Growth of rumen papillae in weaned calves is associated with lower expression of insulin‐like growth factor‐binding proteins 2, 3, and 6

This study aimed to characterize the relationship between the growth of rumen papillae in calves and the mRNA expression of insulin‐like growth factor‐binding proteins (IGFBPs) in the rumen papillae. The length of rumen papillae, the mRNA expression of IGFBPs in rumen papillae by quantitative real‐time PCR, and the presence of insulin‐like growth factors I and II (IGF‐I and II) by immunohistochemistry (IHC) were analyzed in nine Holstein calves divided into three groups: suckling (2 weeks, n = 3), milk‐continued (8 weeks, n = 3), and weaned (8 weeks, n = 3). The length of rumen papillae was greater (p < 0.01) in weaned calves than in suckling and milk‐continued calves, whereas the expressions of IGFBP2, IGFBP3, and IGFBP6 genes were lower (p < 0.05) in the rumen papillae of weaned calves than in milk‐continued calves. Thus, rumen papillae length and IGFBP2, 3, and 6 expressions were negatively correlated. The IHC analysis showed that IGF‐I and IGF‐II were present in the rumen epithelium of calves. These results suggested that the growth of rumen papillae after weaning is associated with the induction of IGFs by the low levels of IGFBP2, IGFBP3, and IGFBP6.     

Gene expression of resistin and TNF-α in adipose tissue of Japanese Black steers and Holstein steers

The aim of the present study was to compare the expression of adipose tissue mRNA related to glucose metabolism between Japanese Black steers (n = 5) and Holstein steers (n = 5). We examined the expression of the resistin, tumor necrosis factor‐α (TNF‐α), glucose transporter 1 (GLUT1) and growth hormone receptor (GHR) genes using real‐time polymerase chain reaction of cDNA in adipose tissue. The cDNA sequence identified by 5′/3′‐rapid amplification of cDNA and the deduced amino acid sequence were highly conserved in human, porcine and murine resistin. Expression of resistin mRNA was significantly greater in Holstein steers than in Japanese Black steers. In contrast, expression of TNF‐α mRNA was slightly greater in Japanese Black steers. Expression of GHR mRNA was significantly greater in Japanese Black steers compared with the Holstein steers, although there was no significant difference in the expression of GLUT1 mRNA. However, the plasma non‐esterified fatty acid (NEFA), glucose, insulin and growth hormone concentrations did not differ between Japanese Black and Holstein steers. The present results show that there is a difference in the expression level of mRNA related to glucose metabolism between Japanese Black steers and Holstein steers.     

Mechanism of insulin action on glucose metabolism in ruminants

This review presents a brief overview on the mechanism of insulin action on glucose metabolism at the molecular basis in ruminants. For ruminants, an exact mechanism of insulin on glucose metabolism is still rudimentary, but it is clear that originally, if not all, the mechanism of insulin action in ruminants was the same as in other species. Like non‐ruminants, the insulin‐sensitive glucose transporter GLUT 4 is thought to be a key‐protein in the control of glucose uptake and metabolism in ruminants, and insulin regulates glucose transport by stimulating the translocation of GLUT 4 from an intracellular membrane pool to the plasma membrane in adipocytes and muscles. Moreover, insulin‐induced GLUT 4 translocation is activated through the common intracellular signaling pathway of insulin phosphatidylinositol 3‐kinase (PI3‐kinase) signaling pathway rather than the mitogen activated protein kinase (MAP kinase)‐dependent signaling pathway. However, GLUT 4 mRNA and protein, and insulin‐induced GLUT 4 translocation on adipocytes and muscles in ruminants are lower than those in rodents and human subjects. Furthermore, insulin‐induced PI3‐kinase activation is reduced concomitantly with the lower content of insulin receptor substrate‐1 (IRS‐1) in ruminants. In spite of normal status, a resistance to the stimulatory action of insulin on glucose metabolism in ruminants as compared to non‐ruminants may be due to, at least in part, the lower content of GLUT 4 and the lower capacity of insulin signal transduction, resulting to the lower glucose transport activity.     

