Can a mock medication-taking learning activity enable pharmacy students to experience the range of barriers and facilitators to medication adherence? An analysis informed by the Theoretical Domains Framework and COM-B model.

Background: Pharmacy professionals are well-placed to provide medication adherence support to patients. The Capability, Opportunity, Motivation-Behaviour (COM-B) and Theoretical Domains Framework (TDF) are two complementary models previously applied to medication-taking behaviour. Understanding the patient-specific barriers and facilitators to adherence using psychological frameworks from the early stages of pharmacy education enables the design and delivery of effective interventions. Objectives: To examine whether a novel 'mock medicine' learning activity enabled students to experience the range of barriers and facilitators to medication adherence using the COM-B and TDF. Methods: A mock medicine activity was conducted with students at pharmacy schools in three universities in the UK, Norway, and Australia over one week. Percentage adherence was calculated for five dosing regimens; theoretical framework analysis was applied to map reflective statements from student logs to COM-B and TDF. Results: A total of 349 students (52.6%) returned completed logs, with high overall mean adherence (83.5%, range 0-100%). Analysis of the 277 (79.4%) students who provided reflective statements included barriers and facilitators that mapped onto one (9%), two (29%) or all three (62%) of the COM-B components and all fourteen TDF domains (overall mean = 4.04; Uni 1 = 3.72; Uni 2 = 4.50; Uni 3 = 4.38; range 1-8). Most frequently mapped domains were 'Environmental context and resources' (n = 199; 72%), 'Skills' (n = 186; 67%), 'Memory, attention and decision-making' (184; 66%) and 'Beliefs about capabilities' (n = 175; 63%). Conclusions: This is the first study to utilise both COM-B and TDF to analyse a proxy measure of medication adherence in pharmacy education. Data mapping demonstrated that students experienced similar issues to patients when prescribed a short course of medication. Importantly, all the factors influencing medication-taking reported by students were captured by these two psychological frameworks. Future educational strategies will involve students in the mapping exercise to gain hands-on experience of using these psychological constructs in practice.

An evaluation of central nervous system cross-contamination due to carcass splitting in commercial beef-packing plants.

Four experiments were conducted in commercial beef-packing facilities The objectives of these experiments were to: (i) determine and validate a carcass sampling technique and location to determine if central nervous system (CNS) cross-contamination exists/occurs; (ii) determine if residual CNS tissue contamination remains on splitting saws after sanitation procedures; (iii) determine the prevalence of CNS cross-contamination in commercial slaughter facilities; (iv) determine whether washing treatments reduce or eliminate CNS tissue presence in carcass-splitting saws; (v) determine the effectiveness of commercial spray-washing systems in removing CNS tissue from beef carcasses; and (vi) compare residual CNS tissue levels on the blade and in the housings of the Jarvis Buster IX and Buster IV carcass-splitting saws. CNS tissue remained, albeit at very low levels, in the housings and on the blades of carcass-splitting saws after carcass splitting and operational sanitation. Additionally, after splitting carcasses, CNS tissue remaining in the splitting saw housings and on saw blades was found to cross-contaminate subsequent carcasses during splitting. Most splitting saw operational sanitation procedures reduced the amount of CNS tissue remaining in the splitting saw housings and on splitting saw blades, but no treatment eliminated CNS tissue from either to levels below the detection limit of the assay (6 ng/100 cm2). Washing in carcass spray-washing cabinets at three of the five commercial beef-packing facilities reduced, but did not eliminate, presence of CNS tissue in the aitch bone area of carcasses. Carcass spray washing in cabinets at three of the five facilities reduced (P < 0.05) the concentration of CNS tissue in the fourth thoracic vertebra area. While extremely low concentrations of CNS tissue remained in the splitting saw housings, on the splitting saw blades, and on carcasses, it is unknown whether these levels would pose a human food safety risk because the exact amount of bovine spongiform encephalopathy-infected spinal cord capable of transmitting the disease to humans is dependent on the infectivity titer, which is not readily known.

