• View in gallery

    Photomicrographs of sections of pancreas from an adult (A) and a neonatal (B) camelid after immunostaining specific for GLUT-2 molecules. Darkly (red) stained cells contain high amounts of immunoreactive GLUT-2. Note the many individual stained cells (open arrows) that are not associated with the islets of Langerhans (closed arrows). Bar = 50 μm.

  • View in gallery

    Photomicrograph of a section of brain from a neonatal camelid after immunostaining specific for GLUT-1 molecules. The darkly (red) stained cells (solid arrows) represent vascular endothelium, and the intensity of staining reflects high amounts of immunoreactive GLUT-1. Bar = 100 μm.

  • View in gallery

    Photomicrograph of a section of pancreas from an adult camelid after immunostaining specific for GLUT-2 molecules. The darkly (red) stained cells in the center of the image are within a pancreatic islet, and the intensity of staining reflects high amounts of immunoreactive GLUT-2. Bar = 20 μm.

  • 1

    Gould GW, Holman GD. The glucose transporter family: structure, function and tissue specific expression. Biochem J 1993;295:329341.

  • 2

    Torres SH. Obesity, insulin resistance and skeletal muscle characteristics. Acta Cient Venez 1999;50:3441.

  • 3

    Dahlborn K, Benlamlih S, Wallsten C, et al.Glucose regulation in the camel. In: Allen WR, Higgins AJ, Mayhew IG, eds. Proceedings of the 1st International Camel Conference. Newmarket, UK: R&D Publications, 1992;414415.

    • Search Google Scholar
    • Export Citation
  • 4

    Elmahdi B, Sallmann HP, Fuhrmann H, et al.Comparative aspects of glucose tolerance in camels, sheep, and ponies. Comp Biochem Physiol A Physiol 1997;118:147151.

    • Search Google Scholar
    • Export Citation
  • 5

    Cebra CK, Tornquist SJ, Van Saun RJ, et al.Glucose tolerance testing in llamas and alpacas. Am J Vet Res 2001;62:682686.

  • 6

    Ommaya AK, Atwater I, Yañez A, et al.Lama glama (the South American camelid, llama): a unique model for evaluation of xenogenic islet transplants in a cerebral spinal fluid driven artificial organ. Transplant Proc 1995;27:33043307.

    • Search Google Scholar
    • Export Citation
  • 7

    Cebra CK, McKane SA, Tornquist SJ. Effects of exogenous insulin on glucose tolerance in alpacas. Am J Vet Res 2001;62:15441547.

  • 8

    Arraya AV, Atwater I, Navia MA, et al.Evaluation of insulin resistance in two kinds of South American camelids: llamas and alpacas. Comp Med 2000;50:490494.

    • Search Google Scholar
    • Export Citation
  • 9

    Cebra CK. Hyperglycemia, hypernatremia, and hyperosmolarity in 6 neonatal llamas and alpacas. J Am Vet Med Assoc 2000;217:17011704.

  • 10

    Garry F. Clinical pathology of llamas. Vet Clin North Am Food Anim Pract 1989;5:5570.

  • 11

    Stehman SM, Morris LI, Weisensel L, et al.Case report: picornavirus infection associated with abortion and adult onset diabetes mellitus in a herd of llamas, in Proceedings. 40th Annu Meet Am Assoc Vet Lab Diagn 1997;43.

    • Search Google Scholar
    • Export Citation
  • 12

    Khatim MS, Gumaa KA, Petersson B, et al.The structure and hormone content of the endocrine pancreas of the one-humped camel (Camelus dromedarius). Anat Anz 1985;159:181186.

    • Search Google Scholar
    • Export Citation
  • 13

    Adeghate E. Immunohistochemical identification of pancreatic hormones, neuropeptides and cytoskeletal proteins in pancreas of the camel (Camelus dromedarius). J Morphol 1997;231:185193.

    • Search Google Scholar
    • Export Citation
  • 14

    Abdel-Wahab MF, Abdel-Moneim Hussein M, Abdo MS. Preparation and purification of 131I-labelled insulin from the pancreas of domestic animals. Endokrinologie 1968;53:128135.

    • Search Google Scholar
    • Export Citation
  • 15

    al-Attas OS. Comparative studies on the major features of insulin receptors in mammalian and non-mammalian liver membranes. Comp Biochem Physiol B 1989;93:125133.