Glucose transport and milk secretion during manipulated plasma insulin and glucose concentrations and during LPS‐induced mastitis in dairy cows

In dairy cows, glucose is essential as energy source and substrate for milk constituents. The objective of this study was to investigate effects of long‐term manipulated glucose and insulin concentrations in combination with a LPS‐induced mastitis on mRNA abundance of glucose transporters and factors involved in milk composition. Focusing on direct effects of insulin and glucose without influence of periparturient endocrine adaptations, 18 dairy cows (28 ± 6 weeks of lactation) were randomly assigned to one of three infusion treatments for 56 h (six animals each). Treatments included a hyperinsulinemic hypoglycaemic clamp (HypoG), a hyperinsulinemic euglycaemic clamp (EuG) and a control group (NaCl). After 48 h of infusions, an intramammary challenge with LPS from E. coli was performed and infusions continued for additional 8 h. Mammary gland biopsies were taken before, at 48 (before LPS challenge) and at 56 h (after LPS challenge) of infusion, and mRNA abundance of genes involved in mammary gland metabolism was measured by RT‐qPCR. During the 48 h of infusions, mRNA abundance of glucose transporters GLUT1, 3, 4, 8, 12, SGLT1, 2) was not affected in HypoG, while they were downregulated in EuG. The mRNA abundance of alpha‐lactalbumin, insulin‐induced gene 1, κ‐casein and acetyl‐CoA carboxylase was downregulated in HypoG, but not affected in EuG. Contrary during the intramammary LPS challenge, most of the glucose transporters were downregulated in NaCl and HypoG, but not in EuG. The mRNA abundance of glucose transporters in the mammary gland seems not to be affected by a shortage of glucose, while enzymes and milk constituents directly depending on glucose as a substrate are immediately downregulated. During LPS‐induced mastitis in combination with hypoglycaemia, mammary gland metabolism was more aligned to save glucose for the immune system compared to a situation without limited glucose availability during EuG.     

Ruminal epithelial insulin‐like growth factor‐binding proteins 2, 3, and 6 are associated with epithelial cell proliferation

The aim of this study was to identify factors that regulate ruminal epithelial insulin‐like growth factor‐binding protein (IGFBP) expression and determine its role in rumen epithelial cell proliferation. Primary bovine rumen epithelial cells (BREC) were incubated with short‐chain fatty acids (SCFAs) at pH 7.4 or 5.6, lactate, lipopolysaccharide (LPS), insulin‐like growth factor‐I (IGF‐I), ‐II (IGF‐II), or recombinant bovine IGFBP2 (rbIGFBP2). The mRNA expression levels of IGFBP in BREC were analyzed using quantitative real‐time polymerase chain reaction (qRT‐PCR). The proliferation rate of BREC was analyzed using a WST‐1 assay. IGFBP2 gene expression tended to be lower with SCFA treatment (p < .1), and IGFBP6 gene expression was significantly lower with SCFA treatment (p < .05). IGFBP3 and IGFBP6 gene expression tended to be higher with d ‐Lactate treatment (p < .1). IGFBP3 gene expression was significantly higher (p < .05) with LPS treatment. BREC treated with IGF‐I grew more rapidly than vehicle control‐treated cells (p < .01); however, recombinant bovine rbIGFBP2 inhibited IGF‐I‐induced proliferation. IGF‐II and/or rbIGFBP2 did not affect BREC proliferation. Taken together, SCFA treatment decreased IGFBP2 and IGFBP6 expression in rumen epithelial cells, and lower expression of these IGFBP might promote rumen epithelial cell proliferation by facilitating IGF‐I.     

Insulin Sensitivity during Late Gestation in Ewes Affected by Pregnancy Toxemia and in Ewes with High and Low Susceptibility to this Disorder

Background

Insulin resistance during late gestation might act as 1 etiologic factor causing pregnancy toxemia in ewes.

