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Oxidized linoleic acid metabolites (OXLAM) are products of adipocyte lipolysis with the potential to modulate adipose tissue (AT) lipid metabolism and inflammation. In periparturient cows, linoleic acid is preferentially mobilized from AT during lipolysis by hormone-sensitive lipase (HSL) compared with other polyunsaturated fatty acids. Enzymatic and nonenzymatic reactions generate OXLAM from linoleic acid. Among OXLAM, 9-, 10-, and 12-hydroxy-octadecadienoic acids (HODE) are associated with pro-inflammatory responses, whereas 9- and 13-oxo-octadecadienoic acids (oxoODE) and 13-HODE can facilitate inflammation resolution and promote lipogenesis. This study evaluated the effect of HSL activity on OXLAM biosynthesis using subcutaneous AT explants collected from multiparous dairy cows at 10 d before and again at 10 and 24 d after calving. Explants were treated for 3 h without or with the β-adrenergic agonist isoproterenol (ISO; 1 µM; MilliporeSigma, Burlington, MA) to induce HSL activity. The contribution of HSL to OXLAM biosynthesis was determined by inhibiting its activity with CAY10499 (2 µM; Cayman Chemical, Ann Arbor, MI). After treatments, media and explants were collected for lipidomic analysis using HPLC-tandem mass spectroscopy. Results indicated that ISO increased the biosynthesis of 9-, 12-, and 13-HODE and 9-oxoODE, and this effect was reduced at 24 d after calving. Inhibiting HSL activity partially reversed ISO effects on HODE and 9-oxoODE. Our ex vivo model demonstrated for the first time a direct effect of HSL activity on the biosynthesis of OXLAM in AT, especially at 10 d before and 10 d after calving. The biosynthesis of anti-inflammatory OXLAM is limited during the first weeks after parturition and may promote AT inflammation and lipolytic responses to negative energy balance. These results indicate that HSL activity releases linoleic acid for OXLAM biosynthesis in concentrations of a magnitude that may bypass the need for the activation of phospholipases linked with the inflammatory cascade and thus supports, in part, lipolysis-driven inflammation within AT of periparturient cows.

A 10-y-old giraffe ( Giraffa camelopardalis reticulata) bull developed colic after a 3-mo history of reduced feed consumption. Physical examination and management were performed with 2 standing sedations. The giraffe developed metabolic alkalosis and progressive pre-renal azotemia followed by compensatory respiratory acidosis and paradoxical aciduria. A metallic “ping” sound was auscultated on the left side near ribs 10–12. The giraffe was euthanized given the grave prognosis, and postmortem examination confirmed left displacement of the abomasum (LDA) with fluid sequestration (150–190 L [40–50 gal]) within the rumen. Dental disease was evident at postmortem examination and perimortem skull computed tomography. To ensure cases of LDA are not overlooked, the position of the abomasum must be noted during postmortem examination prior to removal of the gastrointestinal tract. The risk factors for the development of LDA in giraffes are not known, and associations such as those of dairy cattle (hypocalcemia, high-concentrate low-fiber diet, and indoor housing) remain to be elucidated.

Bovine coliform mastitis presents treatment challenges because of systemic inflammation and oxidative stress. Soluble epoxide hydrolase (sEH) is a promising therapeutic target in conditions characterized by inflammation and oxidative stress but has not been evaluated in cattle. We compared sEH activity and oxidant status in healthy Holstein dairy cows to those with systemic coliform mastitis (n = 5/group) using complementary approaches. First, the activity of sEH on [3H]-trans-diphenyl-propene oxide (tDPPO) was assessed ex vivo using tissue homogenates (mammary, liver, and kidney). Second, the concentrations of sEH substrates and metabolites in plasma, milk, and urine were determined as an index of in vivo sEH activity. Oxidant status was assessed in serum and milk. Data were analyzed by non-parametric methods. Metabolism of tDPPO was greater in mammary tissues from cows with coliform mastitis compared to controls. In contrast, ratios of sEH substrates and metabolites predicted lower sEH activity in cows with coliform mastitis than controls. Milk oxidant status showed greater prooxidant levels in coliform mastitis cows. Cows with coliform mastitis exhibit increased sEH activity in mammary tissue, at the same time, milk oxidant status is increased. Future studies should characterize sEH activity and oxidant status patterns and explore therapies targeting sEH during coliform mastitis.

