Pentoxifylline (PTX)--an alternative treatment in Graves' ophthalmopathy (inactive phase): assessment by a disease specific quality of life questionnaire and by exophthalmometry in a prospective randomized trial.

PURPOSE: To investigate the effect of pentoxifylline (PTX) in subjects with inactive Graves' ophthalmopathy (GO) through a specific quality of life (QOL) questionnaire and exophthalmometry readings. METHODS: Eighteen females were randomly divided in two groups. Group A (n=9) was treated with PTX 1200 mg orally/day for 6 months. Group B (n=9) received placebo during the initial 6 months and then PTX for another 6 months. Proptosis measurements were carried out every 3 months and a questionnaire graded from 0 to 10 according to the severity of the symptoms was performed at baseline and after placebo and PTX administration. RESULTS: At baseline, Group A questionnaire score values were 5.5 (median; range 3.5 to 8.0), and 5.0 after 6 months (3.0 to 6.0; p=0.01). In Group B, baseline values were not significantly different after 6 months of placebo: 6.0 (4.5 to 7.0) and 5.5 (4.5 to 7.0), respectively. However, a significant change was observed 6 months after PTX: 4.0 (2.0 to 5.0; p<0.001). Patients in Group A had a progressive improvement of proptosis during PTX: at baseline, 23 mm (median; range 20 to 32); after 3 months, 23 mm (18 to 30; p=0.02); and after 6 months, 23 mm (18 to 30; p=0.005). In Group B, proptosis remained stable during placebo: at baseline, 23 mm (21 to 25); after 3 months, 23 mm (20 to 25); and after 6 months, 23.5 mm (20 to 25). A significant change was observed after 3 and 6 months of PTX: 22 mm (19 to 24; p=0.0006) and 20.8 mm (17 to 25; p=0.0003), respectively. CONCLUSIONS: Pentoxifylline seems to improve the QOL of patients in the inactive phase of GO. The objective findings of the proptosis readings corroborate to suggest that PTX may be an effective and promising drug in the inactive phase of GO.

Pentoxifylline (PTX): an alternative treatment in Graves ophthalmopathy (inactive phase): assessment by a disease specific quality of life questionnaire and by exophthalmometry in a prospective randomized trial.

PURPOSE: To investigate the effect of pentoxifylline (PTX) in subjects with inactive Graves ophthalmopathy (GO) through a specific quality of life (QOL) questionnaire and exophthalmometry readings. METHODS: Eighteen females were randomly divided in two groups. Group A (n=9) was treated with PTX 1200 mg orally/day for 6 months. Group B (n=9) received placebo during the initial 6 months and then PTX for another 6 months. Proptosis measurements were carried out every 3 months and a questionnaire graded from 0 to 10 according to the severity of the symptoms was performed at baseline and after placebo and PTX administration. RESULTS: At baseline, Group A questionnaire score values were 5.5 (median; range 3.5 to 8.0), and 5.0 after 6 months (3.0 to 6.0; p=0.01). In Group B, baseline values were not significantly different after 6 months of placebo: 6.0 (4.5 to 7.0) and 5.5 (4.5 to 7.0), respectively. However, a significant change was observed 6 months after PTX: 4.0 (2.0 to 5.0; p<0.001). Patients in Group A had a progressive improvement of proptosis during PTX: at baseline, 23 mm (median; range 20 to 32); after 3 months, 23 mm (18 to 30; p=0.02); and after 6 months, 23 mm (18 to 30; p=0.005). In Group B, proptosis remained stable during placebo: at baseline, 23 mm (21 to 25); after 3 months, 23 mm (20 to 25); and after 6 months, 23.5 mm (20 to 25). A significant change was observed after 3 and 6 months of PTX: 22 mm (19 to 24; p=0.0006) and 20.8 mm (17 to 25; p=0.0003), respectively. CONCLUSIONS: Pentoxifylline seems to improve the QOL of patients in the inactive phase of GO. The objective findings of the proptosis readings corroborate to suggest that PTX may be an effective and promising drug in the inactive phase of GO. (Eur J Ophthalmol 2004; 14: 277-83).

