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Biomedical subjects

J M Holly

Publications and source records attributed to J M Holly.

At least 19 recordsLinked to original sources
PubMed2001

Alterations in the insulin-like growth factor system during treatment with diethylstilboestrol in patients with metastatic breast cancer.

Alterations in the insulin-like growth factor (IGF)-system were evaluated in 16 patients treated with diethylstilboestrol 5 mg 3 times daily. Fasting blood samples were obtained before treatment and after 2 weeks, 1 month and/or 2-3 months on therapy. Insulin-like growth factor (IGF)-I, IGF-II, free IGF-I, IGF-binding protein (IGFBP)-1, IGFBP-2 and IGFBP-3 were measured by radioimmuno-/immunoradiometric-assays. All samples were subjected to Western ligand blotting as well as immunoblotting for IGFBP-3. We observed a significant decrease (percentage of pretreatment levels with 95 confidence intervals of the mean) in IGF-I [2 weeks 63% (49-79); 1 month 56% (44-73); 2-3 months 66% (53-82)], IGF-II [2 weeks 67% (56-80); 1 month 60% (52-68); 2-3 months 64% (55-75)], free IGF-I [2 weeks 29% (19-42); 1 month 25% (18-36); 2-3 months 31% (21-46)], IGFBP-2 [2 weeks 53% (18-156); 1 month 69% (61-78); 2-3 months 66% (57-78)], IGFBP-3 [2 weeks 74% (63-85); 1 month 69% (62-76); 2-3 months 71% (63-80)], as well as IGFBP-3 protease activity [2 weeks 71% (54-95); 1 month 78% (64-94); 2-3 months 71% (54-93)]. Contrary, the plasma levels (percentage of pretreatment levels with 95 confidence intervals of the mean) of IGFBP-1 [2 weeks 250% (127-495); 1 month 173% (138-542); 2-3 months 273% (146-510)] and IGFBP-4 [2 weeks 146% (112-192); 1 month 140% (116-169); 2-3 months 150% (114-198)] increased significantly. While this study confirms previous observations during treatment with oral oestrogens in substitution doses, the reduction in plasma IGF-II, free IGF-I, IGFBP-2 and -3 are all novel findings. A profound decrease in free IGF-I suggests a reduced bioavailability of IGFs from plasma to the tissues. These observations may be of significance to understand the mechanisms of the antitumour effect of diethylstilboestrol in pharmacological doses.

Aged↗
PubMed2001

Plasma insulin-like growth factor binding protein-3 proteolysis is increased in primary breast cancer.

Fasting blood samples were obtained before definitive surgery or biopsy in 128 patients referred to the department of surgery with suspected or manifest breast cancer. Insulin-like growth factor (IGF)-I, IGF-II and free IGF-I were measured by radioimmunoassay/immunoradiometric assay, while IGFBP-3 proteolysis was evaluated by Western immunoblot. 12 patients had ductal carcinoma in situ benign conditions, while staging revealed metastatic disease in 15 of 16 patients with invasive cancers. IGFBP-3 proteolysis above the normal range was recorded in 19 patients with invasive cancers, but in none of the patients suffering from DCIS/benign conditions. Increased IGFBP-3 proteolysis was most frequently recorded in patients harbouring large tumours and metastatic disease (Stage I: 0/19, 0%; Stage II: 3/45, 7%, Stage III: 9/37, 24%, and Stage IV: 7/15, 47%). IGFBP-3 proteolysis was significantly higher in Stage III (P =0.01) and IV (P< 0.001) patients compared to the other stage groups (P = 0.001). IGF-I and IGF-II correlated negatively to IGFBP-3 proteolysis and age. Plasma levels of IGF-I and -II were significantly lower in patients with elevated IGFBP-3 proteolysis compared to those within the normal range. Our findings reveal alterations in the IGF-system among a substantial number of patients with large primary breast cancers.

Adult↗
PubMed2001

Insulin-like growth factors (IGF-I and IGF-II) inhibit C2 skeletal myoblast differentiation and enhance TNF alpha-induced apoptosis.

