progressive disease and failure to respond to treatment. In this case, the TZL exhibited histologic features of a higher grade neoplasm. This case series highlights a unique presentation of TZL and identifies a new differential diagnosis for lingual neoplasia. In this study, we characterize the clinical presentation, diagnostic features and patient outcomes of 12 dogs with lingual TZL.

Increased frequency of CD45 negative T cells (T zone cells) in older Golden retriever dogs.

T zone lymphoma (TZL) is characterized by the clonal expansion of T cells lacking expression of the pan-leukocyte antigen CD45 (TZ cells). A strong breed predisposition is observed in Golden retrievers. This study aimed to confirm aberrant CD45 mRNA expression and determine if Golden retrievers without clinical lymphoma have an increased frequency of circulating TZ cells. Gene expression analysis on confirmed TZL cases showed a significant decrease in CD45 expression compared to normal dogs. Peripheral blood samples from senior dogs, 242 Golden retrievers and 42 non-Golden retrievers, without evidence of lymphoproliferative disease were assessed for the presence of TZ cells by flow cytometry. Thirty-one percent of Golden retrievers had TZ cells compared to 14% of non-Golden retrievers. Thirty-four percent of Golden retrievers with TZ cells had a clonal T cell receptor gamma (TRG) gene rearrangement. Interestingly, 20% of Golden retrievers without TZ cells also had a clonal TRG rearrangement. Golden retrievers may have an increased risk of TZL due to an increased frequency of TZ cells.

Breed Distribution and Clinical Characteristics of B Cell Chronic Lymphocytic Leukemia in Dogs.

BACKGROUND: B-cell chronic lymphocytic leukemia (B-CLL) is the most common hematopoietic malignancy in humans in the developed world and the primary risk factor is genetic. Dogs also develop B-CLL, but there is no systematic description of the disease in dogs. Understanding the epidemiology of B-CLL in dogs may help practitioners recognize the disease and position the dog as a model for future genetic studies. OBJECTIVES: To describe B-CLL presentation in dogs, its clinicopathologic findings, and breed predisposition. ANIMALS: Four hundred and ninety-one dogs with B-CLL and 5,673 control dogs with suspicion of a lymphoproliferative disorder (LPD). METHODS: Retrospective cross-sectional study of dogs for which samples were submitted to the Colorado State University Clinical Immunology Laboratory for immunophenotyping between 2010 and 2014. To assess breed predilection, dogs with B-CLL were compared to those with suspicion of other LPDs using logistic regression. RESULTS: The median age was 11 years with no sex predilection. Half of the dogs presented with peripheral lymphadenopathy or splenomegaly and 26% had anemia. Eleven small-breed dogs had significantly increased odds of B-CLL. In addition, English Bulldogs had an increased risk and a unique presentation: these dogs were diagnosed at a median of 6 years and expressed lower class II MHC and CD25. CONCLUSIONS: B-cell chronic lymphocytic leukemia is overrepresented in small-breed dogs. Future genetic studies of these breeds may identify genetic risk factors. The unique presentation of English Bulldogs provides evidence of multiple forms of this disease. Additional studies are necessary to determine whether presenting signs are associated with survival.

Canine T-zone lymphoma: unique immunophenotypic features, outcome, and population characteristics.

BACKGROUND: Canine T-cell lymphoma (TCL) is clinically and histologically heterogeneous with some forms, such as T-zone lymphoma (TZL), having an indolent course. Immunophenotyping is an important tool in the classification of TCL in people, and can be equally useful in dogs. HYPOTHESIS/OBJECTIVES: We hypothesized that loss of expression of the CD45 antigen is a specific diagnostic feature of TZL. ANIMALS: Twenty dogs with concurrent histology and immunophenotyping by flow cytometry were studied in depth. An additional 494 dogs diagnosed by immunophenotyping were used to characterize the population of dogs with this disease. METHODS: Lymph node biopsies from 35 dogs with TCL were classified by 2 pathologists using WHO criteria. Twenty lymph nodes were from dogs with CD45- TCL and 15 were from CD45+ TCL. The pathologists were blinded to the flow cytometry findings. Outcome information was sought for the 20 dogs with CD45- lymphoma, and population characteristics of the additional 494 dogs were described. RESULTS: All 20 CD45- cases were classified as TZL. The 15 CD45+ cases were classified as aggressive TCL and are described in an accompanying paper. TZL cases had a median survival of 637 days. Examination of 494 additional dogs diagnosed with TZL by immunophenotyping demonstrated that 40% of cases are in Golden Retrievers, are diagnosed at a median age of 10 years, and the majority have lymphadenopathy and lymphocytosis. CONCLUSIONS: TZL has unique immunophenotypic features that can be used for diagnosis.