Suppressors of cytokine signaling-1 and -6 associate with and inhibit the insulin receptor. A potential mechanism for cytokine-mediated insulin resistance.

Insulin resistance contributes to a number of metabolic disorders, including type II diabetes, hypertension, and atherosclerosis. Cytokines, such as tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6, and hormones, such as growth hormone, are known to cause insulin resistance, but the mechanisms by which they inhibit the cellular response to insulin have not been elucidated. One mechanism by which these agents could cause insulin resistance is by inducing the expression of cellular proteins that inhibit insulin receptor (IR) signaling. Suppressors of cytokine signaling (SOCS) proteins are negative regulators of cytokine signaling pathways, the expression of which is regulated by certain cytokines. SOCS proteins are therefore attractive candidates as mediators of cytokine-induced insulin resistance. We have found that SOCS-1 and SOCS-6 interact with the IR when expressed in human hepatoma cells (HepG2) or in rat hepatoma cells overexpressing the human IR. In SOCS-1-expressing cells, insulin treatment increases the extent of interaction with the IR, whereas in SOCS-6-expressing cells the association with the IR appears to require insulin treatment. SOCS-1 and SOCS-6 do not inhibit insulin-dependent IR autophosphorylation, but both proteins inhibit insulin-dependent activation of ERK1/2 and protein kinase B in vivo and IR-directed phosphorylation of IRS-1 in vitro. These results suggest that SOCS proteins may be inhibitors of IR signaling and could mediate cytokine-induced insulin resistance and contribute to the pathogenesis of type II diabetes.

Differential expression of suppressors of cytokine signalling genes in response to nutrition and growth hormone in the septic rat.

GH treatment during critical illness and sepsis may increase mortality. A family of negative regulators of cytokine signalling, the suppressors of cytokine signalling (SOCS), have been characterised. SOCS provide a mechanism for cross-talk between the cytokine receptors, including GH. Here, we have investigated the impact of nutrition and GH treatment on GH receptor, SOCS1, SOCS-2, SOCS-3 and cytokine-inducible SH2-containing protein (CIS) hepatic mRNA expression in a rat model of sepsis, caecal ligation and puncture (CLP). Four groups of rats were studied: control (food given ad libitum, n=7), CLP only (n=8), CLP and total parenteral nutrition (TPN) (n=9), and CLP, TPN and GH (n=10). CLP rats underwent surgery and 18 h later received saline or TPN or TPN+GH for 6 h before they were killed. Serum IGF-I levels were lower in all CLP groups (P<0.001). The combination of TPN and GH treatment increased IGF-I levels compared with the saline-treated CLP rats (P<0.01), but IGF-I levels remained lower than control animals (P<0.001). GH receptor and GH-binding protein expression in liver was reduced in animals subjected to CLP and was unaffected by nutrition or GH treatment. Hepatic SOCS-1 was detectable in normal rats, induced in all CLP animals but was unaffected by nutrition and GH. Hepatic SOCS-2 expression was difficult to detect in normal and CLP rats but was greatly induced in CLP rats treated with GH. Hepatic SOCS-3 expression was only just detectable in the control group but was elevated in all CLP groups and unaffected by nutrition and GH. Hepatic CIS expression was difficult to detect in normal rats, was not induced by CLP but was induced by both nutrition and GH. In conclusion, CLP induced low IGF-I levels associated with increased expression of SOCS-1 and SOCS-3, both of which are known to inhibit GH receptor signalling. GH induced SOCS-2 and CIS in the CLP rat despite resistance with respect to IGF-I generation, and parenteral feeding induced CIS in the CLP rat. Thus, there is potential for a complex interaction between GH and cytokine signalling at the level of SOCS expression whereby the inflammatory response may alter GH signalling and GH may influence the inflammatory response.