IGF-I and IGF-II are thought to be unique in their ability to promote muscle cell differentiation. Murine C2 myoblasts differentiate when placed into low serum media (LSM), accompanied by increased IGF-II and IGF binding protein-5 (IGFBP-5) production. Addition of 20 ng/ml TNF alpha on transfer into LSM blocked differentiation, IGF-II and IGFBP-5 secretion and induced apoptosis. We, therefore, wished to assess whether IGFs could protect against the effects of TNF alpha. Neither inhibition of differentiation or induction of apoptosis was rescued by co-incubation with IGF-I or IGF-II. A lower dose of TNF alpha (1 ng/ml) while not inducing apoptosis still inhibited myoblast differentiation by 56% +/- 12, (P < 0.001), indicating that induction of apoptosis is not the sole mechanism by which TNF alpha inhibits myoblast differentiation. Addition of IGF-I or IGF-II alone reduced differentiation by 49% +/- 15 and 33% +/- 20, respectively, (P < 0.001), although neither induced apoptosis. For muscle cells to differentiate, they must arrest in G0. We established that addition of IGF-I, IGF-II or TNF alpha to the myoblasts promoted proliferation. The myoblasts could not exit the cell cycle as efficiently as controls and differentiation was thus reduced. Unexpectedly, co-incubation of IGF-I or IGF-II with 1 ng/ml TNF alpha enhanced the inhibition of differentiation and induced apoptosis. In the absence of apoptosis we show an association between IGF-induced inhibition of differentiation and increased IGFBP-5 secretion. These results indicate that the effects of the IGFs on muscle may depend on the cytokine environment. In the absence of TNF alpha, the IGFs delay differentiation and promote myoblast proliferation whereas in the presence of TNF alpha the IGFs induce apoptosis.

Animals↗
PubMed2001

The changes of IGF binding proteins after rhGH administration to patients totally dependent on parenteral nutrition.

The objective was to study the effect of recombinant human growth hormone (rhGH) administration to patients with chronic malnutrition maintained on total parenteral nutrition (TPN) on the levels of insulin-like growth factor-I (IGF-I) and IGF binding proteins (IGFBPs) during a double-blind trial. After 1 week of TPN the patients were randomized into group I (placebo) or group II (rhGH). Samples were collected on the first day (start of the TPN) to measure basal values, the seventh day to study the effect of TPN and the 10th, 14th and 21st days to evaluate the rhGH effect. Basal laboratory evaluation, nutritional status and glucose tolerance were assessed using standard laboratory techniques. Radioimmunoassays were used to analyse IGF-I, free IGF-I (fIGF-I) and IGFBP1-3. Electrophoresis with Western ligand blotting and Western immunoblotting was applied to find the pattern of IGFBPs. TPN had no effect on the circulating IGF-I concentration and the pattern of IGFBPs present in the studied groups of patients. The rhGH administration led to significant increases of IGF-I, total IGFBP-3, glycosylated IGFBP-3 (39, 42 kDa) and the 29 kDa fragment of IGFBP-3 and the decrease of IGFBP-2 during the trial (P<0.05). The mean levels of IGFBP-1, fIGF-I and the parameters of nutritional status in group II during the trial were not significantly influenced by rhGH. However, it has been found that IGFBP-1 and fIGF-I levels were correlated with the levels of the weekly nitrogen balance of each patient in group II at the end of the trial. In spite of the significant changes of IGF-I, IGFBP-2, total IGFBP-3 and IGFBP-3 (29 kDa proteolytic fragment) after rhGH administration to patients with malnutrition, maintained on parenteral nutrition, the increase of nitrogen balance was seen only in patients who decreased their IGFBP-1 and increased bioavailable IGF-I as reflected by measurement of fIGF-I. The levels of IGFBP-1 may provide a useful marker of IGF-I bioavailability in monitoring the efficiency of the rhGH therapy in malnourished patients.

Adult↗
PubMed2001

The insulin-like growth factor system in human immunodeficiency virus infection: relations to immunological parameters, disease progression, and antiretroviral therapy.

