Metabolic profile distinguishes laminitis-susceptible and -resistant ponies before and after feeding a high sugar diet.

BACKGROUND: Insulin dysregulation (ID) is a key risk factor for equine endocrinopathic laminitis, but in many cases ID can only be assessed accurately using dynamic tests. The identification of other biomarkers could provide an alternative or adjunct diagnostic method, to allow early intervention before laminitis develops. The present study characterised the metabolome of ponies with varying degrees of ID using basal and postprandial plasma samples obtained during a previous study, which examined the predictive power of blood insulin levels for the development of laminitis, in ponies fed a high-sugar diet. Samples from 10 pre-laminitic (PL - subsequently developed laminitis) and 10 non-laminitic (NL - did not develop laminitis) ponies were used in a targeted metabolomic assay. Differential concentration and pathway analysis were performed using linear models and global tests. RESULTS: Significant changes in the concentration of six glycerophospholipids (adj. P ≤ 0.024) and a global enrichment of the glucose-alanine cycle (adj. P = 0.048) were found to characterise the response of PL ponies to the high-sugar diet. In contrast, the metabolites showed no significant association with the presence or absence of pituitary pars intermedia dysfunction in all ponies. CONCLUSIONS: The present results suggest that ID and laminitis risk are associated with alterations in the glycerophospholipid and glucose metabolism, which may help understand and explain some molecular processes causing or resulting from these conditions. The prognostic value of the identified biomarkers for laminitis remains to be investigated in further metabolomic trials in horses and ponies.

Paraneoplastic Secretion of Multiple Phosphatonins From a Deep Fibrous Histiocytoma Causing Oncogenic Osteomalacia.

CONTEXT: Literature suggests that oncogenic osteomalacia is usually caused by a benign mesenchymal tumor secreting fibroblast growth factor subtype-23 (FGF-23), but the involvement of other phosphatonins has only been scarcely reported. We have previously published a seemingly typical case of oncogenic osteomalacia. Following curative neoplasm resection, we now report unique molecular characteristics and biology of this tumor. CASE DESCRIPTION: A 25-year-old man had been diagnosed with severe oncogenic osteomalacia that gradually crippled him over 6 years. 68Ga-DOTA-TATE positron emission tomography/computed tomography scan localized the culprit tumor to his left sole, which on resection revealed a deep fibrous histiocytoma displaying a proliferation of spindle cells with storiform pattern associated with multinucleated giant cells resembling osteoclasts. Circulating FGF-23, which was elevated more than 2-fold, declined to undetectable levels 24 h after surgery. Microarray analysis revealed increased tumor gene expression of the phosphatonins FGF-23, matrix extracellular phosphoglycoprotein (MEPE) and secreted frizzled-related protein subtype 4, with elevated levels of all 3 proteins confirmed through immunoblot analysis. Differential expression of genes involved in bone formation and bone mineralization were further identified. The patient made an astonishing recovery from being wheelchair bound to fully self-ambulant 2 months postoperatively. CONCLUSION: This report describes oncogenic osteomalacia due to a deep fibrous histiocytoma, which coincidentally has been found to induce profound muscle weakness via the overexpression of 3 phosphatonins, which resolved fully upon radical resection of the tumor. Additionally, genes involved in bone formation and bone remodeling contribute to the molecular signature of oncogenic osteomalacia.

Changes of microbial and metabolome of the equine hindgut during oligofructose-induced laminitis.

