Unbound Growth Research Articles

Using UV-Responsive Nanoparticles to Provide In Situ Control of Growth Factor Delivery and a More Constant Release Profile from a Hydrogel Environment.

Nanoparticles are popular delivery vehicles, but their diffusional release results in inconstant drug delivery. Here, we flatten the delivery profile into a more constant, zero-order profile. Brain-derived neurotrophic factor (BDNF) is attached to photoactive titanium dioxide nanoparticles and loaded into a nanofibrous self-assembling peptide (SAP) hydrogel. Different UV exposure conditions show three distinct profiles, including a counterintuitive decrease in release after UV exposure. We propose that the adsorption of the freed growth factor onto the hydrogel nanofibers affects release. Nanoparticles diffuse from the hydrogel readily, carrying the bound growth factor, but the freed growth factor (released from the nanoparticles by UV) instead interacts with─and is released less readily from─the hydrogel. UV shifts growth factor from nanoparticles to the hydrogel, therefore changing the diffusional release. Through midpoint UV exposure, we achieve a flattened delivery profile─unusual for diffusion─by changing in situ the amount of growth factor bound to the diffusing nanoparticles. With nanoparticle diffusion alone, we observed an increasing release profile with 36% of release in the first 6 h and 64% in the second 6 h. With midway UV exposure, this was controlled to 49 and 51%, respectively. The release of an unbound (soluble) control growth factor, glial cell-line derived neurotrophic factor (GDNF), was not affected by UV treatment, demonstrating the potential for independent control of temporal delivery profiles in a multiagent material.

A Novel Fluorescence-Labeled Curcumin Conjugate: Synthesis, Evaluation and Imaging on Human Cell Lines

Curcumin, as the main ingredient of the curcuma spice, has increasingly become the target of scientific research. The turmeric root where the spice is obtained from has been widely used in the traditional medicine. Moreover, scientific studies have found that curcumin has anti-inflammatory, anti-cancer, anti-angiogenic effects as well as antibacterial properties. Recently, curcumin has gathered interest as a potential therapeutic agent in the research on Alzheimer's disease. A consistent problem in the investigative and therapeutic applications of curcumin is its poor solubility in aqueous solutions. In the present study, we synthesized a conjugate of curcumin, the amino acid lysine and the fluorescent dye fluorescein. This conjugate was soluble in cell culture medium and facilitated the examination of curcumin with fluorescence imaging methods. We studied the cell growth impact of unmodified curcumin on seven different human cell lines and then analyzed the uptake and cellular localization of our curcumin conjugate with confocal laser scanning imaging and flow cytometry on the seven cell lines. We found that unbound curcumin inhibited cell growth in vitro and was not taken up into the cells. The curcumin conjugate was internalized into the cell cytoplasm in a dot-like pattern and cellular uptake correlated with the cell membrane damage which was measured using propidium iodide. The CAL-72 osteosarcoma cell exhibited 3-4fold increased conjugate uptake and a strong uniform fluorescein staining in addition to the dot-like pattern observed in all cell lines. In conclusion, we successfully synthesized a novel water-soluble fluorescent curcumin conjugate which showed a strong preference for CAL-72 osteosarcoma cells in vitro.

Software metric evaluation on cloud based applications

Unbound growth in the cloud computing service models have motivated the companies building traditional software to be migrated into the clouds. During the high demand of the traditional applications, the performance and quality of the software were evaluated by the popular and globally accepted metrics. Nevertheless, after the migration of the same applications into the cloud, the expectation and definition of performance and quality has been changed. The beneficiaries of these applications are setting new milestones for the applications. Hence, the recent demand of the research trend is to build new software metric models to match the trade of between the new expectations from the beneficiaries and the software quality policies for organization or individual or state. Thus this work makes an attempt to understand the traditional software quality metrics and try to justify the applicability of these parameters in the trend of cloud based software applications. This work also proposes a novel metric method for performance evaluation for the migrated applications into the cloud, with the intension of formalizing and standardizing the cloud based metric methods unlike the recent trends.

Measurement of Cetuximab and Panitumumab-Unbound Serum EGFR Extracellular Domain Using an Assay Based on Slow Off-Rate Modified Aptamer (SOMAmer) Reagents

BackgroundResponse to cetuximab (Erbitux®) and panitumumab (Vectibix®) varies among individuals, and even those who show response ultimately gain drug resistance. One possible etiologic factor is differential interaction between the drug and target. We describe the development of an assay based on Slow Off-rate Modified Aptamer (SOMAmer™) reagents that can distinguish drug-bound from unbound epidermal growth factor receptor (EGFR).MethodsThis quantitative assay uses a SOMAmer reagent specific for EGFR extracellular domain (ECD) as a capturing reagent. Captured SOMAmer is quantitated using PCR. Linearity and accuracy (recovery) of the assay were assessed using normal sera and purified EGFR ECD.ResultsThis EGFR ECD assay showed linearity between 2.5 and 600 ng/mL. Average recovery was 101%. The assay detected EGFR but showed little cross-reactivity to other ErbB proteins: 0.4% for ErbB2, 6.9% for ErbB3, and 1.3% for ErbB4. Preincubation of normal serum with either cetuximab or panitumumab resulted in a dose-dependent decrease in EGFR ECD levels measured using the SOMAmer assay; preincubation did not affect measurement with an ELISA.ConclusionsThis SOMAmer-based serum EGFR ECD assay accurately and specifically measures EGFR in serum. Detection of significant amounts of drug-unbound EGFR in patients undergoing cetuximab or panitumumab treatment could be an indicator of poor drug response. Further studies are needed to evaluate the utility of the assay as an indicator of drug efficacy or as a guide to dosing.