    • Search Google Scholar
    • Export Citation
  • 16

    Zhao FQ, Glimm DR, Kennelly JJ. Distribution of mam-malian facilitative glucose transporter messenger RNA in bovine tissues. Int J Biochem 1993;25:18971903.

    • Search Google Scholar
    • Export Citation
  • 17

    Snedecor GW, Cochran WG. Statistical methods. 7th ed. Ames, Iowa: Iowa State University Press, 1980;144.

  • 18

    Fowler ME. Digestive system. In: Medicine and surgery of South American camelids. 2nd ed. Ames, Iowa: Iowa State University Press, 1998;327328.

    • Search Google Scholar
    • Export Citation
  • 19

    Pearson EG, Snyder SP. Pancreatic necrosis in New World camelids: 11 cases (1990–1998). J Am Vet Med Assoc 2000;217:241244.

  • 20

    Hamir AN, Habecker PL, Tillman C. Pancreatic necrosis in a llama. Vet Rec 1998;142:644645.

  • 21

    Hazelwood RL. Embryology and anatomical organization of the vertebrate pancreas. In: The endocrine pancreas. Englewood Cliffs, NJ: Prentice-Hall Inc, 1989;915.

    • Search Google Scholar
    • Export Citation
  • 22

    Xu RJ, Wang T, Zang SH. Functional structure and growth of the pancreas in postnatal growing animals. In: Pierzynowski SG, Zabielski R, eds. Biology of the pancreas in growing animals. New York: Elsevier Press, 1999;1525.

    • Search Google Scholar
    • Export Citation
  • 23

    Bonner-Weir S, Like AA. A dual population of islets of Langerhans in bovine pancreas. Cell Tissue Res 1980;206:157170.

  • 24

    Jackson HD, VanDewark SD, VanVleet JF. Blood chemical and pancreatic histologic alterations in alloxan-diabetic ewes and their fetuses. Am J Vet Res 1970;31:15771587.

    • Search Google Scholar
    • Export Citation
  • 25

    Guillam MT, Hümmler E, Schaerer E, et al.Early diabetes and abnormal postnatal pancreatic islet development in mice lacking Glut-2. Nat Genet 1997;17:327330.

    • Search Google Scholar
    • Export Citation
  • 26

    Zisman A, Peroni OD, Abel ED, et al.Targeted disruption of the glucose transporter 4 selectively in muscle causes insulin resistance and glucose intolerance. Nat Med 2000;6:924928.

    • Search Google Scholar
    • Export Citation
  • 27

    Graziotti GH, Rios CM, Rivero JL. Evidence for three fast myosin heavy chain isoforms in type II skeletal muscle fibers in the adult llama (Lama glama). J Histochem Cytochem 2001;49:10331044.

    • Search Google Scholar
    • Export Citation
  • 28

    Smerdu V, Karsch-Mizrachi I, Campione M, et al.Type IIx myosin heavy chain transcripts are expressed in type IIb fibers of human skeletal muscle. Am J Physiol 1994;267:C1723C1728.

    • Search Google Scholar
    • Export Citation
  • 29

    Talmadge RJ, Grossman EJ, Roy RR. Myosin heavy chain composition of adult feline (Felis catus) limb and diaphragm muscles. J Exp Zool 1996;275:413420.

    • Search Google Scholar
    • Export Citation
  • 30

    Tanabe R, Muroya S, Chikuni K. Sequencing of the 2a, 2x, and slow isoforms of the bovine myosin heavy chain and the different expression among muscles. Mamm Genome 1998;9:10561058.

    • Search Google Scholar
    • Export Citation

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Microanatomic features of pancreatic islets and immunolocalization of glucose transporters in tissues of llamas and alpacas

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  • 1 Department of Clinical Sciences, College of Veterinary Medicine, Oregon State University, Corvallis, OR 97331-4802.
  • | 2 Department of Biomedical Sciences, College of Veterinary Medicine, Oregon State University, Corvallis, OR 97331-4802.
  • | 3 Department of Biomedical Sciences, College of Veterinary Medicine, Oregon State University, Corvallis, OR 97331-4802.

Abstract

Objective—To describe the microanatomic features of pancreatic islets and the immunohistochemical distribution of glucose transporter (GLUT) molecules in the pancreas and other tissues of New World camelids.

Animals—7 healthy adult New World camelids, 2 neonatal camelids with developmental skeletal abnormalities, and 2 BALB/c mice.