Suppressor of cytokine signaling (SOCS)-3 protein interacts with the insulin-like growth factor-I receptor.

SOCS proteins are a class of proteins that are negative regulators of cytokine receptor signaling via the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway. In a yeast two-hybrid screen of a human fetal brain library, we have previously identified SOCS-2 as a binding partner of the activated IGF-I receptor (IGFIR). To test whether or not SOCS-3 also binds to the IGFIR, we cloned human SOCS-3 by reverse transcription-polymerase chain reaction from human skeletal muscle mRNA. SOCS-3 mRNA was expressed in many human fetal and adult tissues and in some human cancer cell lines (Hela, A549 pulmonary adenocarcinoma and G361 human melanoma). We found that human SOCS-3 protein interacts directly with the cytoplasmic domains of the activated IGFIR and the insulin receptor (IR) in the yeast two-hybrid assay. In GST-SOCS-3 pull-down experiments using IGFIR from mammalian cells and in immunoprecipitation experiments in which IGFIR and FLAG-SOCS-3 were transiently expressed in human embryonic kidney 293 cells, we found that SOCS-3 interacts constitutively with IGFIR in vitro and in intact cells. Unlike SOCS-2, hSOCS-3 was phosphorylated on tyrosines in response to IGF-I addition to 293 cells. We conclude that SOCS-3 binds to the IGFIR and may be a direct substrate for the receptor tyrosine kinase.

Effect of combined treatment with calcitonin on bone densitometry of patients with treated hypothyroidism.

INTRODUCTION: Thyroid hormones (TH) may affect bone metabolism and turnover, inducing a loss of bone mass among hyperthyroid and in hypothyroid patients under hormone replacement treatment. Thyroid dysfunction leads to changes in the dynamics of parathyroid hormone (PTH) and calcitonin (CT) secretion. OBJECTIVE: The objective of the study was to determine the usefulness of CT as adjuvant therapy in the prevention of bone loss during the treatment of hypothyroidism. MATERIAL AND METHODS: We studied 16 female patients with recently diagnosed primary hypothyroidism, divided into two groups: group G1 (n = 8) submitted to treatment with thyroxine (L-T4), and Group 2 (n = 8) that, in addition to being treated with L-T4, received a nasal CT spray. All patients were submitted to determination of TSH, free T4, bone mineral densitometry (BMD) and total bone calcium (TBC) at the time of diagnosis, after 6 to 9 months of treatment, and after 12 months of treatment. RESULTS: No statistical significant differences were detected in either group between the total BMD values obtained for the femur and lumbar spine before and after treatment. However, group G1 presented a statistical significant TBC loss after 12 months of treatment compared to initial values. In contrast, no TBC loss was observed in the group treated with LT-4 in combination with CT, a fact that may suggest that CT was responsible for the lower bone reabsorption during treatment of hypothyroidism.

Effect of combined treatment with calcitonin on bone densitometry of patients with treated hypothyroidism

INTRODUCTION: Thyroid hormones (TH) may affect bone metabolism and turnover, inducing a loss of bone mass among hyperthyroid and in hypothyroid patients under hormone replacement treatment. Thyroid dysfunction leads to changes in the dynamics of parathyroid hormone (PTH) and calcitonin (CT) secretion. OBJECTIVE: The objective of the study was to determine the usefulness of CT as adjuvant therapy in the prevention of bone loss during the treatment of hypothyroidism. MATERIAL AND METHODS: We studied 16 female patients with recently diagnosed primary hypothyroidism, divided into two groups: group G1 (n=8) submitted to treatment with thyroxine (L-T4), and Group 2 (n=8) that, in addition to being treated with L-T4, received a nasal CT spray. All patients were submitted to determination of TSH, free T4, bone mineral densitometry (BMD) and total bone calcium (TBC) at the time of diagnosis, after 6 to 9 months of treatment, and after 12 months of treatment. RESULTS: No statistical significant differences were detected in either group between the total BMD values obtained for the femur and lumbar spine before and after treatment. However, group G1 presented a statistical significant TBC loss after 12 months of treatment compared to initial values. In contrast, no TBC loss was observed in the group treated with LT-4 in combination with CT, a fact that may suggest that CT was responsible for the lower bone reabsorption during treatment of hypothyroidism.

