Eran Dvir

Clinical differentiation between dogs with benign and malignant spirocercosis

Spirocerca lupi is a nematode infesting the canine oesophagus, where it induces the formation of a nodule that may transform into a malignant sarcoma. The current, retrospective study compared the clinical presentation, haematology, serum albumin and globulin and radiology of benign cases (n=31) and malignant cases (n=31) of spirocercosis. Dogs with spirocercosis-induced sarcoma were significantly older (6.4+/-1.91 years) than benign cases (4.93+/-2.87). In the malignant cases there were significantly (p=0.03) more sterilized females (10/31) and fewer intact males (4/31) compared to 2/31 and 13/31, respectively, in the benign cases. Hypertrophic osteopathy was observed in 38.7% of malignant cases and in none of the benign cases (p=0.0002). Common clinical signs included weight loss, regurgitation, anorexia, pyrexia (T>or=39.5 degrees ), respiratory complications and salivation but did not differ in prevalence between groups. On haematology, the malignant group had significantly (p<0.05) lower haematocrit (0.34+/-0.08 vs. 0.41+/-0.07) and higher white cell count (31.6+/-27.83 vs. 17.71+/-13.18 x 10(3)microl(-1)), mature neutrophil count (26.06+/-26.08 vs. 12.23+/-9.96 x 10(3)microl(-1)) and thrombocyte count (493.15+/-151.61 vs. 313.27+/-128.54 x 10(9)microl(-1)). There were no differences in the mean corpuscular volume and immature neutrophil count. On radiology, the mass length was not significantly different, but the height and the width of the malignant masses were significantly larger (62.59+/-15.15 mm and 73.93+/-20.94 mm) compared to the benign group (46.43+/-23.62 and 49.29+/-25.56, respectively). Spondylitis was more prevalent in the malignant group (67.86% vs. 38.46%, p=0.03). Examining secondary pulmonary changes revealed significantly higher prevalence of bronchial displacement in the malignant group (52% vs. 17%, p=0.008). Hypertrophic osteopathy appeared to be a very specific but relatively rare (poor sensitivity) marker of malignancy. Female gender, anaemia, leukocytosis, thrombocytosis, spondylitis and bronchial displacement are significantly more common in malignant cases, but appear in benign cases as well. However, if found together in a specific case, they should increase the index of suspicion for malignancy in a diagnosed spirocercosis case.

Proposed histological progression of the Spirocerca lupi-induced oesophageal lesion in dogs.

This study aims to outline the histological progression of the Spirocerca lupi nodule from infection to neoplastic transformation. Sixty-two spirocercosis-induced nodules, 42 non-neoplastic and 20 neoplastic, were stained with HE. Ten non-overlapping high power fields per nodule were examined and non-neoplastic and neoplastic nodules were compared. Inflammation was scored 0-3 and revealed a score of 1.91+/-0.52 in the non-neoplastic and 0.97+/-0.5 in the neoplastic cases (p<0.01). In most non-neoplastic cases the inflammatory infiltrate was lymphoplasmacytic and in the neoplastic cases neutrophils predominated. Necrosis was scored 0-3 and revealed a score of 0.88+/-0.41 in the non-neoplastic and 1.47+/-0.5 in the neoplastic cases (p<0.01). The average number of mitoses over 10 high power fields per nodule was 1.31+/-1.55 in the non-neoplastic compared to 42.85+/-30.79 in the neoplastic cases (p<0.01). The average number of multinucleated giant cells over 10 high power fields per nodule was 0.9+/-1.45 in the non-neoplastic compared to 13.9+/-14.66 in the neoplastic cases (p<0.01). In the non-neoplastic cases, collagen, immature fibroblasts and fibroblast activation (excessively plump euchromatic nuclei with single or multiple prominent nucleoli) were scored 0-3 and a combined score, fibroblasts+activation score-collagen was calculated. The non-neoplastic cases were divided into a combined score of <or=1 (n=15) or >1 (n=27). The 2 groups had similar scores for inflammation and necrosis, but were significantly different (p<0.01) in mitotic index (0.26+/-0.46 vs. 1.89+/-1.65) and number of multinucleated cells (0 vs. 1.4+/-1.6). These results indicate 2 stages in the non-neoplastic nodules: early inflammation, characterized by fibrocytes and abundant collagen, and a pre-neoplastic stage, characterized by activated fibroblasts and reduced collagen.