Procedure—Samples of pancreas, liver, skeletal muscle, mammary gland, brain, and adipose tissue were collected postmortem from camelids and mice. Pancreatic tissue sections from camelids were assessed microscopically. Sections of all tissues from camelids and mice (positive control specimens) were examined after staining with antibodies against GLUT-1, -2, -3, and -4 molecules.

Results—In camelids, pancreatic islets were prominent and lacked connective tissue capsules. Numerous individual endocrine-type cells were visible distant from the islets. Findings in neonatal and adult tissues were similar; however, the former appeared to have more non–islet-associated endocrine cells. Via immunostaining, GLUT-2 molecules were detected on pancreatic endocrine cells and hepatocytes in camelids, GLUT-1 molecules were detected on the capillary endothelium of the CNS, GLUT-3 molecules were detected throughout the gray matter, and GLUT-4 molecules were not detected in any camelid tissues. Staining characteristics of neonatal and adult tissues were similar.

Conclusions and Clinical Relevance—In New World camelids, microanatomic features of pancreatic islets are similar to those of other mammals. Data suggest that the poor glucose clearance and poor insulin response to hyperglycemia in adult camelids cannot be attributed to a lack of islet cells or lack of GLUT molecules on the outer membrane of those cells.

In virtually all mammalian cells, most if not all glucose uptake from the blood is achieved by members of the GLUT family of molecules.1 The different members of this family have different characteristics with regard to affinity, preferred substrate (glucose or another sugar), directionality (preference for uptake, export, or a concentration-dependent process), and responsiveness to hormonal stimuli, which appear to be based on the host tissue's need for glucose. Thus, the characteristics of glucose transport in a specific tissue can be estimated by determining which GLUT molecules populate the outer membranes of its cells.

Of the more common GLUT molecules, insulinindependent transporters can be widely distributed (eg, GLUT-1 molecules) or concentrated on tissues with important roles in glucose synthesis or regulation (eg, GLUT-2 molecules located on pancreatic beta cells, hepatocytes, and renal tubular cells) or high glucose requirements (eg, GLUT-3 molecules located on nervous tissue).1 Because these molecules represent constitutive uptake of glucose to provide basal requirements, their numbers and activity are fairly constant in these tissues. Their absence or inactivity would likely result in severe tissue dysfunction before or simultaneous to any noticeable effect on glucose clearance.

The insulin-dependent GLUT-4 molecule is most common on tissues with glucose requirements that vary considerably with activity (eg, skeletal muscle) or those in which the rate of glucose uptake is altered considerably in response to availability (eg, skeletal muscle and adipose tissue).1 Except in instances of overt obesity, skeletal muscle is a greater user of systemically available glucose, compared with adipose tissue.2 Thus, slow glucose clearance can often be linked directly or indirectly to slow glucose uptake by skeletal muscle.

The rate of glucose clearance in camelids is slower than rates in many other domestic mammals.3–8 This may be attributable in part to a smaller insulin response to exogenous glucose and in part to peripheral insulin resistance in camelids. The smaller insulin response could potentially be a result of a lack of insulin-producing beta islet cells, a lack of GLUT molecules on these cells (a function ascribed to GLUT-2 in other species), a defect in intracellular signaling, or some other defect in intracellular insulin synthesis or release. Peripheral insulin resistance could be attributable to a lack or failure of insulin receptors or a lack of insulin-sensitive GLUT-4 molecules, among other causes. Slow glucose clearance may be contributory to some of the hyperglycemic conditions detected in camelids, including hyperosmolar disorder,9 stress hyperglycemia,10 and persistent hyperglycemia11 (a diabetes mellitus–like condition).

Some of these aspects of glucose metabolism have been investigated in camels, which in evolutionary terms are the closest relatives to New World camelids.

The density and distribution of beta islet cells within the pancreas of camels appear to be similar to those of other animals,12 except that the pancreas of camels has additional isolated insulin-immunoreactive cells that are not associated with the islets.13 Also, camels appear to have insulin14 and an insulin receptor15 with morphologic and functional characteristics similar to those of other mammalian species. To our knowledge, the microscopic anatomic features of the pancreas have not been examined in New World camelids and the distribution of GLUT molecules has not been assessed in any species of camelid.