Objective

Evaluation of pancreatic insulin secretion and peripheral insulin sensitivity in ewes with differing susceptibility to pregnancy toxemia and in ketotic ewes.

Animals

Pregnant ewes suffering from (PT, n = 5) and ewes with high (HR, n = 7) and low risk (LR, n = 5) of being affected by pregnancy toxemia.

Methods

In a case‐control study, the pancreatic insulin release and the peripheral insulin sensitivity were assessed by means of the intravenous glucose tolerance test with subsequent measurement of the plasma concentrations of glucose, insulin, nonesterified fatty acids (NEFA), and β‐hydroxybutyrate (β‐HB). The ewes were tested during late pregnancy within 5 and 15 days antepartum.

Results

The insulin secretion after glucose administration was significantly lower in the HR and PT than in the LR ewes. The baseline rate of lipolysis was significantly increased in the HR ewes, but the NEFA clearance was similar in both risk groups, albeit delayed in the PT ewes. The baseline β‐HB concentration was significantly higher in the PT than in the HR and LR ewes. In the HR and in the PT ewes, the plasma β‐HB concentrations did not decrease after glucose administration.

Conclusion and Clinical Importance

There is reduced pancreatic first‐phase insulin response and impaired insulin‐dependent inhibition of the ketone body formation during late pregnancy in the HR and PT ewes. This insulin resistance might represent 1 causative factor in the pathogenesis of ovine pregnancy toxemia.     

Ganglioneuroblastoma in a cat: a rare neoplasm treated with electrochemotherapy

An 8-year-old male castrated cat was referred for sudden onset of lameness. Physical examination revealed a 1x2x1cm mass originating from a footpad of the right hind leg. A diagnosis of ganglioneuroblastoma was suggested by the tumour appearance following histopathological staining with haematoxylin and eosin and haematoxylin/van Gieson. Immunohistochemical staining for glial fibrillary acidic protein (GFAP), vimentin, neuron-specific enolase (NSE), neurofilament and S100 further confirmed the diagnosis. The staging process did not indicate metastatic spread. The cat was treated with three sessions of electrochemotherapy (ECT) 1 week apart, following local injection of bleomycin. The tumour had completely regressed within 1 week of the third ECT application and remained in remission for 402 days at which time a small recurrence was noted. The animal was given a further session of ECT using intra-lesional cisplatin and again went into remission. It remained tumour free at 450 days. Electrochemotherapy is considered a safe and effective treatment for localised neoplasms of cats and dogs and warrants further investigation.     

Effect of nutrient intake on intramuscular glucose metabolism during the early growth stage in cross‐bred steers (Japanese Black male × Holstein female)

The objective was to investigate the impact of nutrient intake during the early growth period on the expression of glucose metabolism‐related genes in skeletal muscle of cross‐bred cattle. From 1.5 to 5 months of age, group H (n = 7) animals were intensively fed a high‐protein and low‐fat milk replacer [crude protein (CP) 28%; ether extracts (EE) 18%; max: 2.0 kg, 12 l/day], and group R (n = 7) animals were fed a restricted amount of normal milk replacer (CP 25%; EE 23%; max 0.5 kg, 4 l/day). From 6 to 10 months of age, group H cattle were fed a high‐nutrition total mixed ration mainly prepared from grain feed, and group R cattle were fed only roughage. Blood samples were taken from each animal at three biopsy times (1.5, 5 and 10 months of age), and the blood plasma concentration of glucose and insulin was analysed. In glucose concentration, there were no significant differences; however, the concentrations of insulin were higher in group H than in group R at 5 and 10 months of age. Muscle samples were taken by biopsy from longissimus thoracis muscle (LT) at 1.5, 5 and 10 months of age. We analysed mRNA expression levels using the quantitative real‐time polymerase chain reaction (PCR) assay for glucose transporters (GLUT1 and GLUT4), insulin receptor, phosphatidylinositol 3‐kinase (PI‐3K), protein kinase B (PKB, also known as Akt), hexokinase 1 (HK1) and tumour necrosis factor alpha (TNFα). Although no differences were detected at 1.5 and 5 months of age, at 10 months of age, GLUT1, HK1 and TNFα mRNA expression levels were significantly higher in group H than in group R. These results suggested Glut1 that affects insulin‐independently mediated glucose uptake was more responsive to improved nutrition during early growth stage than GLUT4 that insulin‐dependently mediated glucose uptake in LT of cattle.     