The veterinary pharmacopeia available to treat pain and inflammation is limited in number, target of action and efficacy. Inhibitors of soluble epoxide hydrolase (sEH) are a new class of anti-inflammatory, pro-resolving and analgesic drugs being tested in humans that have demonstrated efficacy in laboratory animals. They block the hydrolysis, and thus, increase endogenous concentrations of analgesic and anti-inflammatory signaling molecules called epoxy-fatty acids. Here, we screened a library of 2,300 inhibitors of the sEH human against partially purified feline, canine and equine hepatic sEH to identify inhibitors that are broadly potent among species. Six very potent sEH inhibitors (IC50 < 1 nM for each enzyme tested) were identified. Their microsomal stability was then measured in hepatic extracts from cat, dog and horse, as well as their solubility in solvents suitable for the formulation of drugs. The trans-4-{4-[3-(4-trifluoromethoxy-phenyl)-ureido]-cyclohexyloxy}-benzoic acid (t-TUCB, 1,728) appears to be the best compromise between stability and potency across species. Thus, it was selected for further testing in veterinary clinical trials of pain and inflammation in animals.

Oxidative stress contributes to dysfunctional immune responses and predisposes dairy cattle to several metabolic and inflammatory-based diseases. Although the negative effects of oxidative stress on transition cattle are well established, biomarkers that accurately measure oxidative damage to cellular macromolecules are not well defined in veterinary medicine. Measuring 15-F2t-isoprostane, a lipid peroxidation product, is the gold standard biomarker for quantifying oxidative stress in human medicine. The aim of our study was to determine whether changes in 15-F2t-isoprostane concentrations in plasma and milk could accurately reflect changes in oxidant status during different stages of lactation. Using liquid chromatography-tandem mass spectrometry, 15-F2t-isoprostane concentrations were quantified in milk and plasma of 12 multiparous Holstein-Friesian cows that were assigned to 3 different sampling periods, including the periparturient period (1-2 d in milk; n = 4), mid lactation (80-84 d in milk; n = 4), and late lactation (183-215 d in milk; n = 4). Blood samples also were analyzed for indicators of oxidant status, inflammation, and negative energy balance. Our data revealed that 15-F2t-isoprostane concentrations changed at different stages of lactation and coincided with changes in other gauges of oxidant status in both plasma and milk. Interestingly, milk 15-F2t-isoprostane concentrations and other indices of oxidant status did not follow the same trends as plasma values at each stage of lactation. Indeed, during the periparturient period, systemic 15-F2t-isoprostane increased significantly accompanied by an increase in the systemic oxidant status index. Milk 15-F2t-isoprostane was significantly decreased during the periparturient period compared with other lactation stages in conjunction with a milk oxidant status index that trended lower during this period. The results from this study indicate that changes in 15-F2t-isoprostane concentrations in both milk and plasma may be strong indicators of an alteration in redox status both systemically and within the mammary gland.

Periparturient dairy cows experience an increased incidence and severity of several inflammatory-based diseases such as mastitis and metritis. Factors associated with the physiological adaptation to the onset of lactation can impact the efficiency of the inflammatory response at a time when it is most needed to eliminate infectious pathogens that cause these economically important diseases. Oxidative stress, for example, occurs when there is an imbalance between the production of oxygen radicals during periods of high metabolic demand and the reduced capabilities of the host's antioxidant defenses. The progressive development of oxidative stress in early lactation cows is thought to be a significant underlying factor leading to dysfunctional inflammatory responses. Reactive oxygen species (ROS) are also produced by leukocytes during inflammation resulting in positive feedback loops that can further escalate oxidative stress during the periparturient period. During oxidative stress, ROS can modify polyunsaturated fatty acids (PUFA) associated with cellular membranes, resulting in the biosynthesis of oxidized products called oxylipids. Depending on the PUFA substrate and oxidation pathway, oxylipids have the capacity of either enhancing or resolving inflammation. In mediating their effects, oxylipids can directly or indirectly target sites of ROS production and thus control the degree of oxidative stress. This review discusses the evidence supporting the roles of oxylipids in the regulation of oxidative stress and the subsequent development of uncontrolled inflammatory responses. Further, the utility of some of the oxylipids as oxidative stress markers that can be exploited in developing and monitoring therapies for inflammatory-based diseases in dairy cattle is discussed. Understanding of the link between some oxylipids and the development or resolution of oxidative stress could provide novel therapeutic targets to limit immunopathology, reduce antibiotic usage, and optimize the resolution of inflammatory-based diseases in periparturient dairy cows.