Thyroid suppression test with a single oral dose of levothyroxine in the diagnosis of functional thyroid autonomy.

OBJECTIVE: To propose a modified form of thyroid suppression test with use of a single oral dose of levothyroxine (35 mg/kg). METHODS: After a baseline scintigram, 23 patients with nodular goiter suspected of autonomous function (warm or hot nodules, subnormal or undetectable thyrotropin levels, or both findings) and 14 normal subjects underwent a repeated scintigram 4 days after administration of levothyroxine. We evaluated triiodothyronine (T(3)), free thyroxine, and thyrotropin before and on the first, second, third, fourth, and seventh days after administration of the individualized dose of levothyroxine. RESULTS: The 99th percentile of postsuppression uptake in normal subjects was determined, and an uptake >12.4%, a 131 I concentration restricted to the nodule, or both factors were adopted as the criteria for diagnosis of an autonomously functioning thyroid nodule. Twelve patients were considered to have autonomously functioning nodules, and 11 patients were considered to have nonautonomous nodules. Baseline thyrotropin levels in patients with autonomous nodules did not differ significantly from those in patients with nonautonomous nodules. No signs or symptoms of toxicity were detected during the test, but all study subjects had increased free thyroxine values, and seven had high levels of T(3). CONCLUSION: The thyroid suppression test with 35 mg/kg of levothyroxine is an effective method for the diagnosis of an autonomously functioning thyroid nodule, is nontoxic, and avoids the inaccurate use of the medication occasionally observed with T(3). Even sensitive methods of thyrotropin determination cannot replace this test in the evaluation of autonomous thyroid function.

Interaction of human suppressor of cytokine signaling (SOCS)-2 with the insulin-like growth factor-I receptor.

SOCS (suppressor of cytokine signaling) proteins have been shown to be negative regulators of cytokine receptor signaling via the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway. We have cloned a member of this family (hSOCS-2) by utilizing the insulin-like growth factor I receptor (IGF-IR) cytoplasmic domain as bait in a yeast two-hybrid screen of a human fetal brain library. The hSOCS-2 protein interacted strongly with the activated IGF-IR and not with a kinase negative mutant receptor in the two-hybrid assay. Mutation of receptor tyrosines 950, 1250, 1251, and 1316 to phenylalanine or deletion of the COOH-terminal 93 amino acids did not result in decreased interaction of the receptor with hSOCS-2 protein. hSOCS-1 protein also interacted strongly with IGF-IR in the two-hybrid assay. Glutathione S-transferase-hSOCS-2 associated with activated IGF-IR in lysates of mouse fibroblasts overexpressing IGF-IR. Human embryonic kidney cells (293) were transiently transfected with vectors containing IGF-IR and FLAG epitope-tagged hSOCS-2. After IGF-I stimulation, activated IGF-IR was found in anti-FLAG immunoprecipitates and, conversely, FLAG-hSOCS-2 was found in anti IGF-IR immunoprecipitates. Thus, hSOCS-2 interacted with IGF-IR both in vitro and in vivo. HSOCS-2 mRNA was expressed in many human fetal and adult tissues with particularly high abundance in fetal kidney and adult heart, skeletal muscle, pancreas, and liver. These results raise the possibility that SOCS proteins may also play a regulatory role in IGF-I receptor signaling.

Cloning of human p55 gamma, a regulatory subunit of phosphatidylinositol 3-kinase, by a yeast two-hybrid library screen with the insulin-like growth factor-I receptor.