Urinary and faecal N-methylhistamine concentrations do not serve as markers for mast cell activation or clinical disease activity in dogs with chronic enteropathies

Background

This study sought to correlate faecal and urinary N-methylhistamine (NMH) concentrations with resting versus degranulated duodenal mast cell numbers in dogs with chronic enteropathies (CE), and investigate correlations between intestinal mast cell activation and clinical severity of disease as assessed by canine chronic enteropathy clinical activity index (CCECAI), and between urinary and faecal NMH concentrations, mast cell numbers, and histopathological scores. Twenty-eight dogs with CE were included. Duodenal biopsies were stained with haematoxylin and eosin (H&E), toluidine blue, and by immunohistochemical labelling for tryptase. Duodenal biopsies were assigned a histopathological severity score, and duodenal mast cell numbers were counted in five high-power fields after metachromatic and immunohistochemical staining. Faecal and urinary NMH concentrations were measured by gas chromatography–mass spectrometry.

Results

There was no correlation between the CCECAI and faecal or urinary NMH concentrations, mast cell numbers, or histopathological score – or between faecal or urinary NMH concentration and mast cell numbers. Post hoc analysis revealed a statistically significant difference in toluidine blue positive mast cells between two treatment groups (exclusion diet with/without metronidazole versus immunosuppression (IS)), with higher numbers among dogs not requiring IS.

Conclusion

Faecal and urinary NMH concentrations and duodenal mast cell numbers were not useful indicators of severity of disease as assessed by the CCECAI or histological evaluation. The number of duodenal mast cells was higher in dogs that did not need IS, i.e. in dogs responding to an exclusion diet (with/without metronidazole), than in dogs requiring IS. Further studies comparing the role of mast cells in dogs with different forms of CE are needed.     

The canine hair cycle - a guide for the assessment of morphological and immunohistochemical criteria

The hair follicle has a lifelong capacity to cycle through recurrent phases of controlled growth (anagen), regression (catagen) and quiescence (telogen), each associated with specific morphological changes. A comprehensive classification scheme is available for mice to distinguish the cycle stages anagen I-VI, catagen I-VIII and telogen. For dogs, such a classification system does not exist, although alopecia associated with hair cycle arrest is common. We applied analogous morphological criteria and various staining techniques to subdivide the canine hair cycle stages to the same extent as has been done in mice. Of all the staining techniques applied, haematoxylin and eosin stain, Sacpic, Masson Fontana and immunohistochemistry for vimentin and laminin proved to be most useful. To evaluate the applicability of our criteria, we investigated skin biopsies from healthy beagle dogs (n=20; biopsies from shoulder and thigh) kept in controlled conditions. From each biopsy, at least 50 hair follicles were assessed. Statistical analysis revealed that 30% of the follicles were in anagen (12% early and 18% late), 8% in catagen (2% early, 5% late and 1% not determinable) and 27% in telogen. Thirty-five per cent of hair follicles could not be assigned to a specific cycle stage because not all follicles within one biopsy were oriented perfectly. In conclusion, this guide will not only be helpful for the investigation of alopecic disorders and possibly their pathogenesis, but may also serve as a basis for research projects in which the comparison of hair cycle stages is essential, e.g. comparative analysis of gene expression patterns.     