Dairy cows experience increased oxidative stress during periods of transition such as at the cessation of lactation and around the periparturient period, thus increasing disease risk. Despite routine supplementation of transition cow diets with certain vitamins in an attempt to mitigate oxidative stress, there is no currently available data directly linking vitamin supplementation with antioxidant potential (AOP) in transition cows. The objective of this study was to determine the association between serum vitamins and biomarkers of oxidative stress in healthy cows. Blood samples were collected from 240 cows at dry off (DO), close up (CU), and 2–10 days post-calving (DIM2-10). Blood samples were analyzed for vitamins (A, D, E), β-carotene, reactive oxygen species (ROS), and AOP. Spearman correlations and mixed linear regression models were used to assess associations between vitamins and measures of oxidant status. Vitamin D concentrations were positively associated with AOP at the CU and DIM2-10. Based on the positive association with AOP, additional in-vitro studies were conducted that showed vitamin D mitigated barrier integrity loss in endothelial cells during oxidative stress. These results indicate for the first time that vitamin D may have a role in promoting antioxidant potential in transition dairy cows.

Unregulated inflammation during bovine mastitis is characterized by severe mammary tissue damage with systemic involvement. Vascular dysfunction underlies tissue pathology because of concurrent oxidative stress mediated by several inflammatory mediators. We recently demonstrated increased production of 20-hydroxyeicosatetraenoic acid (20-HETE), a cytochrome P450-derived (CYP) oxylipid that correlated with oxidative stress during severe bovine coliform mastitis. The hypothesis for this study was that 20-HETE-induced oxidative stress disrupts barrier function of endothelial cells. Primary endothelial cells from the bovine aorta were utilized to investigate the effects of 20-HETE on barrier integrity in an in-vitro model of oxidative stress. The effects of various antioxidants on modulating the 20-HETE barrier integrity effects also were investigated. Our results showed that 20-HETE decreased endothelial barrier integrity, which was associated with increased reactive metabolite production and decreased total glutathione. The antioxidant, vitamin E, partially delayed the loss of endothelial resistance upon exposure to 20-HETE but did not prevent complete loss of barrier integrity. The decrease in barrier resistance due to 20-HETE was neither associated with oxidative stress as assessed by oxidative protein or lipid damage nor endothelial cell apoptosis; however, selenium supplementation conferred resistance to loss of barrier integrity suggesting a role for shifts in redox status. Specific mechanisms by which 20-HETE alters vascular barrier integrity require further investigation to identify targets for therapy during inflammatory conditions with enhanced CYP450 activity.

Oxidative stress contributes to dysfunctional immune responses and predisposes dairy cattle to several metabolic and inflammatory-based diseases. Although the negative effects of oxidative stress on transition cattle are well established, biomarkers that accurately measure oxidative damage to cellular macromolecules are not well defined in veterinary medicine. Measuring 15-F

Tube cystostomy is a surgical method used for managing obstructive urolithiasis and involves placement of a Foley catheter into the urinary bladder. We identified and evaluated the antibiotic resistance patterns of bacteria isolated from indwelling Foley catheters following tube cystostomy in goats with obstructive urolithiasis. Urine samples collected over a 10-y period from catheter tips at the time of removal were submitted for bacteriologic culture and antibiotic susceptibility testing. Resistance patterns to antibiotics, trends in the resistance patterns over the study period, and the probability of a bacterial isolate being resistant as a function of the identity of the isolate and antibiotic tested were determined. A total of 103 urine samples from 103 male goats with obstructive urolithiasis managed surgically with tube cystostomy were included in the study. Aerococcus (36.9%) and Enterococcus (30.1%) were isolated most frequently. The susceptibility patterns of all bacteria isolated did not change over the study period ( p > 0.05). Proportions of isolates resistant to 1, 2, and ≥3 antibiotics were 36.9%, 18.5%, and 23.3%, respectively. Thus, 41.8% of bacterial isolates were resistant to 2 or more antibiotics tested. The probability of Aerococcus spp., Escherichia coli, and Pseudomonas aeruginosa isolates to be resistant to ampicillin, ceftiofur, erythromycin, penicillin, or tetracycline ranged from 0.59 to 0.76.