A Novel Phospholipid Derivative of Indomethacin, DP-155 [Mixture of 1-Steroyl and 1-Palmitoyl-2-{4-[1-(p-chlorobenzoyl)-5-methoxy-2-methyl-3-indolyl acetamido]butanoyl}-sn-glycero-3-phosophatidyl Choline], Shows Superior Safety and Similar Efficacy in Reducing Brain Amyloid beta in an Alzheimer's Disease Model

Indomethacin has been suggested for the treatment of Alzheimers disease (AD), but its use is limited by gastrointestinal and renal toxicity. To overcome this limitation, D-Pharm Ltd. (Rehovot, Israel) developed DP-155 (mixture of 1-steroyl and 1-palmitoyl-2-{4-[1-(p-chlorobenzoyl)-5-methoxy-2-methyl-3-indolyl acetamido] hexanoyl}-sn-glycero-3-phosophatidyl choline), a lecithin derivative of indomethacin. Safety was tested by daily oral administration of DP-155 or indomethacin to rats in a dose range of 0.007 to 0.28 mmol/kg. The prevalence of gastrointestinal ulceration was significantly lower (10-fold) for DP-155 than for indomethacin, and the ulcerations were delayed. Signs of renal toxicity, namely reduced urine output and increased urine N-acetyl glycosaminidase to creatinine ratio, were 5-fold lower for DP-155. Indomethacin, but not an equimolar dose of DP-155, reduced urine bicyclo-prostaglandin E2. An equimolar oral dose of DP-155 or indomethacin, administered every 4 h for 3 days, was equally efficacious in reducing the levels of Aβ42 in the brains of Tg2576 mice. Indomethacin was the principal metabolite of DP-155 in the serum. After DP-155 oral administration, indomethacins half-life in the serum and the brain was 22 and 93 h, respectively, compared with 10 and 24 h following indomethacin oral administration. The brain to serum ratio was 3.5 times higher for DP-155 than indomethacin. This finding explains the efficacy of DP-155 in reducing Aβ42 brain levels, despite the low systemic blood concentrations of indomethacin derived from DP-155. In conclusion, compared with indomethacin, DP-155 has significantly lower toxicity in the gut and kidney while maintaining similar efficacy to indomethacin in lowering Aβ42 in the brains of Tg2576 mice. This superior safety profile highlights DP-155s potential as an improved indomethacin-based therapy for AD.

Electrocardiographic changes and cardiac pathology in canine babesiosis.

The main purpose of this study was to describe electrocardiographic (ECG) changes in canine babesiosis, and to relate these to clinical severity, outcome and cardiac pathology. Four groups of dogs with babesiosis were studied: mild to moderate anemia, severe anemia, concurrent autoagglutination and concurrent complications. Lead II ECG was recorded at admission for 1 minute in all dogs (121). A six lead ECG was recorded in 88 dogs. Full necropsy was performed on 16 dogs (5 died on arrival, 11 had ECG recording). The following ECG changes were recorded in relatively high prevalence: sinoatrial blocks or sinus arrest (7%), ventricular premature complexes (7%), low R-amplitude (23%), prominent Q (13%), axis deviations (40%), prolonged QRS (32%), ST depression and coving (28%), large T (42%), and notched R (28%). Differences between groups were minor. There was a significantly higher prevalence of sinus bradycardia and irregular rhythm in the non-survivors. Gross pathological changes were pericardial effusion and hemorrhages. Histological changes were hemorrhages, necrosis, inflammation and fibrin microthrombi. The only correlation between pathology and ECG was low R-amplitude and pericardial effusion. The ECG changes were similar to the pattern described for myocarditis and myocardial ischemia, and together with the histopathological findings indicated that the heart suffers from the same pathological processes described in other organs in canine babesiosis, namely inflammation and hypoxia. As the clinical application of the ECG changes found in this study was limited, cardiovascular assessment should be based on functional monitoring rather than ECG.

