Karen E. Russell

Cerebrospinal fluid myelin basic protein as a prognostic biomarker in dogs with thoracolumbar intervertebral disk herniation.

BACKGROUND Release of myelin basic protein (MBP) into the cerebrospinal fluid (CSF) is associated with active demyelination and correlates with outcome in various neurological diseases. HYPOTHESIS/OBJECTIVES To describe associations among CSF MBP concentration, initial neurological dysfunction, and long-term ambulatory outcome in dogs with acute thoracolumbar intervertebral disk herniation (IVDH). ANIMALS Fifty seven [corrected] dogs with acute thoracolumbar IVDH and 16 clinically normal dogs. METHODS Prospective case series clinical study. Signalment, initial neurological dysfunction as determined by a modified Frankel score (MFS), and ambulatory outcome at >3-month follow-up were recorded. Cisternal CSF MBP concentration was determined by an ELISA. Associations were estimated between CSF MBP concentration and various clinical parameters. RESULTS Dogs with thoracolumbar IVDH that did not ambulate at follow-up had a higher CSF MBP concentration (median, 3.56 ng/mL; range, 0.59-51.2 ng/mL) compared with control dogs (median, 2.22 ng/mL; range, 0-3.82 ng/mL) (P=.032). A CSF MBP concentration of >or=3 ng/mL had a sensitivity of 78% and specificity of 76% to predict an unsuccessful outcome based on receiver-operating characteristics curve analysis (area under the curve=0.688, P=.079). Affected dogs with a CSF MBP concentration>or=3 ng/mL had 0.09 times the odds of ambulation at follow-up compared with affected dogs with CSF MBP concentration<3 ng/mL when adjusted for initial MFS (95% confidence interval 0.01-0.66, P=.018). CONCLUSIONS AND CLINICAL IMPORTANCE These results would suggest that CSF MBP concentration may be useful as an independent prognostic indicator in dogs with thoracolumbar IVDH.

Primary and secondary hemostatic functionalities of rehydrated, lyophilized platelets

BACKGROUND: The rehydrated, lyophilized (RL) platelet (PLT) is being developed as a hemostatic infusion agent for the control of active bleeding. The key to the method for preparing RL PLTs is a mild aldehyde stabilization that allows for freezing and lyophilizing without cellular rupture. RL PLTs have been shown to be effective at rapidly controlling bleeding in animal models of cardiopulmonary bypass induced PLT dysfunction and washout thrombocytopenia, yet the rehydrated cells have proved to be safe with respect to induction of pathologic intravascular coagulation.

Thrombus Formation with Rehydrated, Lyophilized Platelets

Abstract Stored human platelets are frequently used in hemorrhagic emergencies, but have limited immediate utility for controlling bleeding due to storage lesion and are frequently contaminated with microorganisms. The development of paraformaldehyde-treated, lyophilized and rehydrated (RL) platelets, which are sterile and have a prolonged shelf life (years), ameliorate the efficacy and sterility problems with stored platelets. RL platelets have been shown to have many native functions of fresh platelets in vitro and to mediate hemostasis in vivo in large animal models of hemorrhagic shock and cardiopulmonary bypass induced platelet dysfunction. To further evaluate the functional properties of this transfusion product, we studied the role of RL platelets in three aspects of thrombus formation and lysis. First, the interaction between RL platelets and fibrinogen was investigated. The surface density of unligated GPIIb-IIIa on RL and fresh platelets were, respectively 30,000 and 70,000 molecules per cell as detected with the monoclonal antibody 10E-5. Freezing, lyophilization and rehydration steps in the preparation of RL platelets resulted in the surface presentation of 120,000 molecules of fibrinogen per cell from alpha granule sources. After ADP activation, RL platelets bound exogenous 125I-labeled fibrinogen in a dose-dependent manner with an affinity that is similar to that of fresh platelets and was inhibited by RGD peptides. 125I-Labeled fibrinogen binding to RL and fresh platelets, respectively, saturated at 14,000 and 32,000 molecules per cell. Scanning electron microscopic ultrastructural analysis showed that fibrin strands interacted with the surface of RL platelets in a normal manner. The second set of studies investigated the ability of RL platelets to catalyze and amplify the clot formation process in an activation-dependent manner. We showed that RL platelets undergo degranulation in fibrin in clots and functioned as thrombogenic surfaces for the generation of activated coagulation factors and fibrin generation. A final set of studies was performed to investigate fibrin of clots that contained RL platelets. RL platelet clots were lysed in the presence of tissue plasminogen activator with a similar time course as clots without platelets, and lysis occurred faster than when fresh platelets were included in the fibrin mass. The results of these three studies demonstrate that RL platelets are capable of mediating thrombus formation and do not inhibit lysis. Our results help explain how RL platelets restore hemostasis in vivo, and indicate that these cells might be a viable alternative to fresh stored platelets in transfusion medicine.