Alternative cutting methods to minimize transfer of nervous system tissue during steak preparation from bone-in short loins.

Fresh beef products, such as steaks, may become contaminated with potential specified risk materials (SRMs), such as central nervous system tissue, during the fabrication of bone-in loin subprimals. The objective of this study was to evaluate current and alternative cutting methods that could be used to minimize the transfer of nervous system tissue (NST) tissue during preparation of steaks from bone-in short loins. Bone-in short loins were cut according to three methods. (i) Cutting method I-The vertebral column bones were removed prior to cutting the loin into steaks from the medial (vertebral column) to lateral (flank) side. (ii) Cutting method II--The loin was cut into steaks from the vertebral column side to the flank side prior to removal of the vertebral column bones. (iii) Cutting method III--The loin was cut into steaks from the flank side to the vertebral column side prior to removal of the vertebral column bones. Results indicated that surface areas along the vertebral column cutting line had detectable (0.10 and 0.22% NST/100 cm2) and, thus, higher potential SRM contamination than resulting steak surfaces or the cutting blade. Overall, there were no detectable (<0.10% NST/100 cm2) differences in NST contamination of steaks produced by the three cutting methods. Immunohistochemical evaluation of areas on excised and ground steak surfaces indicated that regardless of cutting method, there was generally "no" to "moderate" staining, suggesting that detectable (0.137 to 0.201% NST) contamination from these samples was most likely due to peripheral nerve detection. These results imply that steaks may be cut from bone-in short loins prior to removal of the vertebral column bones without affecting the transfer of NST to resulting steaks at concentrations <0.10% NST/100 cm2.

Verification of different glial fibrillary acidic protein (GFAP) analyses as accurate detectors of central nervous system tissue in advanced meat recovery (AMR) products.

A glial fibrillary acidic protein (GFAP) fluorescent enzyme linked immunosorbant assay (ELISA) was compared with an ELISA test kit for GFAP to determine the level of central nervous system (CNS) tissue in advanced meat recovery (AMR) products. The test kit results were highly correlated (r=0.975) with the fluorescent ELISA. Meat cuts and AMR were analyzed on site at 14 meat plants utilizing the test kits. In seven of the plants all AMR samples had less than 1 ng GFAP. Seven of the plants had greater than 1 ng GFAP in AMR samples. Development of proper process controls to eliminate inclusion of spinal cord in AMR materials should bring all values to less than 1 ng GFAP, a level slightly above background.

Cellular mechanisms by which oxytocin mediates ovine endometrial prostaglandin F2alpha synthesis: role of G(i) proteins and mitogen-activated protein kinases.

Oxytocin stimulates a rapid increase in ovine endometrial prostaglandin (PG) F2alpha synthesis. The overall objective of these experiments was to investigate the cellular mechanisms by which oxytocin induces endometrial PGF2alpha synthesis. The objective of experiment 1 was to determine whether G(i) proteins mediate oxytocin-induced PGF2alpha synthesis. Uteri were collected from four ovary-intact ewes on Day 14 postestrus. Caruncular endometrial explants were dissected and subjected to in vitro incubation. Pertussis toxin, an inhibitor of G(i) proteins, had no effect on the ability of oxytocin to induce PGF2alpha synthesis (P > 0.10). The objective of experiment 2 was to determine whether any of the three mitogen-activated protein kinases (MAPKs), extracellular signal regulated protein kinase (ERK1/2), c-Jun N-terminal/stress-activated protein kinase (JNK/SAPK), or p38 MAPK, mediate oxytocin-induced PGF(2alpha) synthesis. Eleven ovary-intact ewes were given an injection of oxytocin (10 IU; i.v.; n = 5) or physiological saline (i.v.; n = 6) on Day 15 postestrus. Uteri were collected 15 min after injection and caruncular endometrium was dissected. Endometrial homogenates were prepared and subjected to Western blotting. Membranes were probed for both total and phosphorylated forms of all three classes of MAPK. All classes of MAPK were detected in ovine endometrium, but oxytocin treatment had no effect on the expression of these proteins (P > 0.10). ERK1/2 was the only phosphorylated MAPK detected and its concentrations were higher in oxytocin-treated ewes (P < 0.01). The objective of experiment 3 was to further investigate the role of ERK1/2 during oxytocin-induced PGF2alpha synthesis. Uteri were collected from four ovary-intact ewes on Day 14 postestrus. Caruncular endometrial explants were dissected and subjected to in vitro incubation. PD98059, a specific inhibitor of ERK1/2 activity, blocked the ability of oxytocin to stimulate PGF(2alpha synthesis in a dose-dependent manner (P < 0.05). These results indicate that the ovine oxytocin receptor is not coupled to G(i) proteins. These results indicate that oxytocin induces phosphorylation of ERK1/2 and that this MAPK appears to mediate oxytocin-induced PGF2alpha synthesis in ovine endometrium.