Effects of intermittent Pringle's manoeuvre on cirrhotic compared with normal liver.

BACKGROUND: Although patients with liver cirrhosis are supposed to tolerate ischaemia-reperfusion poorly, the exact impact of intermittent inflow clamping during hepatic resection of cirrhotic compared with normal liver remains unclear. METHODS: Intermittent Pringle's manoeuvre was applied during minor hepatectomy in 172 patients with a normal liver, 59 with chronic hepatitis and 97 with liver cirrhosis. To assess hepatic injury, delta (D)-aspartate aminotransferase (AST) and D-alanine aminotransferase (ALT) (maximum level minus preoperative level) were calculated. To evaluate postoperative liver function, postoperative levels of total bilirubin, albumin and cholinesterase (ChE), and prothrombin time were measured. RESULTS: Significant correlations between D-AST or D-ALT and clamping time were found in each group. The regression coefficients of the regression lines for D-AST and D-ALT in patients with normal liver were significantly higher than those in patients with cirrhotic liver. Irrespective of whether clamping time was 45 min or less, or at least 60 min, D-AST and D-ALT were significantly lower in patients with cirrhosis than in those with a normal liver. Parameters of hepatic functional reserve, such as total bilirubin, prothrombin time, albumin and ChE, were impaired significantly after surgery in patients with a cirrhotic liver. CONCLUSION: Patients with liver cirrhosis had a smaller increase in aminotransferase levels following portal triad clamping than those with a normal liver. However, hepatic functional reserve in those with a cirrhotic liver seemed to be affected more after intermittent inflow occlusion.

Impact of a left-lobe graft without modulation of portal flow in adult-to-adult living donor liver transplantation.

In adult-to-adult living donor liver transplantation (LDLT), left-lobe grafts can sometimes be small-for-size. Although attempts have been made to prevent graft overperfusion through modulation of portal inflow, the optimal portal venous circulation for a liver graft is still unclear. Hepatic hemodynamics were analyzed with reference to graft function and outcome in 19 consecutive adult-to-adult LDLTs using left-lobe grafts without modulation of graft portal inflow. Overall mean graft volume (GV) was 398 g, which was equivalent to 37.8% of the recipient standard liver volume (SV). The GV/SV ratio was less than 40% in 13 of the 19 recipients. Overall mean recipient portal vein flow (PVF) was much higher than the left PVF in the donors. The mean portal contribution to the graft was markedly increased to 89%. Average daily volume of ascites revealed a significant correlation with portal vein pressure, and not with PVF. When PVP exceeds 25 mmHg after transplantation, modulation of portal inflow might be required in order to improve the early postoperative outcome. Although the study population was small and contained several patients suffering from tumors or metabolic disease, all 19 patients made good progress and the 1-year graft and patient survival rate were 100%. A GV/SV ratio of less than 40% or PVF of more than 260 mL/min/100 g graft weight does not contraindicate transplantation, nor is it necessarily associated with a poor outcome. Left-lobe graft LDLT is still an important treatment option for adult patients.

Conversion of elective laparoscopic to open cholecystectomy between 1993 and 2004.

BACKGROUND: Laparoscopic cholecystectomy (LC) is the standard treatment for symptomatic gallbladder disease. The identification of factors that reliably predict the likely need to convert LC to an open procedure would provide short-term benefits in terms of patient education and postoperative expectations. METHODS: Between 1993 and 2004, 1179 elective LCs were attempted from a total of 1339 elective cholecystectomies. The change in conversion rate between 1993-1999 and 2000-2004 was analysed. Factors predictive of higher risk for conversion were also identified. RESULTS: Eighty-nine LCs (7.5 per cent) required conversion. Gallbladder wall thickness and a history of common bile duct (CBD) stones, treated by preoperative endoscopic sphincterotomy, were predictors of conversion. The proportion of patients who underwent LC was the same in 1993-1999 (87.5 per cent) and 2000-2004 (88.8 per cent), but the conversion rate increased significantly from 5.3 to 10.6 per cent in these two time intervals. In addition, the proportion of patients with a history of CBD stones rose significantly, from 6.4 per cent in 1993-1999 to 11.0 per cent in 2000-2004. CONCLUSION: The conversion rate increased over the 12-year interval of the study. A history of preoperative endoscopic sphincterotomy and a thickened gallbladder wall contributed to the likelihood of conversion.

Clinical significance of immunohistochemically detectable lymph node metastasis in adenocarcinoma of the ampulla of Vater.