Serum C‐Reactive Protein Concentration in Benign and Malignant Canine Spirocercosis

BACKGROUND Spirocerca lupi is a nematode of canids that forms a nodule in the esophagus that can undergo neoplastic transformation. C-reactive protein (CRP) is a major acute phase protein in the dog that has been used for treatment, monitoring, and prognostication in inflammatory and neoplastic disease. HYPOTHESIS/OBJECTIVES The objectives of this study were to determine if serum CRP concentration (1) is increased in canine spirocercosis, (2) can be used to determine neoplastic transformation, and (3) can be used to monitor response to treatment in benign spirocercosis. ANIMALS Forty-two dogs naturally infected with S. lupi and 21 control dogs. METHODS A retrospective study was performed. The infected cases were divided into benign (n = 28) or malignant (n = 14) spirocercosis. CRP was performed on all of the spirocercosis and control cases at presentation. Statistical analysis was done by the one-way analysis of variance and Students t-test. RESULTS The mean CRP concentration in the benign cases was 60.4 ± 48.0 mg/L and that of the malignant cases was 76.5 ± 44.8 mg/L; both values were significantly higher (P < .001) than those of the control group where the mean was 13.4 ± 17.9 mg/L. The mean CRP concentration for the convalescent sera in the benign group was lower than the pretreatment concentrations (P= .01). CONCLUSION AND CLINICAL IMPORTANCE CRP cannot be used to differentiate between benign and malignant spirocercosis. There is a decrease in CRP concentration in dogs with benign spirocercosis once treatment has commenced. Serial CRP measurement can be used to monitor response to treatment in benign spirocercosis.

Plasma IL-8 concentrations are increased in dogs with spirocercosis

The nematode Spirocerca lupi (S. lupi) induces sarcoma in the dog oesophagus in about 25% of cases. The aim of this study was to compare the differences in the cytokine milieu between dogs with neoplastic (n=29) and non-neoplastic disease (n=49) and age- and gender-matched healthy controls (n=25). We measured IL-2, IL-4, IL-6, IL-8, IL-10, IL-18, GM-CSF and MCP-1 in a specific canine multiplex immunoassay kit. Cytokine concentrations were compared between the different groups using the Kruskal-Wallis test followed by Dunns test. Only IL-8 and IL-18 showed significant differences in their plasma concentration among the three groups. Kruskal-Wallis test revealed a significant (p=0.001) difference in IL-8 concentration between the neoplastic group (634pg/ml), the non-neoplastic (429 pg/ml) and the control groups (150 pg/ml). Post-test analysis revealed a significance difference between the two S. lupi groups and the control group (p<0.01). The highest IL-18 concentration was found in the non-neoplastic group (53 pg/ml), followed by the control group (46 pg/ml) and finally the neoplastic group (33 pg/ml). IL-18 concentrations were significantly higher in the non-neoplastic group than in the neoplastic group (p=0.05). The increased IL-8 in the spirocercosis groups is consistent with the neutrophilic infiltrate in spirocercosis lesions and in those of other inflammatory-induced neoplasias such as Barrets oesophagus and Helicobacter gastritis. IL-18 showed negative regulatory effect in several worm infections and it is possible that it plays the same role in spirocercosis, allowing the worm to evade the host response and to induce neoplastic transformation.

NT-ProBNP and cardiac troponin I in virulent canine babesiosis

Although cardiac pathology and consequently elevated serum cardiac troponin I (cTnI) have been reported, clinically it remains difficult to diagnose cardiac involvement in canine babesiosis. Thus the use of cardiac biomarkers would be useful in determining if a dog with babesiosis also has concurrent cardiac dysfunction. The objectives of this study were to determine plasma N terminal brain natriuretic peptide (NT-proBNP) in canine babesiosis and if it is correlated with cTnI. Three groups of dogs with babesiosis were used: mild uncomplicated (Group 1), severe uncomplicated (Group 2), and complicated (Group 3), and a control group (Group 4) with 15 dogs per group. Each animal had the following determined: serum urea and creatinine, urea: creatinine ratio, cystatin-C, cTnI, blood lactate, plasma NT-proBNP, fractional shortening (FS), and blood pressure. The median NT-proBNP value in Groups 1-4 was 246, 650, 638, and 106 pmol/l. All 3 babesiosis groups had a statistically elevated NT-proBNP level compared to the control group and Groups 2 and 3 showed significantly higher values compared to Group 1. Median cTnI in Group 1-3 was 0.39, 0.4, and 1.45 ng/ml, respectively with the control group having concentrations below the detection limit (0.2 ng/ml). There was a significant difference in cTnI concentration between the control group and group 3 but no statistical difference between the other babesiosis groups. The study concluded that dogs with babesiosis showed elevated levels NT-proBNP and the more severe the disease process the greater the elevation. This elevation is earlier or independent of the increased cTnI.