The detection of central nervous system tissue on beef carcasses and in comminuted beef.

We report the development and validation of a fluorescent enzyme-linked immunosorbent assay (ELISA) for glial fibrillary acidic protein (GFAP), which can be used as a rapid and sensitive method to detect CNS tissue in meat products. The fluorometric assay is sensitive to 0.2 ng GFAP and has an intra-assay coefficient of variation (CV) of 2.0% and an interassay CV of 14.1%. Bovine spinal cord and brain demonstrate dose-response curves that are parallel to GFAP standards, whereas peripheral sciatic nerve and cervical ganglia also cross-react at high tissue levels. The use of another central nervous system marker, syntaxin 1-B, was not effective for neural tissue detection. Less than 1.0 ng GFAP per mg tissue was found on most beef subprimals and advanced meat recovery (AMR) product. Occasional samples contained higher levels of GFAP, probably because of contamination by the carcass-splitting saw, incomplete removal of the spinal cord, or a chance sampling of a major nerve. Further reduction of CNS content was facilitated by removal of the cervical vertebrae and the spinal canal prior to processing beef chuck bones through AMR equipment. The presence of GFAP was very low (0.037 ng/mg) in beef patties collected from major processors throughout the USA. The presence of normal sausage ingredients or heating the product to 80 degrees C for 60 min did not affect the detection of GFAP. Heating the product to 115 degrees C for 100 min eliminated the detectability of GFAP.

Improved recovery of insulin-like growth factors (IGFs) from bovine colostrum using alkaline diafiltration.

Studies to develop a rapid, bioprocess-compatible method to recover low-molecular-mass growth factors from bovine colostrum are reported. Defatted bovine colostrum was subjected to tangential-flow filtration (TFF) using two different filters [polyether sulphone (PES) and regenerated cellulose (RC)] at pH 5.8, pH 8.0 and pH 8.0+0. 01 M NaCl. Single-pass TFF at pH 5.8 using a 100 kDa RC filter resulted in the loss of approx. 90% of insulin-like growth factor I (IGF-I) to non-specific filter adsorption. Comparison of 30 kDa RC and PES filters under single-pass conditions showed that yields of IGF-1 and IGF-II were highest with RC filters. Yields of IGF-I and protein from both filter types were increased at pH 8.0 and were greatest for the 30 kDa RC filter. Effects of adding large diluent volumes continuously during TFF (diafiltration) were tested at pH 5. 0 and 8.0. The use of 10 diafiltrate vols. at pH 8.0 resulted in the recovery of 15-28% of colostral IGF-1 from the RC 30 kDa permeates, 2-4-fold greater than under acidic conditions. Yields of IGF-II (39.6%) were unaffected by pH and at least 97% of total protein was retained by the 30 kDa filter at pH 8.0. Denaturing SDS/PAGE analysis of the alkaline RC 30 kDa permeates demonstrated two major regions of stained proteins at 10-13 kDa and 17-19 kDa. Acidic TFF permeates contained additional stained proteins at approximately 90, 48 and 37 kDa. Isoelectric focusing of these samples demonstrated the presence of proteins with isoelectric points of 8.2 and 8.6. The current study demonstrates a one-step bioprocess-compatible technique for the recovery of low-molecular-mass polypeptides from bovine colostrum. By using alkaline diafiltration with RC filters TFF provided optimal recovery of IGF-1 from colostrum.