Objective—To determine whether dietary supplementation with ammonium chloride would affect urine pH or urinary fractional excretion (FE) of electrolytes in goats fed grass hay. Design—Clinical trial. Animals—15 yearling castrated male goats. Procedures—In the dose response study, 3 yearling goats fed orchard grass hay and water ad libitum were administered ammonium chloride at either 200, 400, or 500 mg/kg (91, 182, or 227 mg/lb), PO, every 24 hours. In the FE study, 8 goats fed orchard grass hay were randomly divided into either a treatment (n = 4) or a control group (4). In the treatment group, ammonium chloride was administered at 450 mg/kg (2.25% of dry matter intake [DMI]), PO, every 24 hours for 8 days. The FE of electrolytes was compared between groups; FE measurements were also determined for 4 client-owned goats fed alfalfa hay. Results—Ammonium chloride administered at 450 mg/kg (2.25% of DMI) achieved and maintained urine pH < 6.5 for 24 hours. Goats fed orchard grass hay with ammonium chloride supplementation had significantly higher FE of calcium and chloride than did goats fed orchard grass hay without supplemental ammonium chloride. Conclusions and Clinical Relevance—Dietary ammonium chloride supplementation at a dose of 450 mg/kg may be necessary to achieve a urine pH < 6.5 in goats. Further studies of ammonium chloride supplementation and urolithiasis in goats fed low-calcium diets are indicated.

The periparturient period of dairy cows is characterized by intense lipolysis in adipose tissues (AT), which induces the release of free fatty acids (FFA) into circulation. Among FFA, polyunsaturated fatty acids are susceptible to oxidation and can modulate inflammatory responses during lipolysis within AT. Linoleic and arachidonic acid oxidized products (oxylipids) such as hydroxy-octadecadienoic acids (HODE) and hydroxy-eicosatetraenoic acids (HETE), were recently identified as products of lipolysis that could modulate AT inflammation during lipolysis. However, the effect of lipolysis intensity during the transition from gestation to lactation on fatty acid substrate availability and subsequent AT oxylipid biosynthesis is currently unknown. We hypothesized that in periparturient dairy cows, alterations in AT and plasma fatty acids and oxylipid profiles coincide with changes in lipolysis intensity and stage of lactation. Blood and subcutaneous AT samples were collected from periparturient cows at -27±7 (G1) and -10±5 (G2) d prepartum and at 8±3 d postpartum (PP). Targeted lipidomic analysis was performed on plasma and AT using HPLC-MS/MS. We report that FFA concentrations increased as parturition approached and were highest at PP. Cows exhibiting high lipolysis rate at PP (FFA>1.0 mEq/L) had higher body condition scores at G1 compared to cows with low lipolysis rate (FFA

Objective—To determine associations between age, sex, breed, and month and year of admission and the diagnosis of lead toxicosis in cattle. Design—Retrospective case-control study. Sample Population—Records of all cattle evaluated at North American veterinary teaching hospitals during the years 1963 to 2002, which were available through the Veterinary Medical Database. Procedures—Logistic regression was used to evaluate the associations between postulated risk factors and the occurrence of lead toxicosis in cattle and predict the occurrence of the diagnosis of lead toxicosis in cattle. Results—413 cases of lead intoxication and 202,363 control cattle were identified and met the inclusion criteria. Cattle < 4 years of age were at increased risk for the diagnosis of lead intoxication relative to cattle ≥ 4 years of age. Cattle ≥ 2 months and < 6 months of age had the greatest risk for lead intoxication (odds ratio, 12.3). Angus cattle were at greater risk for toxicosis (odds ratio, 1.95), compared with other breeds. The risk of lead toxicosis was greater before 1985 (odds ratio, 1.94) than the risk thereafter. The risk of lead toxicosis diagnosis was greatest in the months of May, June, July, and August. Conclusions and Clinical Relevance—Lead toxicosis in cattle was associated with age < 4 years and the Angus breed. A seasonal pattern existed with peak occurrence in the late spring and summer. The occurrence of lead toxicosis has declined over time.