BACKGROUND: The aim of this study was to assess the impact of immunohistochemically identified lymph node metastasis on survival in patients with carcinoma of the ampulla of Vater. METHODS: Three hundred and twenty-six regional lymph nodes dissected from pancreatoduodenectomy specimens from 25 patients with ampulla of Vater carcinoma were immunostained with anticytokeratin antibody (CAM 5.2). The clinicopathological significance of immunohistochemically detectable lymph node metastasis was evaluated and compared with that of other potential prognostic factors. RESULTS: The frequency of lymph node involvement in relation to the total number of dissected lymph nodes increased from 5.5 per cent (18 of 326) using haematoxylin and eosin staining to 9.5 per cent (31 of 326) using cytokeratin immunostaining (P < 0.001). Lymph node involvement was revealed by haematoxylin and eosin staining in eight of 25 patients and by cytokeratin immunostaining in 11 of 25 patients (P = 0.006). Absence of immunohistochemically detectable lymph node metastasis was identified as an independent predictor of improved postoperative survival. CONCLUSION: Immunostaining of dissected lymph nodes adds additional information to data obtained by conventional haematoxylin and eosin staining when determining the prognosis of patients with carcinoma of the ampulla of Vater.

Development of anti-influenza drugs: II. Improvement of oral and intranasal absorption and the anti-influenza activity of stachyflin derivatives.

The in-vivo anti-influenza-virus activity of Stachyflin derivatives (III and its phosphate ester, III-Phos), a new class of haemagglutinin fusion inhibitor, and the improvement of their absorption after oral or intranasal administration were studied in mice, rats, and ferrets. The absorption of III in PEG 4000 and III-Phos aqueous solution increased about three and four fold in AUC after oral administration to uninfected mice compared with that of 0.5% HPMC (hydroxypropyl-methylcellulose) suspension. Using a mouse influenza virus infection model, significant anti-influenza-virus activity was observed in infected mice treated orally with these compounds dissolved in PEG 4000 or distilled water, respectively, but not in mice treated with 0.5% HPMC. The in-vivo anti-influenza-virus activity in ferrets, a good model for influenza virus infection in man, was also studied. Although the concentration of III in plasma was above the IC50 against the influenza virus strain used for 6h after the oral administration of III in PEG 400 to uninfected ferrets, no in-vivo anti-influenza-virus activity was observed at the same dosage given 4 times daily for 3 days. The intranasal administration of III-Phos, which was expected to have a more notable in-vivo anti-influenza-virus activity, was examined. III-Phos, whose intranasal absorption had been improved by the modification of III with phosphate ester in rats, inhibited viral replication in the nasal cavity and suppressed influenza-virus-induced fever when administered intranasally to infected ferrets. This study demonstrates that intranasally administered compounds with anti-influenza-virus activity must permeate the nasal membranes to produce their anti-influenza-virus effect.

Identification of amino acids of influenza virus HA responsible for resistance to a fusion inhibitor, Stachyflin.

We have recently described a novel hemagglutinin (HA) conformational change inhibitor of human influenza virus, Stachyflin (Yoshimoto et al, Arch. Virol., 144, 1-14, 1999). Stachyflin-resistant variants of human influenza A/WSN/33 (H1N1) virus were isolated in vitro and the nucleotide sequences of their HA genes were determined. The relation of amino acid substitutions and Stachyflin resistance was analyzed with in vitro membrane fusion between HA-expressing cells and octadecylrhodamine (R18)-labelled chick erythrocytes (RBC). The amino acid substitutions, lysine to arginine at position 51 or lysine to glutamic acid at position 121 of the HA2 subunit of the HA protein was enough to confer a Stachyflin-resistant phenotype of HA protein. The molecular mechanism of anti-HA conformational change activity of Stachyflin is discussed.

Field infestation of rambutan fruits by internal-feeding pests in Hawaii.