BACKGROUND: Laminitis is a common and serve disease which caused by inflammation and pathological changes of the laminar junction. However, the pathologic mechanism remains unclear. In this study we aimed to investigate changes of the gut microbiota and metabolomics in oligofructose-induced laminitis of horses. RESULTS: Animals submitted to treatment with oligofructose had lower fecal pH but higher lactic acid, histamine, and Lipopolysaccharide (LPS) in serum. Meanwhile, oligofructose altered composition of the hindgut bacterial community, demonstrated by increasing relative abundance of Lactobacillus and Megasphaera. In addition, the metabolome analysis revealed that treatment with oligofructose decreased 84 metabolites while 53 metabolites increased, such as dihydrothymine, N3,N4-Dimethyl-L-arginine, 10E,12Z-Octadecadienoic acid, and asparagine. Pathway analysis revealed that aldosterone synthesis and secretion, regulation of lipolysis in adipocytes, steroid hormone biosynthesis, pyrimidine metabolism, biosynthesis of unsaturated fatty acids, and galactose metabolism were significantly different between healthy and laminitis horses. Furthermore, correlation analysis between gut microbiota and metabolites indicated that Lactobacillus and/or Megasphaera were positively associated with the dihydrothymine, N3,N4-Dimethyl-L-arginine, 10E,12Z-Octadecadienoic acid, and asparagine. CONCLUSIONS: These results revealed that disturbance of gut microbiota and changes of metabolites were occurred during the development of equine laminitis, and these results may provide novel insights to detect biomarkers for a better understanding of the potential mechanism and prevention strategies for laminitis in horses.

Genetics and Signaling Pathways of Laminitis.

Laminitis is a devastating disease with diverse etiologies and few, if any, effective treatments. Gene expression and hypothesis-generating genomic studies have provided a fresh look at the key molecular players at crucial timepoints in diverse experimental and naturally affected tissues. We summarize findings to date, and propose a unifying model of the laminitis disease process that includes several pathogenesis concepts shared with other diseases of epidermal and epithelial tissues. The value of these new pathways as potential therapeutic targets is exciting but will require careful future work to validate new methods and launch systematic clinical trials.

Exotoxins from Staphylococcus aureus activate 5-lipoxygenase and induce leukotriene biosynthesis.

Massive neutrophil infiltration is an early key event in infectious inflammation, accompanied by chemotactic leukotriene (LT)B4 generation. LTB4 biosynthesis is mediated by 5-lipoxygenase (5-LOX), but which pathogenic factors cause 5-LOX activation during bacterial infections is elusive. Here, we reveal staphylococcal exotoxins as 5-LOX activators. Conditioned medium of wild-type Staphylococcus aureus but not of exotoxin-deficient strains induced 5-LOX activation in transfected HEK293 cells. Two different staphylococcal exotoxins mimicked the effects of S. aureus-conditioned medium: (1) the pore-forming toxin α-hemolysin and (2) amphipathic α-helical phenol-soluble modulin (PSM) peptides. Interestingly, in human neutrophils, 5-LOX activation was exclusively evoked by PSMs, which was prevented by the selective FPR2/ALX receptor antagonist WRW4. 5-LOX activation by PSMs was confirmed in vivo as LT formation in infected paws of mice was impaired in response to PSM-deficient S. aureus. Conclusively, exotoxins from S. aureus are potent pathogenic factors that activate 5-LOX and induce LT formation in neutrophils.

Mitochondrial dysfunction is the cause of one of the earliest changes seen on magnetic resonance imaging in Charcot neuroarthopathy - Oedema of the small muscles in the foot.

The hypothesis laid out in this thesis states that the early changes seen on an MR imaging in those with early Charcot neuroarthopathy may be due to mitochondrial dysfunction. In a Charcot foot, there is movement between bones. In an attempt to prevent this movement, the small muscles of the foot contract continuously when the foot is weight bearing. This contraction takes energy in the form of ATP. However, the reduction of glucose transport into the muscle cells due to insulin resistance / insufficiency, leads to reduction in the ATP producing capacity of the mitochondria. The ATP depletion affects the cell membrane gradient leading to mitochondrial and cellular swelling. These early cellular changes could then be picked up with MR imaging as muscle oedema.

Bone penetration of daptomycin in diabetic patients with bacterial foot infections.