Endocrine dysfunctions have previously been reported in human immunodeficiency virus (HIV) infection. In this study we evaluated the relation of immunological parameters, virus load, clinical stage, and wasting to several parameters of the insulin-like growth factor (IGF) system in 76 patients with HIV infection, of whom 37 had developed acquired immune deficiency syndrome (AIDS). A subgroup of 26 untreated patients was followed during longitudinal testing, while the effects of antiretroviral therapy were evaluated in 34 patients (nucleoside analogs in 9, nucleoside analogs in combination with protease inhibitors in 25). Twenty healthy sex- and age-matched controls were analyzed for comparison. IGF-II was decreased (P = 0.03) and IGF-binding protein-2 (IGFBP-2) and IGFBP-3 protease activity were increased (P < 0.001) in AIDS patients compared with other HIV-infected individuals and controls. Plasma levels of IGFBP-2 and IGFBP-3 protease activity correlated positively to virus load (P < 0.001) and tumor necrosis factor-alpha (P < 0.025) and negatively to CD4(+) and CD8(+) cell counts (P < 0.001). AIDS patients with wasting (n = 13) had lower IGF-II levels (P = 0.001) and higher IGFBP-2 levels (P = 0.001) than other AIDS patients. Although no significant change in any of the IGF-parameters was observed in patients during antiretroviral therapy, patients with elevated IGFBP-3 protease activity before therapy (5 of 34) all had a decrease during treatment. During longitudinal testing in patients followed without antiretroviral therapy, disease progression was associated with increases in IGFBP-3 protease activity and IGFBP-2 levels. Our results reveal several alterations in the IGF system during HIV infection with decreased IGF-II levels, increased concentration of IGFBP-2, and an increased IGFBP-3 protease activity in advanced disease.

Adult↗
PubMed2001

The role of IGF-binding proteins in mediating the effects of recombinant human IGF-I on insulin requirements in type 1 diabetes mellitus.

To determine the role of IGF-binding proteins in mediating the direct effects of recombinant human IGF-I on insulin requirements in type 1(insulin-dependent) diabetes mellitus, overnight changes in IGF-I, IGF-II, and IGF-binding protein-1, -2, and -3, collected under euglycemic conditions, were compared in nine subjects after double blind, randomized, sc administration of recombinant human IGF-I (40 microg/kg) or placebo at 1800 h. On both nights a somatostatin analog infusion (300 ng/kg x h) suppressed endogenous GH production, and three timed discrete GH pulses (total, 0.029 IU/kg x night) ensured identical GH levels. After recombinant human IGF-I administration, IGF-I levels and the IGF-I/IGF-binding protein-3 ratio increased [mean +/- SEM:IGF-I, 401 +/- 22 ng/ml; placebo, 256 +/- 20 ng/ml (P = 0.0002); IGF-I, 0.108 +/- 0.006; placebo, 0.074 +/- 0.004 (P = 0.0003), respectively], and insulin requirements decreased (IGF-I, 0.12 +/- 0.03; placebo, 0.23 +/- 0.03 U/kg x min; P = 0.008). The normal within-individual inverse relationships between insulin and IGF-binding protein-1 levels were observed (lag time 2 h: r = -0.34; P < 0.01). Yet despite reduced free insulin levels (8.5 +/- 1.5; placebo, 12.2 +/- 1.2 mU/liter; P = 0.03), IGF-binding protein-1 levels were reduced after recombinant human IGF-I administration (53.7 +/- 6.8; placebo, 82.2 +/- 11.8 ng/ml; P = 0.008). The largest reductions in free insulin levels after recombinant human IGF-I and thus putative improvement in insulin sensitivity occurred in subjects with the smallest increase in the plasma IGF-I/IGF-binding protein-3 ratio (r = 0.7; P = 0.03). Taken together, these data are consistent with the hypothesis that transcapillary movement of IGF-I (perhaps mediated by IGF-binding protein-1), out of the circulation facilitates altered insulin sensitivity. These data have important implications for risk-benefit assessment of recombinant human IGF-I therapy in type 1 diabetes mellitus.

Adolescent↗
PubMed2000

IGFBP-3 prolongs the p53 response and enhances apoptosis following UV irradiation.