We have used the yeast two-hybrid system to identify proteins that interact with the intracellular domain of the insulin-like growth factor-I receptor (IGFIR). In a search of a human fetal brain library we identified a cDNA encoding a protein that is the human homologue of mouse p55PIK, a regulatory subunit of phosphatidylinositol 3-kinase (hp55 gamma). The hp55 gamma protein interacts strongly with the activated IGFIR but not with the kinase-negative mutant receptor. hp55 gamma also interacts with the insulin receptor (IR) in the yeast two-hybrid system. The putative hp55 gamma protein is composed of a unique amino terminal region followed by a proline-rich motif and two Src homology 2 (SH2) domains, which are highly homologous to those in mouse p55PIK, rat p55 gamma, human p85 alpha and bovine p85 beta; it contains no SH3 domain. hp55 gamma mRNAs are expressed in most human fetal and adult tissues with particularly high abundance in adult testis. Splice variant(s) of hp55 gamma, one of which has a deletion of 36 amino acids at the amino terminus and another which has an insertion of 59 amino acids at position 256 between the SH2 domains, were also identified. A GST-hp55 gamma fusion protein interacts in vitro with both the activated IGFIR and IR derived from mammalian cells. Our findings suggest that hp55 gamma interacts with the IGFIR and IR and may be involved in PI 3-kinase activation by these receptors.

Prospective and controlled study of ophthalmopathy after radioiodine therapy for Graves' hyperthyroidism.

The effects of radioiodine (131I) therapy for hyperthyroidism on the ocular process of Graves' disease is controversial. In order to evaluate the outcome of ophthalmopathy after radioiodine therapy for thyrotoxicosis we studied prospectively 30 Graves' hyperthyroid patients, 22 submitted to radioiodine (131I) treatment (group A) and 8 treated with antithyroid drugs (group B). All patients were evaluated by clinical ophthalmologic examination, and ocular proptosis (OP) was measured with both a Hertel exophthalmometer (HE) and computed tomography (CT) before and 4 to 7 months after therapy. No statistical difference was obtained between pre- and post-treatment OP measurements in each eye in either group, and we did not observe worsening in the ophthalmopathy of patients treated with drugs or radioiodine. After therapy, there was an improvement in the clinical signs of ophthalmopathy in 59% of group A and in 37.5% of group B patients. We found a significant correlation between OP measured by HE and by CT. CT findings showed an increase in orbital fat and/or muscle thickening in all patients at baseline, proving to be a useful procedure for ophthalmologic diagnosis in doubtful cases. No patient in either group developed hypothyroidism or elevated TSH levels during the study period; this may explain our good results in the evolution of Graves' ophthalmopathy after treatment with 131I and antithyroid drugs. Euthyroidism seems to be an important factor in the outcome of ophthalmopathy after therapy, whatever the mode of treatment chosen to achieve it.

Evidence for the existence of CCK-producing cells in rat pancreatic islets.

BACKGROUND: Although the existence of cholecystokinin-like immunoreactivity (CCK-LI) in rat pancreas had been reported previously, it was never clearly demonstrated whether CCK is produced in rat pancreatic islets. AIMS: The purpose of this study was to elucidate the source of the CCK-LI, the molecular properties of CCK, and the expression of the CCK gene in islet cells. METHODS: Immunohistochemical studies of rat pancreas were carried out with different rabbit antisera against CCK-8 and CCK-related peptide including N-terminal CCK-33 (1-22) and gastrin-17, and colocalization with known islet hormones including insulin, glucagon, somatostatin, and pancreatic polypeptide was investigated. The major molecular form of CCK in the islets was determined by HPLC. RT-PCR and in situ hybridization were performed to demonstrate the presence of the CCK transcript in the pancreas. RESULTS: CCK-LI was found in the center of the islets, colocalized with insulin in B cells. The major molecular form of CCK in the islets was CCK-8. A 350-nucleotide fragment of PCR-amplified CCK cDNA was detected in the islet as well as the duodenum by RT-PCR. In situ hybridization showed that CCK messenger RNA was located in a large portion of the islets, and this was consistent with the immunohistochemical findings. CONCLUSION: CCK messenger RNA and immunoreactivity are expressed in adult rat pancreatic islets, indicating that CCK-producing cells are present in adult rat islets.