OBJECTIVES: Daptomycin has shown clinical efficacy in diabetic foot infections (DFI). However, only limited data are available on its bone penetration in this particular population. The aim of this study was to determine daptomycin bone concentrations in patients with DFI undergoing surgery after multiple daptomycin infusions and to determine bone daptomycin inhibitory quotients (IQs) for the predominant gram-positive species involved in DFI. METHODS: Fourteen adult patients hospitalized with DFI treated with daptomycin and requiring surgical bone debridement and amputation were included in this single-centre prospective study. Daptomycin concentrations in serum and bone were determined by HPLC at steady state. Bone IQs were then calculated according to different minimum inhibitory concentrations (MICs; range 0.25-4mg/l) that are representative of the main MICs for Staphylococcus aureus, coagulase-negative staphylococci (CoNS), and Enterococcus sp populations. RESULTS: Residual and peak concentrations varied from 4.5mg/l to 39.9mg/l and from 31.8mg/l to 110.9mg/l, respectively. Bone daptomycin concentrations at the moment of surgery varied from 1.2mg/l to 17mg/l. Up to a MIC of 1mg/l, which is the epidemiological cut-off value (ECOFF) and breakpoint value for S. aureus and CoNS, all bone daptomycin IQs were positive. The highest bone IQs were observed with Staphylococcus species. Calculated bone IQs for Enterococcus species were often weak at MIC values near the ECOFF. CONCLUSIONS: Daptomycin penetrates bone well in patients treated for DFI. At an initially recommended dosage of 6mg/kg, bone concentrations are likely to be effective against staphylococcal infections and infections due to low-MIC Enterococcus.

Novel lipoprotein density profiling in laminitic, obese, and healthy horses.

Lipoproteins are water-miscible macromolecules enabling the transport of lipids in blood. In humans, altered proportions of lipoproteins are used to detect and classify metabolic diseases. Obesity and obesity-related comorbidities are common in horses. The pathophysiology of obesity is poorly understood and likely multifactorial. Development of new diagnostic tests to identify horses at risk of developing obesity to implement preventative measures is critical; however, a necessary first step to accomplish this goal is to improve our understanding of the pathophysiology of disease. Thus, the objective of this study was to characterize and compare lipoprotein profiles of horses with normal and excess body conditions, with and without laminitis using a novel method of continuous lipoprotein density profiling (CLPDP). Comparisons were made between 4 groups of horses: (1) laminitic, obese horses (n = 66); (2) laminitic, nonobese horses (n = 35); (3) nonlaminitic, obese horses (n = 41); and (4) nonlaminitic, nonobese horses (n = 95). Lipoprotein profiling, including evaluation of triglyceride-rich lipoprotein (TRL), low-density lipoproteins (LDLs), and high-density lipoproteins (HDLs) was performed using CLPDP, and all 4 groups were compared. A significant difference was observed among groups for the subfractions TRL, LDL1, LDL2, HDL2b, HDL2a, HDL3a, HDL3b, HDL3c, and total HDL. This is the first known description of CLPDP to characterize equine lipid profiles and holds promise as a useful method for lipid characterization of horses.

Detection of endoplasmic reticulum stress and the unfolded protein response in naturally-occurring endocrinopathic equine laminitis.