Neoplastic transformation is characterised by an imbalance in favour of cell growth over programmed cell death (apoptosis). The tumour-suppressor gene p53, responsible for maintaining cell-cycle control, is mutated in the majority of human cancers. Loss of function of the target genes of p53 are therefore important in tumourigenesis. One such target gene is the insulin-like growth factor binding protein-3 (IGFBP-3), an extracellular protein responsible for the carriage of IGF-I but which can act independently of IGF-I, inhibiting cell growth and enhancing apoptosis. Using the KYSE 190 oesophageal carcinoma cell line, we have demonstrated that IGFBP-3 alone has no effect on cell growth or cell survival. However, it significantly enhanced apoptosis, with a 67% increase in the pre-G1 peak on flow cytometry following UV irradiation. The increase in p53 was enhanced and prolonged when cells are stressed in the presence of IGFBP-3. These data suggest an autocrine/paracrine feedback loop exists between IGFBP-3 and p53, which may provide the social control necessary to maintain normal tissue homeostasis.

Apoptosis↗
PubMed2000

Insulin-like growth factor binding protein-3 (IGFBP-3) potentiates paclitaxel-induced apoptosis in human breast cancer cells.

Variability in response to chemotherapy is poorly understood. Paclitaxel-induced apoptosis was assessed in human Hs578T breast cancer cells, using the MTT assay, cell counting, morphological features and flow cytometry. Pre-dosing cells with non-glycosylated insulin-like growth factor binding protein-3 (ngIGFBP-3) had no effect on the cells per se but accentuated paclitaxel-induced apoptosis. The apoptotic pathway was further examined by measuring caspase-3 activity in cell lysates at time points over 48 hr after dosing with paclitaxel. Activity increased significantly, and Western immunoblots for caspase-3 in conditioned media showed that the inactive precursor decreased after incubation with paclitaxel. Endogenous production of IGFBP-3 by the cells after incubation with paclitaxel was evaluated using Western ligand blotting, specific IGFBP-3 immunoblotting and radioimmunoassay. Paclitaxel increased endogenous IGFBP-3, which was further increased if the cells had been pre-dosed with ngIGFBP-3. These findings suggest that IGFBP-3 may be an important modulator of paclitaxel-induced apoptosis.

Antineoplastic Agents, Phytogenic↗
PubMed2000

Differential insulin-like growth factor (IGF)-independent interactions of IGF binding protein-3 and IGF binding protein-5 on apoptosis in human breast cancer cells. Involvement of the mitochondria.

We have demonstrated previously in Hs578T cells that insulin-like growth factor binding protein (IGFBP)-3 can significantly accentuate ceramide (C2)-induced apoptosis, but has no effect on cell death induced by integrin detachment [using an arginine-glycine-aspartic acid (RGD)-containing peptide]. In contrast we found that IGFBP-5 could inhibit apoptosis induced by either C2 or integrin detachment. It is now clear that the mitochondria not only provide the energy required for cell viability, but can also play an important role during the commitment phase to apoptosis. We used a mitochondrial respiratory chain inhibitor, antimycin A, at both apoptotic and nonapoptotic doses to further investigate the IGF-independent actions of IGFBP-3 and IGFBP-5 on C2 and RGD-induced apoptosis in the Hs578T cells. Hs578T cells had one of three treatments. 1: They were incubated with increasing doses of antimycin A for 24 h. 2: They were coincubated with an apoptotic dose of either C2 or RGD together with a nonapoptotic dose of antimycin A for 24 h. 3: They were incubated with a binding protein (100 ng/ml) for 24 h followed by coincubation of the binding protein with an apoptotic dose of antimycin A for a further 24 h. Cell viability was assessed by trypan blue dye exclusion and MTT assay, and apoptosis was confirmed and measured by morphologic assessment and flow cytometry. We found that antimycin A initiated apoptosis at 10 micromol/L and above. We also demonstrated that a nonapoptotic dose of antimycin A (0.1 micromol/L) significantly inhibited C2-induced apoptosis, whereas it significantly accentuated RGD-induced cell death. In addition, we found that cell death induced by antimycin A can be accentuated by IGFBP-3 but is not affected by IGFBP-5. These data indicate that IGFBP-3 can directly enhance apoptosis triggered via the mitochondria; either directly by a mitochondrial inhibitor or by C2 (which we demonstrate to act via effects on the mitochondria in this model). IGFBP-5, however, appears to confer survival effects via a distinct pathway not involving the mitochondria.