COMPUTED TOMOGRAPHIC AND RADIOGRAPHIC CHARACTERISTICS OF AORTIC LESIONS IN 42 DOGS WITH SPIROCERCOSIS

Spirocerca lupi is a common cause of vomiting, regurgitation, and sudden death in dogs that live in tropical or subtropical regions. Sudden death due to aortic rupture may occur with no preceding clinical signs. The purpose of this prospective study was to compare radiographic and computed tomographic (CT) characteristics of aortic lesions in a cohort of 42 dogs with endoscopically confirmed spirocercosis. Dorsoventral and right lateral recumbent thoracic radiographic findings were compared with pre- and postcontrast thoracic CT findings. Aortic mineralization was detected using CT in 18/42 dogs (43%). Three dogs had faint diffuse aortic wall mineralization. Using CT as the reference standard, radiographs had a sensitivity and specificity of 6% and 96%, respectively, for detecting aortic mineralization. A total of 20 aortic aneurysms were detected using CT in 15/42 dogs (36%). Using CT as the reference standard, radiographs had a sensitivity and specificity of 86% and 56%, respectively, for detecting aortic aneurysms. Respiratory motion, aortic displacement by esophageal masses and Spirocerca nodules adjacent to the aorta mimicked aneurysm formation on radiographs. Aortic thrombi were seen in two dogs in postcontrast CT images. Findings from this study indicated that aortic mineralization and aneurysm formation are common in dogs with spirocercosis. Findings also supported the use of pre- and postcontrast CT as effective methods for detecting and characterizing these lesions.

Serum alkaline phosphatase activity is not a marker for neoplastic transformation of esophageal nodules in canine spirocercosis.

BACKGROUND Spirocerca lupi is a nematode of Canidae that matures within the esophageal wall to form fibroblastic nodules with potential for malignant transformation. Diagnosis is based on histopathologic examination, but false-negative results may be obtained from samples collected by endoscopy. Serum alkaline phosphatase (ALP) activity, frequently increased in hepatobiliary disease, is also increased in a variety of neoplastic conditions in dogs, including appendicular osteosarcoma, and has also been reported to be increased in dogs with spirocercosis. OBJECTIVE The aim of this study was to evaluate serum ALP activity as a marker for malignant transformation of esophageal nodules in S. lupi-infected dogs. METHODS In this retrospective study, medical records of dogs diagnosed with spirocercosis from 1991 to 2008 were reviewed, and serum ALP activity determined at presentation was compared between dogs with nonneoplastic and neoplastic nodules. Owing to use of multiple analyzers, ratios of ALP activity to the upper reference interval for ALP were calculated and compared. RESULTS Median ALP activity ratios were 0.65 (0.07-4.00) and 0.86 (0.10-3.40) for dogs with nonneoplastic (n=88) and neoplastic (n=32) nodules, respectively, with no significant difference (P=.18) and substantial overlap between groups. Tumors included osteosarcoma (15 dogs), fibrosarcoma (15 dogs), and anaplastic sarcoma (2 dogs); there was no difference in ALP activity between the dogs with osteosarcoma and fibrosarcoma. CONCLUSION ALP is a poor marker of malignant transformation in canine spirocercosis.

Immunohistochemical characterization of lymphocyte and myeloid cell infiltrates in spirocercosis‐induced oesophageal nodules

Spirocerca lupi is a nematode that infects the dog’s oesophagus and promotes the formation of an inflammatory fibroblastic nodule that progresses to sarcoma in approximately 25% of cases. Spirocercosis‐associated oesophageal sarcoma is an excellent and under‐utilized spontaneous model of parasite‐associated malignancy. The inflammatory infiltrate of paraffin‐embedded, non‐neoplastic oesophageal nodules (n = 46), neoplastic nodules (n = 25) and normal oesophagus (n = 14) was examined by immunohistochemistry using MAC387 (myeloid cells), CD3 (T cells), Pax5 (B cells) and FoxP3 (T regulatory cells) antibodies. Myeloid cells predominated in 70% of nodules, in pockets around the worms’ migratory tracts and in necro‐ulcerative areas in neoplastic cases. T cells predominated in 23% of cases with a focal or diffuse distribution, in the nodule periphery. No significant differences were observed between neoplastic and non‐neoplastic stages. FoxP3+ cells were observed in low numbers, not significantly different from the controls. The inflammation in spirocercosis is characterized by pockets of pus surrounded by organized lymphoid foci. There was no evidence of a local accumulation of FoxP3+ cells, unlike many previous studies that have reported an increase in FoxP3+ T cells in both malignancies and parasite infections. The triggering factor(s) driving the malignant transformation of the spirocercosis‐associated chronic inflammatory nodule warrants further investigation.