Tissue distribution of GLUT molecules can be examined via immunohistochemistry, western blot analysis, or identification of mRNA. Although camelidspecific anti-GLUT antibodies and probes are not available for these investigations, GLUT molecules in most species have relative homology and cross-reactions between species in immunologic and cDNA-based assays are common.1 Compared with other methods of GLUT detection, immunohistochemistry has the advantages that it only detects the transporter molecule itself, rather than untranslated mRNA (which may be present in tissues that do not express the transporter protein),16 and can be performed on fixed tissues. The purpose of the study reported here was to describe the microanatomic features of pancreatic islets and the immunohistochemical distribution of GLUT molecules in various tissues of New World camelids. By doing so, it was our intention to attempt to clarify the mechanisms of slow glucose clearance in camelids. We hypothesized that New World camelids would have pancreatic microanatomic features similar to those of camels and other species, with a lack of GLUT-2 molecules on islet cells (resulting in their poor insulin response to hyperglycemia) and a lack of GLUT-4 molecules on skeletal muscle cells (resulting in their peripheral insulin resistance).

Materials and Methods

Tissue samples

This study was approved by the Institutional Animal Care and Use Committee of Oregon State University. Seven healthy New World camelids (4 llamas and 3 alpacas), 2 neonatal camelids (1 llama and 1 alpaca) with developmental skeletal abnormalities, and 2 BALB/c mice were used in the study. The neonatal camelids had been euthanatized because of poor prognosis for life, whereas the healthy camelids and mice had been euthanatized as control animals in unrelated studies. Samples of pancreas (1 sample from each pole of the organ in adult camelids and 1 randomly selected sample of pancreas from crias and mice), liver, skeletal muscle, mammary gland, brain, and adipose tissue were collected from all animals. All tissues were fixed in neutral-buffered 10% formalin for 48 hours and embedded in paraffin blocks.

For examination of the microanatomic features of pancreatic tissue in camelids, one 7-μm-thick tissue section from each sample was mounted on a glass slide and stained with H&E. Entire tissue sections were assessed microscopically for the presence of lesions prior to more detailed examination of the endocrine component of the tissue. In sections of each pole of the pancreas from adults, the total number of islets of Langerhans identified in 10 randomly selected 25X fields of view was determined; in the pancreatic section from each cria, 10 randomly selected 25X fields of view were similarly examined. In all instances, islets at the edge of the field of view were included in the count. The size (maximum width) of each islet included in the count was also categorized (as ≤ 200 μm or > 200 μm). Each pole of the pancreas was then examined again, and the total number of endocrine cells per islet observed in each of 5 randomly selected islets was counted. Random selection consisted of blindly turning both control knobs of the microscope and choosing the islet closest to the center of the new field. Mouse tissues were not examined in this manner.

Immunohistochemistry was performed by use of the avidin-biotin-peroxidase method. Briefly, tissue sections were deparaffinized in xylene, rehydrated in 100% ethanol, washed in an endogenous enzyme blockera in 100% ethanol for 5 minutes, and then washed twice in 95% ethanol and once in automation bufferb diluted 1:9 in distilled water. High-temperature antigen retrievalc was performed on section slides in a pressure cooker for 5 minutes, after which the slides were allowed to cool to room temperature (approx 20°C) over a period of 20 minutes. The slides were then washed 3 times in automation buffer followed by digestion with proteinase solutiond at room temperature for 5 minutes. The slides were washed in the diluted automation buffer 3 more times, loaded into a slide rack, rinsed, and blotted with buffer to ensure good capillary flow.

Tissue sections were blocked for 20 minutes in normal goat serum diluted with automation bufferb and blotted dry. Primary antibodies (antibodies against GLUT-1, -2, -3, and -4 molecules)e-h were applied, and sections were incubated at 4°C overnight (approx 16 hours). These antibodies were commercially available products and were developed in rabbits against the extracellular domains of rodent GLUT molecules. Antibodies were used at a dilution of 1:200, except for anti–GLUT-4 antibody, which was used at a dilution of 1:400 because of intense staining of the mouse tissues at the 1:200 dilution. Rabbit IgG at a dilution of 1:200 in 2% normal goat serum was used as an isotype control specimen, and BALB/c mouse tissues were used as positive control specimens. After incubation, the slides were washed and blotted 6 times and the biotinylated secondary antibodyi was applied for a 30-minute incubation at room temperature. Section slides were washed in the diluted automation buffer solution and incubated with peroxidase complexi for 30 minutes at room temperature. Nova Redi was used as the chromagen, and Gill hematoxylin (diluted 1:10) was used as the counterstain. Sections were examined via light microscopy.