Changes in the gene expression of adiponectin and glucose transporter 12 (GLUT12) in lactating and non-lactating cows

Glucose delivery and uptake by the mammary gland is a rate‐limiting step in milk synthesis. Insulin resistance is believed to increase throughout the body following the onset of lactation. To study glucose metabolism in peak‐, late‐, and non‐lactating cows we analyzed the expression of an adipokine, namely, adiponectin, decreased insulin resistance, leptin, and a novel insulin‐responsive glucose transporter (GLUT12) in the adipose tissue and mammary gland by using real‐time polymerase chain reaction. Our results demonstrated that the mRNA level of adiponectin in the adipose tissue was greater in non‐lactating cows than in peak‐lactating cows. In the adipose tissue, there were no significant differences in the abundance of GLUT12 mRNA between the peak‐, late‐, and non‐lactating cows. In contrast, in the mammary gland, the mRNA level of GLUT12 was greater in non‐lactating cows than in peak‐ and late‐lactating cows. In the adipose tissue, the mRNA level of leptin and peroxisome proliferator‐activated receptor gamma 2 (PPARγ2) was greater in non‐lactating cows than in peak‐lactating cows. The results of the present study suggest that in lactating cows adiponectin plays an important role in insulin resistance in the adipose tissue; in the mammary gland, GLUT12 expression is believed to be an important factor for insulin‐dependent glucose metabolism.     

Calponin expression and myoepithelial cell differentiation in canine, feline and human mammary simple carcinomas

Calponin is a 34‐kDa smooth muscle‐specific protein that has been shown to be a highly sensitive marker of myoepithelial cells in canine, feline and human mammary tissue and tumours. The expression of calponin was studied in 15 canine, 32 feline and 28 human simple mammary carcinomas using a monoclonal mouse antihuman calponin antibody and the avidin–biotin peroxidase complex (ABC) immunohistochemical technique. Calponin expression was compared with the expression of cytokeratin 14, a marker of normal mammary myoepithelial cells in the three species. Four different types of calponin‐positive cells were identified: (1) Type 1: cytokeratin‐14‐positive pre‐existing myoepithelial cells forming a continuous layer with images of focal disruptions; (2) Type 2: cytokeratin‐14‐positive isolated nests of fusiform, polygonal or round cells without atypia; (3) Type 3: cytokeratin‐14‐positive atypical cells indistinguishable from non‐reactive atypical cells, which should have never been detected in haematoxylin and eosin‐stained sections and (4) Type 4: cytokeratin‐14‐negative stromal fusiform cells around the neoplastic growth or cell nests, identified as myofibroblasts. Calponin‐negative and cytokeratin‐14‐positive atypical neoplastic cells were observed in three canine, 28 feline and two human carcinomas. The latter were indicative of altered expression of high‐molecular‐weight cytokeratins in luminal epithelial‐type simple carcinomas. Our findings show that calponin is a good marker of myoepithelial cell differentiation in feline, human and, particularly, canine simple carcinomas. The high number (six out of 15) of canine tumours with type 3 cells points to the need of both introducing calponin examination in the routine diagnostic schedule and performing further studies on its prognostic significance.     

Effects of a high milk intake during the pre‐weaning period on nutrient metabolism and growth rate in Japanese Black cattle

This study aimed to determine the effects of feeding an increased volume of high‐fat milk during the early post‐natal life on metabolite concentrations in the blood, the expression of key genes regulating intermediary metabolism in the skeletal muscles, and the rate of growth of Japanese Black cattle. All calves were fed a high‐fat milk replacer (crude protein, 26%; crude fat, 25.5%; total dissolved nitrogen, 116%). Control calves (n = 4) were nursed with 500 g milk replacer until 3 months of age, whereas calves in the experimental group (n = 4) were nursed with 1800 g milk replacer until 3 months, and then the volume was gradually reduced until 5 months. Body weight was significantly higher in the experimental group than in the control group at 7 months. Plasma glucose concentrations were significantly lower in the experimental group. Expression of glucose‐transporter‐4 messenger RNA (mRNA) was lower, whereas that of glucose transporter 1, cluster of differentiation 36, and carnitine palmitoyltransferase‐1b mRNA was significantly higher in the Longissimus thoracis of the experimental group. Nutritional status during early post‐natal life appears to strongly influence the growth rate and glucose and lipid metabolism in Japanese Black cattle.     