More than 47,000 mature fruits of nine different varieties of rambutan (Nephelium lappaceum L.) were harvested from orchards in Hawaii to assess natural levels of infestation by tephritid fruit flies and other internal feeding pests. Additionally, harvested, mature fruits of seven different rambutan varieties were artificially infested with eggs or first-instars of Mediterranean fruit fly, Ceratitis capitata (Wiedemann), or oriental fruit fly, Bactrocera dorsalis (Hendel) (Diptera: Tephritidae) to assess host suitability. When all varieties were combined over two field seasons of sampling, fruit infestation rates were 0.021% for oriental fruit fly, 0.097% for Cryptophlebia spp. (Lepidoptera: Tortricidae), and 0.85% for pyralids (Lepidoptera). Species of Cryptophlebia included both C. illepida (Butler), the native Hawaiian species, and C. ombrodelta (Lower), an introduced species from Australia. Cryptophlebia spp. had not previously been known to attack rambutan. The pyralid infestation was mainly attributable to Cryptoblabes gnidiella (Milliere), a species also not previously recorded on rambutan in Hawaii. Overall infestation rate for other moths in the families Blastobasidae, Gracillariidae, Tineidae, and Tortricidae was 0.061%. In artificially infested fruits, both species of fruit fly showed moderately high survivorship for all varieties tested. Because rambutan has such low rates of infestation by oriental fruit fly and Cryptophlebia spp., the two primary internal-feeding regulatory pests of rambutan in Hawaii, it may be amenable to the alternative treatment efficacy approach to postharvest quarantine treatment.

Development of anti-influenza virus drugs I: improvement of oral absorption and in vivo anti-influenza activity of Stachyflin and its derivatives.

PURPOSE: Stachyflin and its derivatives which are active against the influenza virus in vitro, were studied to improve their reduced in vivo activity after oral administration by chemical modification and some vehicles. METHODS: The solubility was examined for different vehicles. The improvement of gastrointestinal absorption was evaluated by the plasma concentration after oral administration to mice or the in situ loop method with rats. The in vivo anti-influenza activity was examined using mice infected with the influenza virus and evaluated based on the virus titer in the lung by TCID50. RESULTS: PEG 400 showed the highest solubility of Stachyflin and its derivative among the vehicles studied. While no viral inhibition was found in the lung after oral administration of 0.5% HPMC suspension of Stachyflin, in vivo anti-influenza virus activity was found with the PEG 400 solution. The absorption of Stachyflin by PEG 400 showed about a fifty-fold increase in AUC compared with that of 0.5% HPMC suspension. Improving the oral absorption of Stachyflin led to an increase in the in vivo anti-influenza virus activity. When the Stachyflin derivative in PEG 4000 was administered orally, there was more enhancement of the oral absorption than with PEG 400. When the aqueous solution of the phosphate ester prodrugs of Stachyflin and its derivative was administered orally, the absorption of the parent compound was improved and in vivo anti-influenza virus activity was found. CONCLUSIONS: When Stachyflin and its derivatives were administered orally to mice with a solution in PEG and an aqueous solution of their phosphate ester, their oral absorption was improved and in vivo anti-influenza virus activity was observed.

Identification of a novel HA conformational change inhibitor of human influenza virus.

Stachyflin is a novel compound having H1 and H2 subtype-specific anti-influenza A virus activity. Stachyflin has no inhibition on H3 subtype influenza A or influenza B viruses. The susceptibility of the reassortant viruses between H1 and H3 subtype influenza A viruses to Stachyflin indicated that its target was virus-encoded hemagglutinin (HA). The results of the timing of Stachyflin addition against in vitro virus infection and virus-mediated hemolysis assay suggested that the drug inhibited the HA-mediated virus-cell fusion process. More directly, Stachyflin interfered with HA conformational change induced by low pH or heat treatment. The effect of Stachyflin could not be eliminated by washing of the Stachyflin-treated virus, which caused very specific virucidal effect. This is a remarkable property among small molecules which inhibit low-pH induced HA conformational change. From these findings, we concluded that the mechanism of Stachyflin action is to inhibit HA conformational change which is necessary for virus-cell membrane fusion. Stachyflin may be used as a tool for a study of molecular mechanism of low-pH induced HA conformational change, and offers potential as a pharmaceutical agent.

Usefulness of p53 protein, Bcl-2 protein and Ki-67 as predictors of chemosensitivity of malignant tumors.

In various types of human malignant tumors, the presence or absence of expression of apoptosis-associated gene products (p53 protein and Bcl-2 protein) and the tumor proliferation activity-related factor (Ki-67) was assessed by immunohistochemical staining and the correlation between this expression and chemosensitivity to anticancer drugs was investigated. Study subjects comprised 55 preoperative patients with untreated malignant tumors (9 with esophageal cancer, 11 with stomach cancer, 11 with colon cancer, 13 with hepatic cancer and 11 with breast cancer). A chemosensitivity test was carried out with the histoculture drug response assay (HDRA) method using 4 drugs, mitomycin C (MMC), 5-fluorouracil (5-FU), doxorubicin hydrochloride (ADM), and cisplatin (CDDP). Immunohistochemical staining was used to assess expression of p53 protein, Bcl-2 protein and Ki-67. The tumor growth inhibition index (I.I.) of the 4 drugs was significantly lower in a group of the patients with p53 protein overexpression-type (mutant p53 protein positive expression-type) tumors than in a group with p53 protein negative expression-type tumors (p<0.05). No significant correlation was found between the expression of the Bcl-2 protein by and the I.I. of any drug studied in any type of cancer. A negative correlation was found between the labeling index (L.I.) for Ki-67 in all cases and I.I. for MMC and ADM and thus, chemosensitivity of the tumors with high growth activity was lower. Furthermore, a positive correlation existed between the L.I. for Ki-67 and that for p53 protein. The patients with p53 protein overexpression-type (mutant p53 protein positive) tumors showed low chemosensitivity. In addition, overexpression of p53 protein is suggested to be one of the factors involved in the lowered chemosensitivity of the tumors with high growth activity. Summarizing these findings, the p53 protein can play an important role in cancer therapy.