BACKGROUND: Laminitis is often associated with endocrinopathies that cause hyperinsulinemia and is also induced experimentally by hyperinsulinemia, suggesting that insulin initiates laminitis pathogenesis. Hyperinsulinemia is expected to activate pro-growth and anabolic signaling pathways. We hypothesize that chronic over-stimulation of these pathways in lamellar tissue results in endoplasmic reticulum stress, contributing to tissue pathology, as it does in human metabolic diseases. We tested this hypothesis by asking whether lamellar tissue from horses with naturally-occurring endocrinopathic laminitis showed expression of protein markers of endoplasmic reticulum stress. RESULTS: Three markers of endoplasmic reticulum stress, spliced XBP1, Grp78/BiP and Grp94, were upregulated 2.5-9.5 fold in lamellar tissues of moderately to severely laminitic front limbs (n = 12) compared to levels in controls (n = 6-7) measured by immunoblotting and densitometry. Comparing expression levels between laminitic front limbs and less affected hind limbs from the same horses (paired samples from 7 to 8 individual horses) demonstrated significantly higher expression for both spliced XBP1 and Grp78/BiP in the laminitic front limbs, and a similar trend for Grp94. Expression levels of the 3 markers were minimal in all samples of the control (n = 6-7) or hind limb groups (n = 7-8). Immunofluorescent localizations were used to identify cell types expressing high levels of Grp78/BiP, as an indicator of endoplasmic reticulum stress. Grp78/BiP expression was highly elevated in suprabasal epidermal keratinocytes and only observed in laminitic front limbs (10/12 laminitic samples, compared to 0/7 in sections from the hind limbs and 0/5 of controls). CONCLUSIONS: These data demonstrate that the endoplasmic reticulum stress pathway is active in naturally occurring cases of laminitis and is most active within a subset of epidermal keratinocytes. These data provide the rationale for further study of endoplasmic reticulum stress in experimental models of laminitis and the links between laminitis and human diseases sharing activation of this stress pathway. Pharmacological options to manipulate the endoplasmic reticulum stress pathway under investigation for human disease could be applicable to laminitis treatment and prevention should this pathway prove to be a driver of disease progression.

Claw horn lesions in mid-lactation primiparous dairy cows under pasture-based systems: Association with behavioral and metabolic changes around calving.

The objectives of this study were to describe the lying behavior of primiparous dairy cows under pasture-based systems during the pre- and postcalving period and characterize the association of lying behavior and analytes related to energy metabolism during this period with claw horn disruption lesion development later in lactation. Our convenience sample included 39 primiparous Holstein cows from 3 commercial farms that were assessed for body condition score (BCS; 5-point scale, 0.25-point increments) and had blood collected at wk -3, -2, -1, 1, 2, and 3 relative to calving date. Blood samples were assayed for nonesterified fatty acids, ß-hydroxybutyrate (BHB), and cholesterol concentrations. Electronic data loggers (HOBO Pendant G Acceleration, Onset Computer Corporation, Bourne, MA) recorded lying behavior at 1-min intervals from 3 wk before calving to 3 wk after calving. Starting at 4 wk after calving and until 16 wk after calving, cows were examined for claw lesions at approximately 4-wk intervals. Sole lesions and white line lesions were scored on a 0 to 10 scale. Of the 39 primiparous cows, 19 cows scored 0 at all exams during the entire study period and 20 cows had at least 1 severe lesion (score ≥4) between 8 and 16 wk after calving. Time spent lying before calving averaged 10.3 ± 0.3 h/d, but declined to 7.3 ± 0.3 h/d after calving (least squares means ± standard error). At calving, we noted an increase in the number of lying bouts (12.9 ± 0.45 bouts/d) compared with the pre- and postcalving averages of 11.6 (±0.53) and 9.1 (±0.47) bouts, respectively. Cows that developed claw lesions later in mid lactation spent less time lying down than cows without lesions during wk 3 after calving compared with healthy cows (7.29 ± 0.22 vs. 8.51 ± 0.16 h/d). Lesion cows had fewer lying bouts per day, and these bouts were of longer duration than no-lesion cows after calving. Increased odds of lesion were found to be associated with shorter lying times and fewer number of lying bouts during wk 3 (odds ratio = 1.23). Nonesterified fatty acids (747 ± 58 vs. 990 ± 86.85 µmol/L) and BHB (0.77 ± 0.06 vs. 0.60 ± 0.04 mmol/L) concentrations during wk 1 were greater in cows that developed claw lesions relative to cows that did not develop lesions. The BHB concentrations also remained higher in wk 2 for cows that developed claw lesions (0.63 ± 0.04 vs. 0.46 ± 0.03 mmol/L) compared with cows that did not develop any lesions. Cows that developed lesions experienced greater losses in BCS from wk -3 to 3 than cows without lesions (0.74 ± 0.01 and 0.61 ± 0.01 BCS change, respectively). In summary, changes in lying behavior and energy metabolic status after calving were associated with claw horn disruption lesions in mid-lactation primiparous cows under pasture-based systems.