Data analysis

Islet counts and number of cells within islets were compared between the caudal and cranial pancreatic poles of adult camelids by use of the Mann-Whitney U test.17 Results were considered significant at a value of P < 0.05.

Results

Pancreatic tissue sections from only 1 adult llama had histologic evidence of disease. The pathologic changes consisted of mild multifocal peripancreatic fat necrosis in the section from each pole, with a few small aggregates of multinucleate giant cells and fibroblasts in the adipose tissue. Glandular and endocrine elements appeared normal. This llama was included in subsequent analyses because of the mild nature of the changes.

Islets were easily located in all sections of pancreatic tissue. Additionally, scattered individual cells with an endocrine morphology (round to polygonal cells with a lightly eosinophilic, nongranular cytoplasm vs the typical columnar to polygonal exocrine cells with a granular cytoplasm) were visible between exocrine acini in all sections (Figure 1). Islets were generally located within the lobules of exocrine tissue, but 2 islets were identified in the interlobular connective tissue of 1 cria. Most islets were round, but oval and irregular shapes were also observed. There was no distinct connective tissue capsule around any islet, and many of the larger islets were traversed by large capillaries. Endocrine cell shapes were mainly polygonal, but columnar cells were often visible as a palisade along capillaries. Differential cytoplasmic staining of cells along the outer rim of many islets suggested a nonhomogeneous cell population.

Figure 1—
Figure 1—

Photomicrographs of sections of pancreas from an adult (A) and a neonatal (B) camelid after immunostaining specific for GLUT-2 molecules. Darkly (red) stained cells contain high amounts of immunoreactive GLUT-2. Note the many individual stained cells (open arrows) that are not associated with the islets of Langerhans (closed arrows). Bar = 50 μm.

Citation: American Journal of Veterinary Research 67, 3; 10.2460/ajvr.67.3.524

The total number of pancreatic islets was determined in twenty 25X fields of view (cranial plus caudal pole sections) for each adult and in ten 25X fields of view for each neonatal camelid. The median number of islets/20 fields in adult camelids was 18 (range, 7 to 21 islets); islets were more numerous in the caudal lobe of the pancreas (median, 9 islets; range, 4 to 17 islets) than in the cranial lobe (median, 5 islets; range, 3 to 11 islets). The difference in islet counts between pancreatic poles in adults was not significant. Islet size in adult camelids ranged from 25 to 400 μm; the maximum width was < 200 μm in 89 of 94 islets examined.

The number of cells comprising individual islets in adults ranged from 5 to 198, but most contained < 50 cells. Median islet cell counts between pancreatic poles in adult camelids were not significantly different; there were 18 cells/islet in the cranial pole and 31 cells/islet in the caudal pole. In the 10 fields examined in the sections from each of the 2 neonatal camelids, the total number of islets encountered was 11 and 15, with all but 2 of these being < 200 μm wide. The largest was 250 μm wide. Considerable variation in the number of cells per islet was also detected in sections from the crias.

Anti–GLUT-1 antibody stained the vascular endothelium of the CNS intensely (Figure 2). Small amounts were visible on the capillary endothelium between adipocytes and myocytes, in the gray matter of the CNS, and on large vessels of the liver. Staining patterns were similar to those observed in mouse tissues.

Figure 2—
Figure 2—

Photomicrograph of a section of brain from a neonatal camelid after immunostaining specific for GLUT-1 molecules. The darkly (red) stained cells (solid arrows) represent vascular endothelium, and the intensity of staining reflects high amounts of immunoreactive GLUT-1. Bar = 100 μm.

Citation: American Journal of Veterinary Research 67, 3; 10.2460/ajvr.67.3.524

In contrast, anti–GLUT-2 antibody stained the sinusoidal membranes of hepatocytes and pancreatic islets intensely (Figure 3). Isolated single pancreatic cells not associated with the islets also had strong affinity for this stain (Figure 1). Compared with findings in adult camelid tissues, pancreatic tissues from neonatal camelids appeared to have fewer islets of high cell density and more single stained cells. Staining specific for GLUT-2 was not detected in any other tissues from adult and neonatal camelids. Staining patterns were similar to those observed in mouse tissues.

Figure 3—
Figure 3—

Photomicrograph of a section of pancreas from an adult camelid after immunostaining specific for GLUT-2 molecules. The darkly (red) stained cells in the center of the image are within a pancreatic islet, and the intensity of staining reflects high amounts of immunoreactive GLUT-2. Bar = 20 μm.