Associations between cerebrospinal fluid biomarkers and long-term neurologic outcome in dogs with acute intervertebral disk herniation

OBJECTIVE To examine associations between CSF biomarkers, initial neurologic dysfunction, and long-term ambulatory outcome in dogs with acute intervertebral disk herniation (IVDH). DESIGN Prospective clinical study. ANIMALS 54 dogs with acute thoracolumbar IVDH and 16 clinically normal dogs. PROCEDURES For each dog, variables, including CSF myelin basic protein (MBP), lactate, calcium, glucose, and total protein concentrations; nucleated cell count; and creatine kinase (CK) and aspartate aminotransferase activities, were measured. For dogs with thoracolumbar IVDH, initial neurologic function was characterized by use of a modified Frankel score (MFS; determined on a scale of 0 to 5, where 0 represented paraplegia with no deep nociception and 5 represented paraspinal hyperesthesia only). Long-term follow-up was assessed. RESULTS Among dogs with thoracolumbar IVDH, those with CSF CK activity ≤ 38 U/L had a 35-fold increase in the odds of long-term ambulation, compared with the odds in dogs with CSF CK activity > 38 U/L, adjusting for neurologic functioning at the evaluation. The CSF lactate, calcium, and glucose concentrations and aspartate aminotransferase activity were not predictive of long-term ambulatory outcome. Data analysis revealed that long-term functional recovery was > 98% for affected dogs, regardless of their initial MFS, when CSF CK activity was ≤ 38 U/L and MBP concentration was ≤ 3 ng/mL. CONCLUSIONS AND CLINICAL RELEVANCE In dogs with acute thoracolumbar IVDH, CSF CK activity and MBP concentration appeared to be prognostic indicators and, along with initial MFS, can be used to predict long-term ambulatory outcome.

Altered inflammatory responses following transforming growth factor-β neutralization in experimental guinea pig tuberculous pleurisy

The predominant extrapulmonary form of tuberculosis, which develops in 10% of diseased individuals, is pleurisy. The immune response mounted against Mycobacterium tuberculosis in the pleural cavity is one that is sufficient for clearing the organism without therapeutic intervention. Thus, examining the role of immune constituents in this context will provide understanding of the vital role they play in controlling tuberculosis. In this study, experimental tuberculous pleurisy was induced in guinea pigs, and anti-TGF-beta was administered intrapleurally to the guinea pigs daily throughout the study (8 days). Neutralizing TGF-beta resulted in a significant reduction in the percentage of lymphocytes and CD8+ cells present in the pleural exudate, decreased proliferative responses of pleural cells to ConA and PPD, and decreased mRNA expression of IFN-gamma and CCL5 in pleural effusion cells. Conversely, the percentage of neutrophils was significantly increased in anti-TGF-beta-treated guinea pigs, along with upregulated mRNA expression of CXCL8. The percentage of macrophages in the pleural exudate, TNF-alpha and IL-12p40 mRNA expression, and the histopathological response were not significantly altered. While TGF-beta is generally thought of as an immunosuppressive cytokine, the results of this study demonstrate its importance in promoting an inflammatory response, and highlight its bipolar nature.

Impact of Supplements on Iron Absorption From Diets Containing High and Low Iron Concentrations in the European Starling (Sturnus vulgaris)

Abstract Iron absorption was compared in European starlings (Sturnus vulgaris) fed diets containing high iron (1585 ppm), high iron (1720 ppm) with a phytate (inosital) and tannic acid, low iron (32–34 ppm), low iron with a meat-based dog food, or low iron with vitamin C. Iron absorption was determined by colorimetric analysis of nonheme liver iron. A dietary iron concentration of 32 ppm was inadequate to meet the physiologic demands of the starlings, even when vitamin C or a meat-based dog food was added. The high-iron diet caused an increase in concentrations of nonheme liver iron to levels similar to those in birds that have died of iron storage disease (ISD). Nonheme liver iron concentrations did not increase when a phytate (inositol) and tannic acid were added the high-iron diet. The optimal dietary iron concentration for starlings was narrowed to the range of 34–125 ppm. Adding a phytate, tannic acid, or both to readily available food stuffs may be a practical alternative to more expensive low-iron diets. Furthermore, a low-iron (32 ppm) diet may be an important adjunct to the treatment of iron storage disease in birds.

Surgical removal of an intramedullary spinal cord foreign body granuloma in a dog

A 2-year-old, spayed female, mixed-breed dog was presented for evaluation of a progressive asymmetric tetraparesis and cranial nerve deficits with a 3-week duration. Computed tomography showed a contrast-enhancing lesion along the left side of the junction of the medulla and the cervical spinal cord. An exploratory surgery determined the presence of an intramedullary lesion of the first cervical spinal cord segment. The mass was removed through a dorsal midline myelotomy. Microscopic examination identified a foreign body granuloma that contained a dense, anisotropic outer wall, supporting the conclusion that the foreign body was of plant origin. The dog recovered to a more improved ambulatory status than prior to surgery.