[Perfil de expresión de Ki67 en lesiones melanocíticas palmoplantares: estudio de casos y controles]. / Expression profile of Ki67 in palmoplantar melanocytic lesions: a case-control study.

INTRODUCCIÓN: El melanoma acral lentiginoso es una neoplasia maligna que afecta a población predominantemente no caucásica. Debido al diagnóstico tardío suele tener mal pronóstico, además de que se considera una neoplasia biológicamente más agresiva, incluso cuando se detecta tempranamente. OBJETIVO: Determinar la expresión de Ki67 en el melanoma acral lentiginoso invasor y compararla con los nevos acrales. MÉTODO: Estudio transversal, descriptivo, observacional. Se realizó inmunohistoquímica con marcador Ki67 en 17 biopsias de melanoma acral lentiginoso invasor (casos) y 17 biopsias de nevos palmoplantares (controles). Se determinó la expresión nuclear de Ki-67 y se comparó entre ambos grupos. RESULTADOS: La media de expresión de Ki67 fue del 8.5% en el grupo control y del 34% en el grupo de melanomas, siendo esta diferencia estadísticamente significativa (p < 0.0001). DISCUSIÓN: La expresión de Ki67 en los melanomas acrales es considerablemente mayor que en los nevos acrales. El valor pronóstico del marcador Ki67 sigue siendo considerado controversial. Sin embargo, hay estudios en los que en combinación con otros marcadores se refuerza su valor pronóstico. CONCLUSIONES: Por la gran diferencia en inmunorreactividad de Ki67 entre melanomas y nevos, la expresión de Ki67, referida como índice proliferativo, podría ser considerada como factor pronóstico incluso más objetivo que el índice mitótico. BACKGROUND: Acral lentiginous melanoma is a malignant neoplasm which appears in hands and feet. Acral lentiginous melanoma has an unclear etiology, and usually affects non-Caucasian population. Because it is frequently diagnosed lately, acral melanoma has bad prognosis; however, it is biologically more aggressive than other clinicopathological types of melanoma, even when diagnosed early. OBJECTIVE: To determine the expression of Ki67 in invasive lentiginous acral melanoma and to compare it with acral nevi. METHOD: Cross-sectional, descriptive, observational study. Immunohistochemistry with Ki67 marker was performed on 17 biopsies of invasive lentiginous acral melanoma (cases) and 17 biopsies of palmoplantar nevi (controls). Nuclear expression of Ki-67 was determined and both were compared between both groups. RESULTS: The mean expression of Ki67 was 8.5% in the control group, and 34% in the melanoma group, which was statistically significant (p < 0.0001). DISCUSSION: Ki67 expression in acral lentiginous melanomas is higher than in acral nevi. Prognostic value of Ki67 is still considered controversial. However, there are several studies where, in combination with other markers, their prognostic value is reinforced. CONCLUSIONS: Due to the wide gap in Ki67 expression between melanomas and nevi showed in this study, Ki67 expression, referred to as a proliferative index, could be considered as a prognostic factor even more objective than the mitotic index.

Metabolic perturbations in Welsh Ponies with insulin dysregulation, obesity, and laminitis.