Antimycin A↗
PubMed2000

Increased p53-dependent apoptosis by the insulin-like growth factor binding protein IGFBP-3 in human colonic adenoma-derived cells.

We investigated the expression of insulin-like growth factor binding protein 3 (IGFBP-3) in normal human colonic epithelium and whether IGFBP-3 is involved in the induction of apoptosis in colonic epithelial cells. A gradient of IGFBP-3 protein expression was observed within the normal colonic crypt, and increased IGFBP-3 expression was coincident with the region of increased differentiation and apoptosis. Treatment of human colonic tumor cell lines with IGFBP-3 alone was shown to have no effect on growth. However, an increase in p53-dependent apoptosis was observed in the presence of 100 ng/ml IGFBP-3 24 h after the induction of DNA damage by gamma-irradiation. These results suggest that IGFBP-3 enhances the p53-dependent apoptotic response of colorectal cells to DNA damage.

Adenocarcinoma↗
PubMed2000

Tumor necrosis factor-alpha-induced apoptosis is associated with suppression of insulin-like growth factor binding protein-5 secretion in differentiating murine skeletal myoblasts.

Wasting of muscle and fat during cachexia exceeds that explained by reduced food intake alone. This wasting may result from an imbalanced cytokine environment, which could lead to increased protein catabolism. Supporting this, tumor necrosis factor-alpha (TNF-alpha) is raised in several animal models of cachectic muscle wasting. Therefore, we assessed the effects of TNF-alpha and its second messenger, ceramide, on the proliferation, differentiation, and survival of murine C2 skeletal myoblasts. Because insulin-like growth factor binding protein-5 (IGFBP-5) and insulin-like growth factor-II (IGF-II) are potent regulators of myoblast proliferation and differentiation, we monitored the ability of exogenous TNF-alpha to manipulate this system. Fibroblast growth factor (FGF) ceramide, or TNF-alpha suppressed differentiation of C2 cells compared with controls. All treatments suppressed IGF-II production but only TNF-alpha blocked IGFBP-5 secretion. TNF-alpha increased apoptotic cell death, which otherwise remained basal (low serum differentiation medium (LSM), FGF) or low (ceramide). Suppression of both IGFBP-5 and IGF-II secretion may explain why of all triggers tested, only TNF-alpha not only blocked differentiation, but also promoted cell death. This suggests a fundamental role of IGFBP-5 for maintaining muscle survival. Supporting this hypothesis, no increase in apoptosis was seen in IGFBP-5 cDNA tranfected C2 cells after TNF-alpha treatment. In summary, the IGF system is essential for maintaining skeletal muscle cell survival and differentiation, and its suppression by TNF-alpha is fundamental regarding muscle wasting, and may be associated in vivo with cancer cachexia.

Animals↗
PubMed2000

Insulin-like growth factor binding proteins (IGFBPs) in breast cancer.

The IGFBP family comprises six proteins with high affinity for the IGFs and several lower affinity IGFBP-related proteins. Their production in the breast is controlled by hormones, other local regulators and in tumors relates to the estrogen receptor status. Their functional activity can also be affected by various post-translational modifications. The IGFs are generally present at levels far in excess of that required for maximal receptor stimulation, and the IGFBPs are critical regulators of cellular action. IGFBPs can affect cell function in an IGF-dependent or independent manner. IGF bioactivity locally in the breast is influenced not only by local tissue expression and regulation of IGFs, IGFBPs, and IGFBP proteases, but also by these factors delivered from the circulation. Changes in the balance of the components of the IGF system may lead to a disruption of tissue homeostasis.

Breast Neoplasms↗
PubMed2000

Endogenous IGFBP-3 is protected from inducible IGFBP-3 protease activity in normal adult serum.