Citation: American Journal of Veterinary Research 67, 3; 10.2460/ajvr.67.3.524

Anti–GLUT-3 antibody lightly and diffusely stained the gray matter of CNS tissue from adult and neonatal camelids. The intensity of this staining was markedly weaker than the anti–GLUT-3 antibody staining detected in mouse CNS tissue; the staining intensity was also markedly weaker than the anti–GLUT-1 antibody staining of the same tissue from these animals. No non-CNS tissues from mice or camelids had affinity for this stain.

In all camelid tissues examined, GLUT-4–specific staining was not detected. Among mouse tissues, anti–GLUT-4 antibody staining was faint throughout the skeletal muscle sections.

Discussion

Macroscopically, the anatomic features of the pancreas of New World camelids are generally similar to those of other mammals; the pancreas is a flattened, lobulated organ that has cranial (head) and caudal (tail) lobes. The pancreatic duct joins the bile duct about 3 cm proximal to the intestinal orifice.18

Reports of pathologic changes in the pancreas of camelids are rare. Severe peripancreatic fat necrosis has been described19,20 as a cause of vague clinical illness in llamas and alpacas. A similar but mild disease was detected microscopically in one of the llamas in the present study, although that llama had no clinical signs attributable to pancreatic malfunction.

In the camelids of the present study, islets of Langerhans were easily identified in both cranial and caudal lobes of the pancreas and appeared randomly located, with no relationship to the ductal system. The distribution of islets between the 2 lobes in adult camelids was not uniform, a feature common to the endocrine pancreatic tissue of many mammals.21,22 Overall, the caudal lobe of the pancreas was found to contain more islets, as is typical of many mammals. However, a more quantitative study12 performed previously revealed that the cranial lobe and body of the pancreas of the 1-humped camel (Camelus dromedarius) both contained slightly more total endocrine hormone activity than the caudal lobe. In our study, the pancreatic endocrine tissue of camelids was located within the exocrine tissue, as reported in camels, and the islets were rarely seen in the interlobular connective tissue. This is in contrast to pancreatic tissue from cattle, which appears to have a population of large islets within the interlobular tissue.23 Another common feature observed in the camelid pancreata was the presence of scattered individual endocrine cells between exocrine acini, which has also been observed in Old World camelids.12,13 These cells were identified in adults and neonatal camelids in our study.

In the present study, the maximum width of islets in the pancreatic tissue of adult camelids was extremely variable, ranging from 25 to 400 μm. Findings from 1-humped camels are similar.13 The distinctive dual population of islets described in cattle23 and fetal sheep24 could not be identified in the tissues examined in our study. Because large islets are reported to be more numerous in fetal pancreatic tissue and in the body of the pancreas, we cannot discount the possibility that such a dual population exists in New World camelids.

In comparison with findings in adult camelids, the pancreatic endocrine tissue of camelid neonates was easily located but the islets were often somewhat less discrete, probably (at least in part) because of the underdevelopment of the exocrine tissue of this age group.22 The size range of islets in neonatal camelids was similar to that detected in adults. Comparable camel data are not available.

Distributions of GLUT molecules were similar to those in other mammals, except for the low intensity of GLUT-3–specific staining of nervous tissue and the absence of GLUT-4–specific staining of skeletal muscle. Absence or difference in intensity of staining could have been a result of either lower numbers of transporter molecules in those camelid tissues or lower affinities of the primary antibodies for the transporter molecules of camelids. Among the primary antibodies used, only anti–GLUT-1 antibody has cross-reactivity to ruminant GLUT-1 moleculese–h and none have been evaluated in camelids.

The presence of GLUT-2 immunoreactivity on pancreatic islet cells suggested that low insulin production in camelids is not related to poor recognition of extracellular glucose concentrations by beta islet cells. Similar to clinically normal camelids, mice lacking this transporter molecule have low insulin production and poor glucose tolerance, but unlike the former, these mice also have evidence of increased fat mobilization and ketone production.25 Immunoreactivity is poorly quantitative and does not address transporter function, so we cannot completely exclude the possibility of impaired glucose transport across beta cell membranes; further investigations of total beta cell mass and intracellular function of those cells in camelids are warranted.