BACKGROUND: Metabolomics, the study of small-molecule metabolites, has increased understanding of human metabolic diseases, but has not been used to study equine metabolic syndrome (EMS). OBJECTIVES: (1) To examine the serum metabolome of Welsh Ponies with and without insulin dysregulation before and during an oral sugar test (OST). (2) To identify differences in metabolites in ponies with insulin dysregulation, obesity, or history of laminitis. ANIMALS: Twenty Welsh Ponies (mean ± SD; 13.8 ± 9.0 years) classified as non-insulin dysregulated [CON] (n = 10, insulin < 30 mU/L) or insulin dysregulated [ID] (n = 10, insulin > 60 mU/L) at 75 minutes after administration of Karo syrup, obese (n = 6) or nonobese (n = 14), and history of laminitis (n = 9) or no history of laminitis (n = 11). METHODS: Case-control study. Metabolomic analysis was performed on serum obtained at 0 minutes (baseline) and 75 minutes during the OST. Data were analyzed with multivariable mixed linear models with significance set at P ≤ .05. RESULTS: Metabolomic analysis of 646 metabolites (506 known) detected significant metabolite differences. At baseline, 55 metabolites (insulin response), 91 metabolites (obesity status), and 136 metabolites (laminitis history) were different. At 75 minutes, 51 metabolites (insulin response), 102 metabolites (obesity status), and 124 metabolites (laminitis history) were different. CONCLUSIONS AND CLINICAL IMPORTANCE: Use of metabolomics could have diagnostic utility for early detection of EMS and provide new knowledge regarding the pathophysiology of metabolic perturbations associated with this condition that might lead to improved clinical management.

Effect of varying the dose of corn syrup on the insulin and glucose response to the oral sugar test.

BACKGROUND: The oral sugar test (OST) is used to identify equine insulin dysregulation (ID); however only a dose of 0.15 mL/kg bwt corn syrup has been evaluated. OBJECTIVES: To determine the effect of varying the dose of corn syrup on insulin and glucose response to the OST and the test's ability to distinguish between ponies with a history of laminitis (PL) and without laminitis (NL). STUDY DESIGN: Randomised crossover experiment. METHODS: After an overnight fast, in a three-way randomised crossover study with a 7-day washout, 0.15, 0.3 or 0.45 mL/kg bwt corn syrup was administered orally to eight ponies (5 PL and 3 NL) and blood obtained between 0 and 120 min. Serum [insulin] and [glucose] were measured using previously validated radioimmunoassay and colorimetric assays respectively. The repeatability of and the effect of continued pasture access on the dose that best distinguished PL and NL ponies were then assessed. The effect of dose, laminitis history and fasting on serum [insulin] and [glucose] responses were assessed using mixed-effects models. RESULTS: The serum [insulin] following 0.15 mL/kg bwt were not significantly different from 0.3 mL/kg bwt at any time point, while serum [insulin] following 0.45 mL/kg bwt significantly (P<0.01) differed from 0.15 and 0.3 mL/kg bwt at all time points apart from 0 min. The serum [insulin] concentration significantly (P<0.01) differed between NL (mean 86 [95% CI 59, 113] µiu/mL) and PL (146 [95% CI 124, 167] µiu/mL) only following 0.45 mL/kg bwt at 60 min. Repeatability of serum [insulin] at 60 min following 0.45 mL/kg bwt dose under fasted conditions was 0.51. Using AUC insulin improved repeatability to 0.83. There was no significant difference between the fasted and at pasture results. MAIN LIMITATIONS: The OST was performed in small numbers of ponies on limited occasions. CONCLUSIONS: A dose of 0.45 mL/kg bwt corn syrup may be preferable to differentiate PL and NL ponies.

Effect of Mitomycin C on recurrence of plantar fibromas.

Although certainly not the first line treatment for plantar fibromas, surgical resection is a treatment option for some patients with have failed exhaustive non-surgical treatment. The use of topical Mitomycin C has been recently shown to reduce the recurrence rate of other fibrous lesions. The purpose of this study was to determine the impact of topical application of Mitomycin C on recurrence rate of plantar fibromas. A retrospective analysis was done from a prospectively gathered database with a total 50 consecutive patients over a 16-month study period. The control group (n = 29) consisted of patients who underwent only surgical resection, while the study group (n = 21) consisted of patients who underwent surgical resection with adjuvant therapy using Mitomycin C. The primary endpoint was local recurrence after the procedure. Secondary end points included complications and toxicity associated with this medication. No patients were lost to follow up. Of the 29 patients in the control group, there were 17 patients (17/29, 58.6%) had recurrence of the plantar fibroma at a mean follow-up of 9.1 months. In contrast, in the experimental study group, all patients were free from local recurrence. No complications or side effects were associated with Mitomycin C use. The results demonstrate that the topical application of Mitomycin C to the tumor bed after surgical resection of plantar fibromas reduced the recurrence rate. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2554-2561, 2018.