Potential for disruption of central nervous system tissue in beef cattle by different types of captive bolt stunners.

The application of pneumatic-powered air injection stunners (PPAISs), pneumatic-powered stunners (PPSs), and cartridge-fired stunners (CFSs) in commercial beef slaughter plants was evaluated to determine the extent of dissemination of central nervous system tissue. Fifteen beef slaughter plants in the western and central United States were visited to observe stunning methods and the condition of the hearts at postmortem inspection. As inspectors performed the normal opening of the hearts, the research observer evaluated the contents of the heart for the presence of clots and/or visible tissue segments in the right ventricle. In eight plants where PPAISs were used, 33% of hearts examined (n = 1,050) contained large clots in the right ventricles. In the four plants where CFSs were used, 1% of the hearts (n = 480) contained detectable clots. In three plants where the newly modified PPSs were used, 12% of the hearts (n = 450) contained detectable clots. Large segments of spinal cord were detected, collected, photographed, and confirmed histologically from two hearts in a plant that used a PPAIS. Most of the material was found in a single right ventricle and was composed of 10 to 13 cm segments of spinal cord.

An enzyme-linked immunosorbent assay for glial fibrillary acidic protein as an indicator of the presence of brain or spinal cord in meat.

The current methods to detect central nervous system (CNS) tissue in blood, lungs, or meat are cumbersome, time consuming, and costly. The objective of this study was to use glial fibrillary acidic protein (GFAP), which is restricted to the CNS, in an enzyme-linked immunosorbent assay (ELISA) for the detection of CNS tissue in blood and muscle from beef cattle. Bovine brain, cerebral cortex, spinal cord, sciatic nerve, diaphragm, blood clots, and other skeletal muscle were obtained from three animals at slaughter. The limit for detection of GFAP was approximately 1.0 ng and the standard curve was linear up to 40 ng. Tissue samples gave responses parallel to the GFAP standard, suggesting that standard and unknown samples were immunoreactively identical. No GFAP was detected in skeletal muscle (ground beef, shoulder clod, and diaphragm) and blood clots. Trace amounts (13.5 to 51 ng/mg) were present in sciatic nerve. In contrast, high levels of GFAP (55 to 220 microg/ mg) were present in spinal cord, cerebral cortex (17 microg/mg), and whole brain (9 to 55 microg/mg). In a storage study using two animals in two separate studies, immunoreactive GFAP was detectable for up to 8 days at 4 degrees C in all tissues containing neural elements. Thus, mixtures of muscle with spinal cord or brain retained almost 80% of their immunoreactivity after 8 days at 4 degrees C, while brain and spinal cord alone retained approximately 50% and 25%, respectively, of their initial activities. In a repeat experiment, 80 to 100% of the initial activity was retained in these tissues after 8 days at 4 degrees C. The results of the current study demonstrate that the GFAP ELISA provides a valid and repeatable method to detect CNS tissue contamination in meat.

Electromyographic evidence for a digastric reflex evoked by perioral stimuli in humans.