Although it could not be proved by findings of our study, it is an intriguing possibility that camelids have substantially fewer insulin-responsive transporters in their skeletal muscle, compared with several other mammalian species. This possibility is also supported indirectly by other data. Selective disruption of skeletal muscle GLUT-4 in mice results in insulin resistance and slow glucose clearance,26 which are characteristics of healthy camelids.3–8 Animals with skeletal muscle that contains a high proportion of type IIB fibers are prone to insulin resistance and obesity,2 and some camelid muscles contain > 80% type IIB fibers.27 Additionally, as many as half of all type IIB fibers in camelids contain the myosin heavy chain type IIB isomer, which is not present in many mammals including humans,28 cats,29 horses,27 and cattle.30 Potentially, the different populations of myosin heavy chain isomers result in different rates of glucose uptake in these species. We postulate that inhibition of muscle glucose uptake provides camelids with a survival advantage in the harsh natural environments where they thrive.

Poor peripheral glucose uptake spares glucose for vital survival functions, including lactation and gestation, the latter of which exceeds 11 months in camelids.

GLUT

Glucose transporter

a

Endo/Blocker, Biomeda, Foster City, Calif.

b

10X automation buffer, Biomeda, Foster City, Calif.

c

Antigen demasker reagent, Biomeda, Foster City, Calif.

d

Pronase reagent, Biomeda, Foster City, Calif.

e

Mouse/rat glucose transporter 1 antibody, Alpha Diagnostic International, San Antonio, Tex.

f

Rat glucose transporter 2 antibody, Alpha Diagnostic International, San Antonio, Tex.

g

Mouse glucose transporter 3 antibody, Alpha Diagnostic International, San Antonio, Tex.

h

Mouse glucose transporter 4 antibody, Alpha Diagnostic International, San Antonio, Tex.

i

Vector Universal Elite ABC kit, Vector Laboratories, Burlingame, Calif.

References

  • 1

    Gould GW, Holman GD. The glucose transporter family: structure, function and tissue specific expression. Biochem J 1993;295:329341.

  • 2

    Torres SH. Obesity, insulin resistance and skeletal muscle characteristics. Acta Cient Venez 1999;50:3441.

  • 3

    Dahlborn K, Benlamlih S, Wallsten C, et al.Glucose regulation in the camel. In: Allen WR, Higgins AJ, Mayhew IG, eds. Proceedings of the 1st International Camel Conference. Newmarket, UK: R&D Publications, 1992;414415.

    • Search Google Scholar
    • Export Citation
  • 4

    Elmahdi B, Sallmann HP, Fuhrmann H, et al.Comparative aspects of glucose tolerance in camels, sheep, and ponies. Comp Biochem Physiol A Physiol 1997;118:147151.

    • Search Google Scholar
    • Export Citation
  • 5

    Cebra CK, Tornquist SJ, Van Saun RJ, et al.Glucose tolerance testing in llamas and alpacas. Am J Vet Res 2001;62:682686.

  • 6

    Ommaya AK, Atwater I, Yañez A, et al.Lama glama (the South American camelid, llama): a unique model for evaluation of xenogenic islet transplants in a cerebral spinal fluid driven artificial organ. Transplant Proc 1995;27:33043307.

    • Search Google Scholar
    • Export Citation
  • 7

    Cebra CK, McKane SA, Tornquist SJ. Effects of exogenous insulin on glucose tolerance in alpacas. Am J Vet Res 2001;62:15441547.

  • 8

    Arraya AV, Atwater I, Navia MA, et al.Evaluation of insulin resistance in two kinds of South American camelids: llamas and alpacas. Comp Med 2000;50:490494.

    • Search Google Scholar
    • Export Citation
  • 9

    Cebra CK. Hyperglycemia, hypernatremia, and hyperosmolarity in 6 neonatal llamas and alpacas. J Am Vet Med Assoc 2000;217:17011704.

  • 10

    Garry F. Clinical pathology of llamas. Vet Clin North Am Food Anim Pract 1989;5:5570.

  • 11

    Stehman SM, Morris LI, Weisensel L, et al.Case report: picornavirus infection associated with abortion and adult onset diabetes mellitus in a herd of llamas, in Proceedings. 40th Annu Meet Am Assoc Vet Lab Diagn 1997;43.

    • Search Google Scholar
    • Export Citation
  • 12

    Khatim MS, Gumaa KA, Petersson B, et al.The structure and hormone content of the endocrine pancreas of the one-humped camel (Camelus dromedarius). Anat Anz 1985;159:181186.

    • Search Google Scholar
    • Export Citation
  • 13

    Adeghate E. Immunohistochemical identification of pancreatic hormones, neuropeptides and cytoskeletal proteins in pancreas of the camel (Camelus dromedarius). J Morphol 1997;231:185193.