A case of malignant melanoma, originating at the base of the left horn, was diagnosed in a white 11-year-old Pygora doe. At necropsy, the goat had an ulcerated, black-pigmented, 2.5-cm mass at the base of the left horn. Firm masses diffusely black on the cut surface were present at the left horn base, subcutaneously over the frontal bones, and infiltrating into the frontal sinuses, the submandibular lymph node, and left parotid salivary gland. The left maxillary premolars and molars were loosened from the periodontia. Multiple black foci of metastasis were observed in the liver. Histologically, the masses consisted predominantly of moderately pleomorphic, polyhedral to spindle cells that contained variable amounts of dark brown intracytoplasmic pigment granules. The tumor cells were positive for Melan A by immunohistochemical staining.

Coliform mastitis is a severe and sometimes fatal disease characterized by an unregulated inflammatory response. The initiation, progression, and resolution of inflammatory responses are regulated, in part, by potent oxylipid metabolites derived from polyunsaturated fatty acids. The purpose of this study was to characterize the biosynthesis and diversity of oxylipid metabolites during acute bovine coliform mastitis. Eleven cows diagnosed with naturally occurring acute systemic coliform mastitis and 13 healthy control cows, matched for lactation number and days in milk, were selected for comparison of oxylipid and free fatty acid concentrations in both milk and plasma. Oxylipids and free fatty acids were quantified using liquid chromatography-tandem mass spectrometry. All polyunsaturated fatty acids quantified in milk were elevated during coliform mastitis with linoleic acid being the most abundant. Oxylipids synthesized through the lipoxygenase and cytochrome P450 pathways accounted for the majority of the oxylipid biosynthesis. This study demonstrated a complex and diverse oxylipid network, most pronounced at the level of the mammary gland. Substrate availability, biosynthetic pathways, and degree of metabolism influence the biosynthesis of oxylipids during bovine coliform mastitis. Further studies are required to identify targets for novel interventions that modulate oxylipid biosynthesis during coliform mastitis to optimize inflammation.

Case Description—6 lactating dairy goats were examined because of acute mastitis. Clinical Findings—Goats were considered to have endotoxemia on the basis of physical examination and clinicopathologic findings. The affected udder halves had gangrenous discolored distal portions with sharp demarcations from grossly normal tissue proximally. Udder secretions from the affected sides were serosanguineous in all cases. A Bacillus sp was isolated in pure cultures in all cases. In 1 case, the Bacillus sp was identified as Bacillus cereus. Treatment and Outcome—Goats were treated for mastitis and endotoxemia with polyionic IV fluid therapy, systemic and intramammary antimicrobial administration, anti-inflammatory drug administration, and other supportive treatment. All goats survived to discharge. All except 1 goat had follow-up information available. The affected udder halves sloughed in 1 to 2 months following discharge. In subsequent lactations after the mastitis episodes, milk production in 2 of 5 goats was above the mean, as determined on the basis of Dairy Herd Improvement records, and 3 of 5 goats were voluntarily withdrawn from lactation. All 5 goats had successful kiddings after the Bacillus mastitis episode. Conclusions and Clinical Relevance—Bacillus sp should be considered as a causative agent in goats with gangrenous mastitis, especially when the Bacillus sp is isolated in a pure culture. Antimicrobial sensitivity testing is recommended for selection of an appropriate antimicrobial for treatment. Prognosis for survival appears to be good, although milk production may be decreased.

Increased incidence of several economically important diseases (i.e. mastitis, metritis, displaced abomasum and ketosis) causes significant animal welfare problems and production losses in transition dairy cattle and decreases the availability of safe and nutritious food for a growing global population. A major underlying factor responsible for the development of transition cow disorders is metabolic stress, which occurs when cows fail to adapt physiologically to an increase in nutrient requirements needed for parturition and the onset of copious milk synthesis and secretion. Metabolic stress can be characterised as resulting from the combined effects of altered nutrient metabolism, dysfunctional inflammatory responses, and oxidative stress. Together, these factors form destructive feedback loops that exacerbate metabolic stress and cause health disorders in transition cows. A better understanding of how nutrition and immunology interact to influence metabolic stress will facilitate the development of control programs to improve transition cow health. The ability to detect signs of metabolic stress early enough in the dry period to implement needed management adjustments before calving will be the key to successful monitoring and intervention programs.