The diagnosis of equine insulin dysregulation.

Insulin dysregulation is the hallmark of equine metabolic syndrome and has received attention because of its direct association with laminitis. In the absence of an adequate treatment for laminitis, a focus on prophylaxis is needed, making early detection of individuals at risk of developing laminitis one of the main challenges in equine endocrinology. Recent studies have shown that insulin dysregulation goes beyond tissue insulin resistance and it is now demonstrated that the equine enteroinsular axis plays a major role in insulin secretion and equine hyperinsulinaemia. In this review, we discuss the different tests currently available to diagnose insulin dysregulation in horses: the ones investigating tissue insulin resistance and those investigating the enteroinsular axis, detailing their goals, practicalities and limitations. This review supports the contention that the diagnosis of equine insulin dysregulation should now be based on the investigation of both tissue insulin resistance and the equine enteroinsular axis. Regardless of the tests used many factors of variation, such as breed, diet, fasting state or season, have been identified and could potentially confound the results of a specific test. Therefore, careful interpretation of the results of a given test in each individual situation is required to optimise the detection of horses at risk of laminitis.

Lamellar events related to insulin-like growth factor-1 receptor signalling in two models relevant to endocrinopathic laminitis.

BACKGROUND: Insulin dysregulation, obesity, and exposure to high-nonstructural carbohydrate (NSC) forage are risk factors for equine metabolic syndrome-associated laminitis (EMSAL); high systemic insulin concentrations in EMSAL are proposed to induce cellular dysregulation in the digital lamellae through activation of the insulin-like growth factor-1 receptor. OBJECTIVES: To use a dietary challenge model (DCM) and a euglycaemic-hyperinsulinaemic clamp (EHC) model to assess lamellar growth factor-related signalling. STUDY DESIGN: Lamellar phospho (P)-protein concentrations of signalling proteins important in growth factor-related signalling were assessed in 2 models: 1) lean and obese ponies on a low- or high-NSC diet; and 2) EHC model using Standardbred horses. METHODS: Ponies stratified for body condition (lean [LN, n = 11] and obese [OB, n = 11]) were exposed to a low-NSC diet (LO, n = 5 per group for LN LO and OB LO) or a high NSC diet (HI, n = 6 per group for LN HI and OB HI groups) for 7 days. For the EHC model, horses were administered insulin (constant rate infusion [6 mIU/kg bwt/min] combined with 50% dextrose, EHC group, n = 8)] or saline (0.57 mL/kg bwt/h, CON group, n = 8) for 48 h. Immunoblotting was employed to assess concentrations of activated/phosphorylated and total protein for members of the PI3K/Akt/mTORC1 and Ras/ERK pathways in lamellar samples from both models. RESULTS: In the DCM, lamellar P-(Ser 240/244) RPS6 was increased in OB HI ponies (vs. OB LO, P<0.05); positive correlations existed (P<0.05; r>0.5) between Day 7 basal serum insulin concentrations and lamellar concentrations of P-p70S6K and P-(Ser 240/244) RPS6. In the EHC model, lamellar concentrations of P-Akt, P-p70S6K, P-ERK 1/2, P-p90RSK, and both P-(Ser 235/236) and P-(Ser 240/244) RPS6 were increased in the EHC group (vs. CON, P<0.05). MAIN LIMITATIONS: The primary limitations of this study are the small number of animals per group in the DCM study, and the fact that many animals did not develop laminitis as that was not the endpoint of either study. CONCLUSIONS: These results support further investigation of mTORC1/RPS6 signalling as a potential therapeutic target(s) in EMSAL. The Summary is available in Chinese - see Supporting Information.