COMPARISON OF TWO ANALYZERS TO DETERMINE SELECTED VENOUS BLOOD ANALYTES OF QUAKER PARROTS (MYIOPSITTA MONACHUS)

Abstract: Point of care devices can assess electrolyte, blood gas, biochemical, and hematologic values in a critical care setting. Although these devices are commonly used in humans and companion mammals, few studies have assessed their use in avian species. This study compares electrolyte, hemoglobin (Hgb), hematocrit (Hct), acid-base, and venous blood gas parameters between the i-STAT® and IRMA TruPoint® blood gas analysis systems for 35 Quaker parrots. Agreement between the two analyzers and the effect of gender, time lag between sample analysis, and cartridge expiration were evaluated. Male birds had increased Hgb and Hct compared with females, independent of analyzer method. In expired i-STAT cartridges, only glucose significantly increased. Packed cell volume determined by centrifugation was higher than Hct, as calculated by either analyzer. The analyzers had good agreement for total carbon dioxide, bicarbonate, pH, and Hgb, fair agreement for potassium (K), ionized calcium (iCa), venous partial pressure of carbon dioxide, and base excess, and poor agreement for sodium (Na), venous partial pressure of oxygen (PO2), and oxygen saturation (SO2). Values for Na, iCa, PO2, and SO2 were significantly higher on the IRMA than the i-STAT, while K was significantly lower on the IRMA when compared with the i-STAT. The time lag between sample analyses on the i-STAT and IRMA did not be correlate to any analyte changes. Despite these differences, both the i-STAT and the IRMA appear to be acceptable clinical tools in avian critical care, although reference ranges for each analyzer should be created.

A Comparison of Four Regimens for Treatment of Iron Storage Disease Using the European Starling (Sturnus vulgaris) as a Model

Abstract European starlings (Sturnus vulgaris) were fed an iron loading diet (3235 ppm) for 31 days to induce nonheme liver iron concentrations approaching those in birds that died with iron storage disease. All birds then were fed a low-iron diet (32–48 ppm) and assigned to 4 treatment groups: 1) low-iron diet only, 2) low-iron diet with phytate (inositol) and tannic acid, 3) low-iron diet and deferoxamine (100 mg/kg SC q24h), and 4) low-iron diet and phlebotomy (1% of body weight q7d). Starlings were treated for 16 weeks. In the groups treated with phlebotomy or with deferoxamine and a low-iron diet, nonheme liver iron concentrations decreased to safe levels after 16 weeks of treatment at similar rates (190 ppm/wk and 163 ppm/wk, respectively). The low-iron diet alone reduced stored liver iron levels at a slower rate (45 ppm/wk). The addition of inositol and tannic acid to the low-iron diet had no impact on stored liver iron concentrations. These results suggest that both phlebotomy and treatment with deferoxamine are effective treatment options for birds with iron storage disease.

Gene expression of innate Th1-, Th2-, and Th17-type cytokines during early life of neonatal foals in response to Rhodococcus equi

Focusing on the first 3 weeks of life, this study examined the mRNA transcript development of different Th-type cytokines in foals in response to Rhodococcus equi infection in vitro. Results demonstrated the significant up-regulation in expression of Th1-, Th2-, and Th17-type cytokines (IFN-γ, IL-4, IL-6, IL-8, IL-12p35, IL-12p40, IL-17, IL-23p19, and TNF-α) in R. equi infection of bronchial alveolar lavage (BAL) cells of 10-day-old foals. Consequently, signature cytokines of 3 Th cell types, IFN-γ (Th1), IL-4 (Th2), and IL-17 (Th17), were used to compare temporal response patterns of circulating peripheral blood mononuclear cells (PBMCs) to stimulation with R. equi. Foals responded to R. equi stimulation by producing similar amounts of IFN-γ mRNA transcripts from birth through 3 weeks of age, suggesting an absence of age-related impairment in Th1-type cytokine response to R. equi during the first 3 weeks of life. It remains debatable whether this Th1 response to R. equi in foals≤3 weeks of age is generally immature relative to older foals or adult horses. IL-4 expression by R. equi-stimulated PBMCs was significantly decreased at birth, and IL-17 expression was relatively reduced during the first week of life. Among all cytokines studied, IL-17 mRNA transcripts were induced with the highest magnitude of fold-change both in BAL cells and in PBMCs. Under the conditions studied, in vivo administration of a CpG failed to modulate the Th1-, Th2-, and Th17-type cytokine expression patterns in PBMCs.