Control of hemorrhage during AVM surgery--with special reference to treatment of dilated capillaries and arteries around the nidus.

Control of hemorrhage during AVM surgery is one of the key issues to prevent NPPB. Inadequate procedures for hemostasis of feeders and drainers, so-called dilated capillaries and arteries (moja moja blood vessels) that are located on the side facing the normal brain, and inappropriate surgical strategies for intraoperative hemorrhage from these blood vessels are frequently the main cause of the difficulty in achieving hemostatic control. We conclude that it is important to aggressively reduce the occurrence of intraoperative hemorrhage and prevent or minimize the dilatation of abnormal capillaries and arteries due to inappropriate surgical procedures on the basis of the fundamental surgical strategy, i.e., feeder-->nidus-->drainer. Adequately securing the length and adequate coagulation of hemorrhagic blood vessel, employing a skillful bipolar coagulator technique aimed at controlling intraluminal pressure and blood flow on the central side, are believed to be key factors in hemorrhage treatment.

Uprighting the mandibular molars stimulates mandibular growth during treatment of class II malocclusion.

We hypothesized that uprighting of the mandibular molars creates a counter-clockwise rotation of the mandible and stimulates mandibular forward growth during the treatment of a Class II malocclusion. This investigation used 33 longitudinal lateral cephalometric radiographs of Class II, Division 1 female patients. All cases were treated with non-extraction. Treatment was started in early adolescence with .018 slot edgewise Alexander appliances. High-pull head-gear and Class II elastics were used. Seventeen cases that showed more than 5 degrees of uprighting of the mandibular first molars were selected as the uprighted group. Cases that showed less than 5 degrees of uprighting of the mandibular first molars were selected as the non-uprighted group. There was a significant correlation coefficient between the uprighted degree of the mandibular first molars and the degree of clockwise rotation of the mandibular plane to FH.

Evidence for direct association of Vpr and matrix protein p17 within the HIV-1 virion.

Vpr is one of the auxiliary proteins of HIV-1 and is selectively incorporated into the virion by a process involving the C-terminal p6 portion of the Gag precursor Pr55. Vpr and the matrix protein p17 are the components of the viral preintegration complex and appear to play important roles in the nuclear transport of proviral DNA in nondividing cells. In the present study, we have demonstrated by coimmunoprecipitation experiments that Vpr associates with matrix protein p17 but not with capsid protein p24 within the HIV-1 virion. Experiments employing the yeast two-hybrid GAL4 assay for protein-protein interactions also demonstrated a direct association between Vpr and the C-terminal region of matrix protein p17. Association of Vpr and the matrix protein p17 within the mature virion is consistent with their collaborative role in the nuclear transportation of the viral preintegration complex in nondividing cells such as macrophages.

Targeting of chrolamphenicol acetyltransferase to human immunodeficiency virus particles via Vpr and Vpx.

Vpr and Vpx are the auxiliary proteins of human immunodeficiency viruses (HIVs) selectively incorporated into mature viral particles. We showed that the bacterial chloramphenicol acetyltransferase (CAT) fused to the N-terminus of HIV-1 Vpr, HIV-2 Vpr, or HIV-2 Vpx was incorporated into mature virions in a type-selective manner. By using chimeric proteins between HIV-1 Vpr and HIV-2 Vpx, we found that the N-terminal side of these proteins was mainly important for type-selective virion incorporation. The C-terminal arginine-rich region of HIV-1 Vpr was also found to transport CAT fusion proteins into virions but without any type selectivity. Furthermore, the corresponding regions of HIV-2 Vpr and HIV-2 Vpx had no such activity. This region of HIV-1 Vpr may interact nonspecifically with viral genomic RNA. Collectively, Vpr and Vpx may provide a means to introduce foreign proteins and other molecules into HIV virions for therapeutic purposes.