Claw disorders in dairy cattle - an unexpected association between energy metabolism and sole haemorrhages.

The present study investigated whether changes of energy metabolism post-partum (pp) are associated with claw health. For this purpose, back-fat-thickness (BFT) was measured and blood samples were taken from 146 cows at four examination times. The serum levels of free fatty acids (FFA), ß-hydroxybutyrate (BHB) and glucose were measured. Furthermore, in the first week postpartum (pp) and at 8 weeks pp, a claw trimming was done and the presence and extent of sole haemorrhages (SH) was recorded. Animals with high BFT at calving and therefore high fat mobilisation and whose FFA and BHB levels in the first week pp exceeded the reference values had fewer pathological changes of the claws than thinner animals whose FFA and BHB levels stayed within reference ranges. The body condition before calving, represented in this study by BFT, plays an important role in non-infectious claw disorders. Poorer body condition was found to be associated with the SH that develop in the first 2 months of lactation.

Vitamin D receptor immunohistochemistry variability in sun-exposed and non-sun-exposed melanomas.

Ultraviolet rays are one of the leading factors in the development of melanoma (MM); however, ultraviolet rays seem not to play a role in non-sun-exposed MM, where systemic immunosuppression, anatomical, and physiological features may contribute toward the development of the malignancy. Our aim was to evaluate vitamin D receptor (VDR) expression in shield-site melanoma (ST-MM) and non-shield-site melanoma (NST-MM) to find features that could explain the different biological behavior of MM according to the area of onset. We reviewed 118 specimens of MM. VDR expression was assayed using immunohistochemistry by dividing the specimens according to the anatomical area. We included MM of the soles, feet, hands, gluteus, scrotum, skin of the penile shaft, and large vaginal labia in the ST-MM group. The NST-MM group was divided into two main categories: NST-MM of chronic sun-exposed areas, including MM of the face, scalp, neck, back of the hands, and NST-MM of intermittent sun-exposed areas, including MM of the trunk, lower, and upper limbs. In shield sites, 66.67% of MMs showed VDR expression; in intermittent sun-exposed areas, 33.3% showed VDR expression; and in chronic sun-exposed areas, only 4.66% showed VDR expression. A similar behavior was observed for Breslow's thickness, where VDR staining intensity was higher in thicker lesions, ranging between 60 and 100%. We found that VDR expression decreased from ST-MM to NST-MM. These findings confirm the hypothesis that different pathways are involved in ST-MM and NST-MM.

Insulin-Like Growth Factor II mRNA-Binding Protein 3 Expression Correlates with Poor Prognosis in Acral Lentiginous Melanoma.

Insulin-like growth factor-II mRNA-binding protein 3 (IMP-3) is an RNA-binding protein expressed in multiple cancers, including melanomas. However, the expression of IMP-3 has not been investigated in acral lentiginous melanoma (ALM). This study sought to elucidate its prognostic value in ALMs. IMP-3 expression was studied in 93 patients diagnosed with ALM via immunohistochemistry. Univariate and multivariate analyses for survival were performed, according to clinical and histologic parameters, using the Cox proportional hazard model. Survival curves were graphed using the Kaplan-Meier method. IMP-3 was over-expressed in 70 out of 93 tumors (75.3%). IMP-3 expression correlated with thick and high-stage tumor and predicted poorer overall, melanoma-specific, recurrence-free and distant metastasis-free survivals (P = 0.002, 0.006, 0.008 and 0.012, respectively). Further analysis showed that patients with tumor thickness ≤ 4.0 mm and positive IMP-3 expression had a significantly worse melanoma-specific survival than those without IMP-3 expression (P = 0.048). IMP-3 (hazard ratio 3.67, 95% confidence intervals 1.35-9.97, P = 0.011) was confirmed to be an independent prognostic factor for melanoma-specific survival in multivariate survival analysis. Positive IMP-3 expression was an important prognostic factor for ALMs.