Electromyographic recordings (EMGs) were made using skin surface electrodes placed over the anterior digastric muscle in seven subjects. In every case, short bursts of electrical stimuli to the upper lip produced a response in the EMG that had a minimum latency of 62.0 +/- 10.8 ms (mean +/- SD). By contrast, no responses were seen when single pulse stimuli were applied. In 6/7 subjects, the minimum stimulation intensity that produced the reflex was described as being sharp or painful. In three additional experiments, single motor units were recorded within the digastric muscle using needle electrodes. In two of these experiments, there was evidence of reflex activity 60-110 ms after the application of painful electrical stimuli to the lip. These findings confirm that perioral stimuli can evoke a digastric reflex in humans and suggest that this reflex requires the summation that results from successive volleys of impulses in a large number of nociceptive afferent neurones.

Insulin-like growth factor (IGF)-I and -II and IGFBP secretion by ovine satellite cell strains grown alone or in coculture with 3T3-L1 preadipocytes.

The current study was designed to examine the effects of muscle and fat stem cell coculture on the secretion of insulinlike growth factor (IGF)-I and -II and IGF binding proteins (IGFBP) by these cells. Two sheep satellite cell strains with negligible or high potential for differentiation (10A and 0(1), respectively) were placed in coculture with 3T3-L1 preadipocytes using a filter support to separate the two cell types. Media conditioned by the cells grown alone or in coculture were analyzed for IGFs by RIA or IGFBPs by ligand blotting. The numbers of satellite cells and preadipocytes declined throughout the 5-d culture period, although coculture slowed the 3T3-L1 decline but hastened the satellite cell decline. The satellite cell strains and 3T3-L1 cells secreted small amounts of IGF-I (< or = 2 ng/ml) and IGF-II (< 10 ng/ml) over the 5-d culture period. Coculture did not increase the amount of IGF-I and -II in conditioned media. The lowly differentiating 10A cells secreted barely detectable amounts of the low molecular weight IGFBP-3 subunit (34 kDa), IGFBP-2 (28 kDa), and IGFBP-4 (18 kDa). Coculture of 10A and 3T3-L1 cells potentiated secretion of IGFBP-2 and -3. Strain 0(1), which readily differentiates, secreted high levels of both IGFBP-3 subunits (34 and 39 kDa) and IGFBP-2 (28 kDa), as well as significant amounts of the 18 kDa IGFBP-4. Coculture did not alter IGFBP secretion of 0(1) cells. This study showed that while IGF-I and -II levels in media conditioned by sheep satellite cell strains are low and relatively invariant, the intensity and complexity of IGFBP patterns increases with time in culture and with the potential for differentiation of the satellite cell strains. Coculture with preadipocytes appeared to potentiate IGFBP secretion while reducing satellite cell viability.

The use of silicone dentures for edentulous patients.

There are a number of occasions when a soft denture base will be of much more benefit to an edentulous patient than a conventional rigid prosthesis. This article discusses three such cases.

Removable partial denture production in Scotland.

Removable partial denture production in Scotland was studied by analyzing photographs taken of consecutive casts and dentures and reviewing prescriptions of 539 cases at a large dental laboratory. Of the casts, 70.4% were maxillary and 29.6% mandibular. The mean number of teeth present was 9.9 in the maxilla and 8.4 in the mandible. Of the partial dentures studied, 33.6% were acrylate dentures without any framework. None of these had occlusal rests, and 5.6% had clasps. Of the framework-retained maxillary dentures, the palatal strap (12.9%), horseshoe (15.8%), and anterior palatal bar (18.9%) designs were the most frequent. In the mandible, lingual bars (32.7%) and lingual plates (32.7%) predominated. Of the acrylic dentures, 89.3% were made without any instruction from the dentist. The corresponding figure for prostheses with cobalt-chromium frameworks was 15.0%.

Twitch interpolation in the assessment of the maximum force-generating capacity of the jaw-closing muscles in man.