Influence of inflammation and coat type on oestrogen receptor immunohistochemistry

The purpose of this study was to evaluate oestrogen receptor alpha staining in a variety of breeds and skin conditions. The influence of inflammation and coat type on the presence and intensity of oestrogen receptor alpha staining was evaluated. Approximately 1700 haematoxylin and eosin (H&E)-stained slides of skin biopsies were screened for presence of small hair bulbs. Slides from 94 cases were submitted for oestrogen receptor alpha immunohistochemistry. H&E-stained skin biopsy tissues were examined for inflammation and hair follicle stages. Oestrogen receptor alpha staining characteristics of telogen follicles, flame follicles, large anagen bulbs, small hair bulbs and early anagen hairs (capped bulbs) were recorded. To assess the influence of inflammation and coat type on oestrogen receptor staining of hair follicle types, chi-square tests, Fisher's exact tests and logistic regression models were performed. Slides were classified as inflammatory (65) and noninflammatory (29). There were no statistically significant differences in oestrogen receptor staining when comparing inflammatory to noninflammatory skin biopsies or skin biopsies from dogs with different coat types. A subset of 13 noninflammatory biopsies from alopecic skin was identified. There was a statistically significant increase in the number of flame follicles in this subset as compared to all others. Comparison of oestrogen receptor staining of hair follicle types from these biopsies and all other biopsies revealed a statistically significant increase in the number of mature telogen hair follicles stained in this subset. No statistical difference in staining of early follicle stages was noted. Therefore, the oestrogen receptor is unlikely to be the controlling factor for the transition from telogen to anagen in the dog.     

Effects at early stage of life of elevated milk replacer feeding on growth rate,plasma IGF‐I concentration and intestinal nutrient transporter expression in Holstein bull calves

In order to evaluate the effects of an elevated amount of modified milk replacer on body weight, daily gain, starter intake, plasma endocrine parameters and expression of nutrient transporters in small intestinal epithelia, Holstein bull calves (n = 24) were fed for 60 days either with the usual amount of 24% crude protein (CP) and 20% fat milk (CF) replacer (C group), or with a double amount of a modified milk replacer of 28% CP and 16% CF (E group). Body weight from D20 to D60 and daily gain before D40 was greater or tended to be greater for the E group than the C group. Plasma concentrations of insulin‐like growth factor‐1 (IGF‐I) and insulin were greater for the E group than the C group on D28 but not on D56, without changing plasma growth hormone levels. Gene expression for sodium‐dependent glucose transporter 1 and fatty acid translocase (CD36) was altered in day‐ and intestine‐dependent manners. From these findings, we conclude that an elevated intake of milk replacer given up to 40 days old is sufficient to enhance body weight, which may be associated with increased plasma IGF‐I concentrations, in Holstein bulls.     

Expression of Genes in the Canine Pre‐implantation Uterus and Embryo: Implications for an Active Role of the Embryo Before and During Invasion

The aim of the present study was to assess genes expressed in maternal uterine tissue and pre‐implantation embryos which are presumably involved in maternal recognition and establishment of canine pregnancy. For this purpose, 10 pregnant bitches were ovariohysterectomized between days 10 and 12 after mating. Four non‐pregnant bitches served as controls. Early pregnancy was verified by flushing the uterine horns with PBS solution. The collected embryos (n = 60) were stored deep‐frozen (?80°C). Uterine tissue was excised, snaps frozen in liquid nitrogen and homogenized using TRI Reagent. All embryos from one litter were thawed together and also homogenized in TRI Reagent. RT‐PCR was performed to prove mRNA expression of progesterone receptor, key enzymes of the prostaglandin synthesis pathway, selected growth factors, cytokines, immune cell receptors, major histocompatibility complex (MHC) and matrix‐metalloproteinases (MMP). Only pregnant uteri revealed the presence of mRNA for interferon (IFN)‐γ, IL‐4 and CD‐8, which resembles the milieu in humans and other mammalians. Similarly, in day 10 embryos, mRNA for transforming growth factor‐β, insulin‐like growth factor‐1,‐2, hepatocyte growth factor, leukaemia inhibitor factor, tumour necrosis factor‐α, interleukin‐1β,‐6,‐8, cyclooxygenase‐2, CD4⁺ cells, and MMP‐2 and ‐9 were detected, but not MHC‐I or ‐II. We therefore suppose that the canine embryo, like its human counterpart, actively initiates measures to prevent attacks from the maternal immune system to prepare its own adhesion, nidation, growth and further development.     