14-3-3 proteins interact with the insulin-like growth factor receptor but not the insulin receptor.

We have used a yeast two-hybrid system to identify proteins which bind to the cytosolic portion of the type 1 insulin-like growth factor (IGF) receptor (IGFIR) but not the insulin receptor (IR). This analysis identified 14-3-3beta and zeta proteins. 14-3-3beta also binds to the IGFIR but not the IR in vitro and 14-3-3-IGFIR complexes are present in insect cells overexpressing the IGFIR cytoplasmic domain. 14-3-3 proteins are substrates of the IGFIR in the yeast system and in vitro. The interaction of 14-3-3 with the IGFIR requires receptor-kinase activity and maps to the C-terminus of the receptor, but does not depend on tyrosine residues in this or the juxtamembrane regions. Instead, the binding maps to serine residue 1283 and requires phosphorylation of this residue. 14-3-3 proteins are phosphoserine-binding proteins which have been shown to interact directly with components of the mitogenic and apoptotic signalling pathways, suggesting that they participate in growth regulation. Our findings suggest that 14-3-3 proteins may play a role in IGFIR signal transduction and may contribute to the differences in IGF and IR signalling capabilities.

The mammalian cell cycle in normal and abnormal growth.

Cell division, a complex array of intracellular events, occurs in a highly ordered and carefully coordinated manner. This regulation is achieved by the sequential activation and deactivation of the members of a family of serine-threonine-specific protein kinases that consist of regulatory and enzymatic subunits, the cyclins and cyclin-dependent kinases. These enzymes, in turn, regulate the activity of other proteins involved in the mitogenic pathway. Mutations in the components of the regulatory pathways can lead to aberrant growth, including malignancies.

Homozygous nonsense mutation in the insulin receptor gene of a patient with severe congenital insulin resistance: leprechaunism and the role of the insulin-like growth factor receptor.

Severe congenital insulin resistance in the syndrome of leprechaunism is caused by mutations in the insulin receptor gene. We report a patient with leprechaunism who was homozygous for a mutation resulting in the absence of cell surface insulin receptors. To determine whether the receptor for Insulin-like growth factor-I (IGF-I) is involved in the phenotype of leprechaunism, we studied the effect of insulin and of IGF-I on cells from this patient. The patient had a homozygous C-->T substitution at base pair 8212 in exon 12 of the insulin receptor gene, creating a premature stop codon. This nonsense mutation is in the extracellular portion of the receptor and truncates the insulin receptor proximal to its transmembrane anchor, resulting in the absence of cell surface insulin receptors. This finding indicates that complete absence of the insulin receptor is compatible with life. Secondly, DNA synthesis was studied in skin derived fibroblasts in response to increasing concentrations of either insulin or Insulin-like growth factor-I (IGF-I), and was assessed by 3H-thymidine incorporation. In this patient's cells, both of these hormones increased 3H-thymidine incorporation, and the effect was blocked by alpha-IR3, a monoclonal antibody that blocks activation of the IGF-I receptor. These findings confirmed the absence of the insulin receptor and indicated that insulin acts here through activation of the IGF-I receptor. These data support the contention that the phenotypic and metabolic abnormalities of leprechaunism result from the combination of lack of insulin receptor action and over-activation by insulin of the type 1 IGF receptor.

Evidence for the direct interaction of the insulin-like growth factor I receptor with IRS-1, Shc, and Grb10.