The method of twitch interpolation was employed to study the maximum potential bite forces of humans. Transcutaneous electrical stimuli were applied to parts of one or both masseter muscles in eight volunteers while they bit with a variable but controlled isometric force on a unidirectional force transducer held between the anterior teeth. In all participants the twitch force produced by a single 1-ms pulse, of 25 50 mA intensity, was inversely and linearly related to the voluntary bite force. For each participant the slope of the regression between twitch force and bite force depended on the stimulus intensity and not on whether the stimulus was applied to one or both masseters. Extrapolation of the regression lines to zero twitch force showed that they converged towards a bite-force value that, for any given participant varied only a small amount between different stimulus intensities. For most participants this bite force lay above the maximum that they produced voluntarily: voluntary maximum bites ranged from 153 to 593 N, while the extrapolations predicted a narrower and higher potential range of 282-629 N. It was concluded that, for the masseters at least, there is often spare force-generating capacity which individuals are either unable or not prepared to utilize. This method is non-invasive and may help to define better the maximum bite-force potential of humans.

Changes in human jaw muscles with age and dental state.

The effects of ageing and dental state on the cross-sectional area and density of two jaw muscles, the masseter and medial pterygoid, were investigated using computed tomography. The study involved 84 male and 70 female subjects between the ages of 20 and 90 years. The cross-sectional area of both muscles showed a significant reduction with age; values for female subjects being found in the lower range of the distribution. When consideration was given to the presence or absence of a natural dentition, the cross-sectional area of both muscles in edentulous subjects showed a greater decrease throughout the age range studied. There was a significant decrease in the density of the muscles with increasing age. Previously, this has been interpreted to indicate a progressive increase in fat and fibrous tissue. Subject gender and the absence of teeth appeared to have little effect on this parameter. Changes in the cross-sectional area and density of these muscles would appear to be consistent with a general age related change of muscle tissue in the body as a whole and may specifically indicate a reduction in the masticatory forces which can be or are being utilised by ageing patients, many of whom have no remaining natural dentition.

The structure and function of the temporomandibular joint.

Although characterised by having a synovial membrane lining the nonarticulating surfaces within the joint capsule, in some ways the temporomandibular joint (TMJ) is an atypical joint. This paper highlights the differences between the TMJ and other movable joints with a description of the structure, innervation blood supply and musculature. Also included are details of how the TMJ moves--the effectors of movement and the various reflexes controlling movement of the joint.

Replacement complete dentures: no friends like old friends.

Provision of replacement complete dentures for an increasingly elderly population presents problems for the dentist. Reduced adaptability in the older person leads to difficulty in learning to use new appliances unless existing skills can be employed. It is therefore helpful to reproduce familiar features of a patient's old appliances, especially if these have been used for some years. This paper describes a method of treatment that provides both clinician and laboratory with the ability to achieve this objective, while retaining freedom to eliminate clinically-identified faults in the old appliances in the process of provision of the new. The technique depends on the substitution of conventional wax-rimmed record blocks with wax replicas of the old appliances. These replica record blocks lend themselves to any - major or minor - alteration during occlusion recording. They also incorporate a rigid baseplate and function as special trays.

Interaction of insulin-like growth factors (IGF) with isolated sheep hepatocytes. I. Binding and degradation of IGF-I.

The binding and degradation of 125I-hIGF-I by isolated sheep hepatocytes have been examined. Hepatocytes were isolated by collagenase perfusion of 32-55 kg wether lambs and were incubated at 20 or 37 C at pH 7.4 in a 95% O2/5% CO2 atmosphere. Maximal binding was obtained at 60 min and declined slightly over the following 60-min period at both 20 and 37 C. Degradation of 125I-hIGF-I by the hepatocytes was minimal with 10-12% degradation over a 120-min period at 37 C. The lysosomal inhibitors chloroquine (0.2 mM), leupeptin and ammonium chloride had no significant effects on 125I-hIGF-I degradation or binding. At 20 C (60-min incubation), half maximal inhibition of 125I-hIGF-I binding was obtained with 8.4 +/- 1.1 nM hIGF-II, 16 +/- 2.4 nM hIGF-I, 36 +/- 6.2 nM oIGF-II, and 60 +/- 5.9 nM oIGF-I. Ovine insulin (0.01-10 uM) had no effect on 125I-hIGF-I binding. These observations suggest that IGF-I binds to the type II IGF receptor. The low molecular weight sheep serum IGF binding protein inhibited binding of 125I-hIGF-I to hepatocytes in a dose-dependent manner with half maximal inhibition occurring at 16.5 micrograms/ml, but did not affect IGF-I degradation. The current studies show that IGF-I interacts with ruminant hepatocytes via type II IGF receptors. The liver is not a major site of IGF-I degradation and the observed degradation is nonlysosomal and independent of receptor interaction.