    • Search Google Scholar
    • Export Citation
  • 14

    Abdel-Wahab MF, Abdel-Moneim Hussein M, Abdo MS. Preparation and purification of 131I-labelled insulin from the pancreas of domestic animals. Endokrinologie 1968;53:128135.

    • Search Google Scholar
    • Export Citation
  • 15

    al-Attas OS. Comparative studies on the major features of insulin receptors in mammalian and non-mammalian liver membranes. Comp Biochem Physiol B 1989;93:125133.

    • Search Google Scholar
    • Export Citation
  • 16

    Zhao FQ, Glimm DR, Kennelly JJ. Distribution of mam-malian facilitative glucose transporter messenger RNA in bovine tissues. Int J Biochem 1993;25:18971903.

    • Search Google Scholar
    • Export Citation
  • 17

    Snedecor GW, Cochran WG. Statistical methods. 7th ed. Ames, Iowa: Iowa State University Press, 1980;144.

  • 18

    Fowler ME. Digestive system. In: Medicine and surgery of South American camelids. 2nd ed. Ames, Iowa: Iowa State University Press, 1998;327328.

    • Search Google Scholar
    • Export Citation
  • 19

    Pearson EG, Snyder SP. Pancreatic necrosis in New World camelids: 11 cases (1990–1998). J Am Vet Med Assoc 2000;217:241244.

  • 20

    Hamir AN, Habecker PL, Tillman C. Pancreatic necrosis in a llama. Vet Rec 1998;142:644645.

  • 21

    Hazelwood RL. Embryology and anatomical organization of the vertebrate pancreas. In: The endocrine pancreas. Englewood Cliffs, NJ: Prentice-Hall Inc, 1989;915.

    • Search Google Scholar
    • Export Citation
  • 22

    Xu RJ, Wang T, Zang SH. Functional structure and growth of the pancreas in postnatal growing animals. In: Pierzynowski SG, Zabielski R, eds. Biology of the pancreas in growing animals. New York: Elsevier Press, 1999;1525.

    • Search Google Scholar
    • Export Citation
  • 23

    Bonner-Weir S, Like AA. A dual population of islets of Langerhans in bovine pancreas. Cell Tissue Res 1980;206:157170.

  • 24

    Jackson HD, VanDewark SD, VanVleet JF. Blood chemical and pancreatic histologic alterations in alloxan-diabetic ewes and their fetuses. Am J Vet Res 1970;31:15771587.

    • Search Google Scholar
    • Export Citation
  • 25

    Guillam MT, Hümmler E, Schaerer E, et al.Early diabetes and abnormal postnatal pancreatic islet development in mice lacking Glut-2. Nat Genet 1997;17:327330.

    • Search Google Scholar
    • Export Citation
  • 26

    Zisman A, Peroni OD, Abel ED, et al.Targeted disruption of the glucose transporter 4 selectively in muscle causes insulin resistance and glucose intolerance. Nat Med 2000;6:924928.

    • Search Google Scholar
    • Export Citation
  • 27

    Graziotti GH, Rios CM, Rivero JL. Evidence for three fast myosin heavy chain isoforms in type II skeletal muscle fibers in the adult llama (Lama glama). J Histochem Cytochem 2001;49:10331044.

    • Search Google Scholar
    • Export Citation
  • 28

    Smerdu V, Karsch-Mizrachi I, Campione M, et al.Type IIx myosin heavy chain transcripts are expressed in type IIb fibers of human skeletal muscle. Am J Physiol 1994;267:C1723C1728.

    • Search Google Scholar
    • Export Citation
  • 29

    Talmadge RJ, Grossman EJ, Roy RR. Myosin heavy chain composition of adult feline (Felis catus) limb and diaphragm muscles. J Exp Zool 1996;275:413420.

    • Search Google Scholar
    • Export Citation
  • 30

    Tanabe R, Muroya S, Chikuni K. Sequencing of the 2a, 2x, and slow isoforms of the bovine myosin heavy chain and the different expression among muscles. Mamm Genome 1998;9:10561058.

    • Search Google Scholar
    • Export Citation

Contributor Notes

Funded by the Oregon Agricultural Experiment Station. Digital photographic equipment was supplied by the Cell and Tissue Analysis Facilities and Services Core, Environmental Health Sciences Center, Oregon State University.

Dr. Cebra.