Comparison of Chondrogenic Potential in Equine Mesenchymal Stromal Cells Derived from Adipose Tissue and Bone Marrow

Objective— To compare the chondrogenic potential of adult equine mesenchymal stem cells derived from bone marrow (MSCs) or adipose tissue (ASCs). Study Design— In vitro experimental study. Animals— Adult Thoroughbred horses (n=11). Methods— BM (5 horses; mean [±SD] age, 4±1.4 years) or adipose tissue (6 horses; mean age, 3.5±1.1 years) samples were obtained. Cryopreserved MSCs and ASCs were used for pellet cultures in stromal medium (C) or induced into chondrogenesis±transforming growth factor‐3 (TGFβ₃) and bone morphogenic factor‐6 (BMP‐6). Pellets harvested after 3, 7, 14, and 21 days were examined for cross‐sectional size and tissue composition (hematoxylin and eosin), glycosaminoglycan (GAG) staining (Alcian blue), collagen type II immunohistochemistry, and by transmission electron microscopy. Pellet GAG and total DNA content were measured using dimethylmethylene blue and Hoechst DNA assays. Results— Collagen type II synthesis was predominantly observed in MSC pellets from Day 7 onward. Unlike ASC cultures, MSC pellets had hyaline‐like matrix by Day 14. GAG deposition occurred earlier in MSC cultures compared with ASC cultures and growth factors enhanced both MSC GAG concentrations (P<.0001) and MSC pellet size (P<.004) after 2 weeks in culture. Conclusion— Equine MSCs have superior chondrogenic potential compared with ASCs and the equine ASC growth factor response suggests possible differences compared with other species. Clinical Relevance— Elucidation of equine ASC and MSC receptor profiles will enhance the use of these cells in regenerative cartilage repair.     

Insulin resistance selectively alters cell-surface glucose transporters but not their total protein expression in equine skeletal muscle

Background: Insulin resistance (IR) has been widely recognized in humans, and more recently in horses, but its underlying mechanisms are still not well understood. The translocation of glucose transporter 4 (GLUT4) to the cell surface is the limiting step for glucose uptake in insulin‐sensitive tissues. Although the downstream signaling pathways regulating GLUT translocation are not well defined, AS160 recently has emerged as a potential key component. In addition, the role of GLUT12, one of the most recently identified insulin‐sensitive GLUTs, during IR is unknown. Hypothesis/Objectives: We hypothesized that cell‐surface GLUT will be decreased in muscle by an AS160‐dependent pathway in horses with IR. Animals: Insulin‐sensitive (IS) or IR mares (n = 5/group). Methods: Muscle biopsies were performed in mares classified as IS or IR based on results of an insulin‐modified frequently sampled IV glucose tolerance test. By an exofacial bis‐mannose photolabeled method, we specifically quantified active cell‐surface GLUT4 and GLUT12 transporters. Total GLUT4 and GLUT12 and AS160 protein expression were measured by Western blots. Results: IR decreased basal cell‐surface GLUT4 expression (P= .027), but not GLUT12, by an AS160‐independent pathway, without affecting total GLUT4 and GLUT12 content. Cell‐surface GLUT4 was not further enhanced by insulin stimulation in either group. Conclusions and Clinical Importance: IR induced defects in the skeletal muscle glucose transport pathway by decreasing active cell‐surface GLUT4.