Interaction of insulin-like growth factors (IGF) with isolated sheep hepatocytes. II. Binding, internalization and degradation of IGF-II.

This study examines the binding and degradation of IGF-II by the ovine liver. Binding and degradation of 125I-IGF-II to isolated hepatocytes was time, temperature and cell number dependent. Ovine and human IGF-II were 2-5 times more effective in inhibiting 125I-hIGF-II binding than were the IGF-I preparations. Insulin did not affect binding. Autoradiographs of 125I-hIGF-II affinity cross-linked to hepatocytes showed a major band of molecular weight 271,000 under reduced conditions. This band was eliminated by 100 nM hIGF-II or oIGF-II but not by excess hIGF-I, oIGF-I or insulin. The internalization of IGF-II was examined by treating the cells with trypsin or sodium acetate to remove surface-bound IGF-II. Both treatments showed that 20-25% of 125I-hIGF-II was internalized. Mannose-6-phosphate at 1, 2 and 4 mM enhanced the binding of 125I-hIGF-II to hepatocytes 3.5, 12.8 and 16.4%, respectively. The lysosomal inhibitors ammonium chloride, chloroquine and leupeptin had no effect on 125I-hIGF-II degradation or cell-associated radioactivity indicating a nonlysosomal pathway of degradation for 125I-hIGF-II in the ovine hepatocyte. The low molecular weight sheep serum IGF binding protein inhibited binding of 125I-hIGF-II in a dose-dependent manner but had no effect on degradation, which suggests that degradation of 125I-hIGF-II is independent of receptor interaction. These studies demonstrate that IGF-II binds to specific high affinity sites in sheep hepatocytes which display the characteristics of type II IGF receptors. A significant fraction of the receptor bound IGF-II is internalized but not degraded by these cells, which suggests that the biological actions of IGF-II may be exerted by an intracellular pathway in sheep hepatocytes.

Characterization of type II insulin-like growth factor (IGF) receptors in sheep liver plasma membranes.

Interactions of insulin-like growth factors (IGFs) from recombinant human and natural ovine sources with sheep liver plasma membranes have been studied. Total specific binding of 125I-hIGF-II (40%) to liver plasma membranes greatly exceeded that of 125I-hIGF-I (1.5%) after incubation at 20 C for 90 min. Binding of 125I-hIGF-II to the plasma membranes was dependent upon time, temperature and membrane concentration of the incubation. Binding of 125I-hIGF-II was only partially reversed by addition of 100 nM IGF-II (18%) or by dilution with excess buffer (36%). Competitive inhibition studies of 125I-hIGF-II binding demonstrated that IGF-II from ovine or recombinant human sources was more effective at inhibiting binding than ovine or human IGF-I. Insulin did not affect binding of 125I-hIGF-II. Plasma membranes were affinity cross-linked to 125I-IGF-II followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis in the presence and absence of the reducing agent dithiothreitol. Following autoradiography, radioactive bands were localized at 274,000 Mr and 210,000-215,000 Mr in the presence and absence of reducing agent, respectively. This pattern was unaffected by 100 nM human or ovine IGF-I or 1,000 nM insulin, but coincubation with 100 nM human or ovine IGF-II eliminated the radioactive band. These data indicate that an IGF-II specific receptor is present in sheep liver plasma membranes which has characteristics similar to those of nonruminant Type II receptors.