We have used the yeast two-hybrid system to study the interaction between the IGF-I receptor and two putative substrates, IRS-1 and Shc. In addition, we have identified Grb10 as a protein that binds to the insulin-like growth factor I (IGF-I) receptor. This two-hybrid system (the interaction trap) utilizes a hybrid protein containing the LexA DNA-binding domain fused to the intracellular portion of the IGF-I receptor (LexA-IGFIR beta) and hybrids containing an activation domain fused to either IRS-1 (Ad-IRS-1), Shc (Ad-Shc), or a cDNA library. A positive interaction of LexA-IGFIR beta with the activation domain hybrid results in activation of reporter genes, LacZ and LEU2, in the yeast. Western blotting of extracts from transformed yeast demonstrated that the LexA-IGFIR beta fusion protein was expressed and phosphorylated on tyrosine residues. Coexpression of LexA-IGFIR beta with Ad-IRS-1 resulted in strong activation of both reporter genes; activation did not occur with a kinase-negative receptor mutant. IRS-1 residues 160-516 were sufficient for this strong interaction. Coexpression of LexA-IGFIR beta with Ad-Shc also resulted in strong activation of both LacZ and LEU2 reporter genes. This interaction was also dependent upon a tyrosine kinase-active receptor and required tyrosine 950 in the juxtamembrane region of the receptor. An N-terminal fragment of Shc (amino acids 1-232) interacted almost as strongly as full-length Shc whereas the Shc SH2 domain only activated the more sensitive LEU2 reporter. Full-length Shc was phosphorylated on tyrosine when coexpressed with IGFIR beta but not when coexpressed with the kinase-negative receptor mutant. To identify additional proteins that interact with the IGFIRs, a human fetal brain cDNA library was screened using the interaction trap system. This analysis identified partial cDNAs for Grb10. Coexpression of LexA-IGFIR beta with Ad-Grb10 resulted in strong activation of both LacZ and LEU2 reporter genes; this interaction was dependent upon a tyrosine kinase-active receptor but did not require tyrosine 950.

Comparison of transdermal and oral estrogen therapy in girls with Turner's syndrome.

Eight girls with Turner's syndrome were given low dose oral ethinyl estradiol or transdermal 17 beta-estradiol in order to compare the effect of the route of administration on selected markers of hepatic metabolism, and various hormonal concentrations. Oral estrogen was given at a dose of 100 ng/kg/day and transdermal estrogen via adhesive skin patch at 0.0125 mg/kg/day. The subjects received one form of estradiol for one month, and after a one month washout period, received the other form. Both oral and transdermal estradiol caused a significant decrease in FSH while only transdermal resulted in a significant decrease in LH. Oral estradiol, though not transdermal estradiol, increased serum high density lipoprotein, thyroxine binding protein and growth hormone binding protein. Urinary growth hormone excretion increased after both forms of therapy, while insulin-like growth factor-I and insulin-like growth factor binding protein-3 remained unchanged. Thus, in girls with Turner's syndrome, estrogen replacement by the transdermal route may have less deleterious effect on hepatic metabolism than oral estrogen.

Insulin receptor signaling is augmented by antisense inhibition of the protein tyrosine phosphatase LAR.

Considerable evidence has shown that most physiologic responses to insulin require activation of the intrinsic tyrosine kinase of the insulin receptor. Biochemical studies have also supported the hypothesis that receptor kinase activity can be modulated by cellular protein tyrosine phosphatases (PTPases), which have not yet been identified. To test the hypothesis that the transmembrane PTPase LAR can modulate insulin receptor signaling in vivo, antisense RNA expression was used to specifically suppress LAR protein levels by 63% in the rat hepatoma cell line, McA-RH7777. Hormone-dependent autophosphorylation of the insulin receptor was increased by approximately 150% in the antisense-expressing cells at all insulin concentrations tested. This increase in autophosphorylation was paralleled by a 35% increase in insulin receptor tyrosine kinase activity. Reduced LAR levels did not alter non-hormone-dependent tyrosine phosphorylation nor basal insulin receptor tyrosine phosphorylation and kinase activity. Most significantly, reduced LAR levels resulted in a 350% increase in insulin-dependent phosphatidylinositol 3-kinase activity. These studies provide unique in vivo evidence that LAR is involved in the modulation of insulin receptor signaling in intact cells.