We have recently demonstrated that the presence or absence of IGFBP-3 protease activity in physiological fluids may not be determined simply by the presence or absence of specific enzymes but rather the presence of inhibitors. In addition, it appears that these inhibitors may not only be associated with the protease(s) but with the IGFBP-3 itself, protecting it from proteolytic cleavage. To provide further evidence for this mechanism of regulation we investigated whether IGFBP-3 protease activity could be unveiled in normal adult serum (NS) and whether the endogenous IGFBP-3 was protected from this activity. The addition of a range of concentrations of heparin, induced IGFBP-3 protease activity in NS. This was comparable to that seen in pregnancy serum (PS) by virtue of the fragmentation pattern and inhibitor profile. While the addition of zinc also revealed IGFBP-3 protease activity in NS the pattern of fragments differed to that seen in PS. Under both conditions, however, the endogenous IGFBP-3 was not proteolytically modified. These results demonstrate that IGFBP-3 protease activity is present in NS and can be activated, although the endogenous IGFBP-3 is relatively protected from such activity.

Adult↗
PubMed2000

Maternal insulin-like growth factor binding protein-1, body mass index, and fetal growth.

AIM: To examine the hypothesis that the maternal insulin-like growth factor system may constrain fetal growth. METHODS: A prospective observational study of maternal serum insulin-like growth factor binding protein-1 (IGFBP-1) and fetal growth was undertaken in neonates with birthweights below the 5th centile. They had been classified either as having fetal growth restriction (FGR) due to placental dysfunction (increased umbilical artery Doppler pulsatility index (PI); n = 25) or as being small for gestational age (SGA; normal umbilical artery PI, growth velocity and amniotic fluid; n = 27). Eighty nine controls had normal birthweights (5th-95th centile), umbilical artery PI, growth velocity, and amniotic fluid. IGFBP-1 was measured by radioimmunoassay. RESULTS: Among the controls, there was no significant correlation between IGFBP-1 and birthweight after allowing for body mass index (BMI). Maternal BMI was high in FGR and after adjusting for this, IGFBP-1 was increased (109 ng/ml) compared with SGA babies (69 ng/ml) and controls (57 ng/ml) and correlated with the umbilical artery PI. CONCLUSIONS: Maternal IGFBP-1 is probably not part of normal placental function. Its increase in FGR could be the cause or consequence of impaired placental perfusion, but high IGFBP-1 concentrations might further reduce the availability of maternal IGF-I to the placenta. This could worsen placental function and so adversely affect fetal growth.

Adult↗
PubMed2000

Alterations in insulin-like growth factor binding protein-3 proteolysis and complex formation in the arthritic joint.

Increased concentrations of insulin-like growth factor (IGF) system components have previously been observed in rheumatoid arthritis (RA) and osteoarthritis (OA); however, disruption of the IGF axis and the implications for the disease process remain largely unaddressed. This study was undertaken to characterise the IGF binding protein (IGFBP)-3 proteolysis and complex formation systems in synovial fluid and to investigate changes in these systems in arthritic disease, and their impact on the availability of IGF. Western blotting or autoradiography of SDS gels was used to visualise IGFBP-3 or its proteolysis. IGF-I and IGFBP-3 concentrations were determined by radioimmunoassays and acid-labile subunit (ALS) was measured by ELISA. A shift in distribution of IGFBP-3 and IGF-I in RA and OA synovial fluids (RASynF, OASynF) and an associated increase in ALS suggested the presence of 150 kDa ternary complexes. IGFBP-3 proteolysis was decreased in RASynF and OASynF, but was apparent in size-fractionated fluid and resembled serum activity. The presence of serum-like inhibitors of IGFBP-3 proteolysis in RASynF was also demonstrated by the ability of this fluid, and 150 kDa fractions from its size fractionation, to inhibit IGFBP-3 proteolysis in the other synovial fluid. A marked disruption in the IGF system was observed, as considerably more IGF-I was retained in ternary complexes. We also classified the IGFBP-3 proteolysis system in synovial fluid and found it to be disturbed in RASynF and OASynF. These changes may be caused by an increased flux of circulatory proteins into synovial fluid, resulting from an inflammation-induced increase in vascular permeability. The net result in RA and OA would be a decrease in IGF availability in arthritic joints, and therefore loss of a potential anabolic stimulus. This disruption to the IGF axis would influence disease progression in RA and OA.

Adult↗