Increased plasma levels of soluble vascular endothelial growth factor receptor 1 (sFlt-1) in women by moderate exercise and increased plasma levels of vascular endothelial growth factor in overweight/obese women

The incidence of breast cancer is increasing worldwide, and this seems to be related to an increase in lifestyle risk factors, including physical inactivity and overweight/obesity. We have reported previously that exercise induced a circulating angiostatic phenotype characterized by increased soluble fms-like tyrosine kinase-1 (sFlt-1) and endostatin and decreased unbound vascular endothelial growth factor (VEGF) in men. However, there are no data on women. The present study determines the following: (a) whether moderate exercise increased sFlt-1 and endostatin and decreased unbound VEGF in the circulation of adult female volunteers and (b) whether overweight/obese women have a higher plasma level of unbound VEGF than lean women. A total of 72 African American and White adult women volunteers ranging in age from 18 to 44 years were enrolled in the exercise study. All the participants walked on a treadmill for 30 min at a moderate intensity (55-59% heart rate reserve), and oxygen consumption (VO(2)) was quantified utilizing a metabolic cart. We obtained blood samples before and immediately after exercise from 63 participants. ELISA assays showed that the plasma levels of sFlt-1 were 67.8±3.7 pg/ml immediately after exercise (30 min), significantly higher than the basal levels, 54.5±3.3 pg/ml, before exercise (P<0.01; n=63). There was no significant difference in the % increase in the sFlt-1 levels after exercise between African American and White (P=0.533) women or between lean and overweight/obese women (P=0.892). There was no significant difference in the plasma levels of unbound VEGF (35.28±5.47 vs. 35.23±4.96 pg/ml; P=0.99) or endostatin (111.12±5.48 vs. 115.45±7.15 ng/ml; P=0.63) before and after exercise. The basal plasma levels of unbound VEGF in overweight/obese women were 52.26±9.6 pg/ml, significantly higher than the basal levels of unbound VEGF in lean women, 27.34±4.99 pg/ml (P<0.05). The results support our hypothesis that exercise-induced plasma levels of sFlt-1 could be an important clinical biomarker to explore the mechanisms of exercise training in reducing the progression of breast cancer and that VEGF is an important biomarker in obesity and obesity-related cancer progression.

Acceleration of particles by black holes as a result of deceleration: Ultimate manifestation of kinematic nature of BSW effect

The recently discovered so-called BSW effect consists in the unbound growth of the energy Ec.m. in the center of mass frame of two colliding particles near the black hole horizon. We consider a new type of the corresponding scenario when one of two particles (“critical”) remains at rest near the horizon of the charged near-extremal black hole due to balance between the attractive and repulsion forces. The other one hits it with a speed close to that of light. This scenario shows in a most pronounced way the kinematic nature of the BSW effect. In the extremal limit, one would gain formally infinite Ec.m. but this does not happen since it would have require the critical massive particle to remain at rest on the null horizon surface that is impossible. We also discuss the BSW effect in the metric of the extremal Reissner–Nordström black hole when the critical particle remains at rest near the horizon.

Corals on the slope (Aptian, Maestrat Basin, Spain)

The term “reef” has been frequently misused when applied to fossil coral communities. Our popular but biased view of coral community structure based on the idyllic picture of recent tropical reefs has failed to recognize that, in many fossil examples, alternative states of community structure with no or limited framework may occur. The Aptian colonial scleractinians analysed in the western Maestrat Basin (eastern Spain) constitute an example of non-reef-building coral populations, which thrived in marly slope settings. These corals developed within the photic zone but below the storm wave-base. All colonies are found well-preserved in life position. They are mostly decimetres in size and mainly occur isolated giving rise to a continuous and uniform (dominated by domal and massive forms) unbound growth fabric with a low to medium degree of development (coral skeletal volume = 5–20%). Occasionally, however, colonies growing on top of each other forming small metre-sized bioherms are also present. A total of 21 species were identified. Coral diversity in each sample location varies between three and nine species. These numbers of species are comparable with those exhibited by coeval coral assemblages from other basins of the Tethys, but are comparatively low when compared with diversities exhibited by many Recent and fossil coral communities. The corals studied apparently found optimal ecological conditions for their development on the marly slopes of the western Maestrat Basin. This is primarily expressed in the unusually large dimensions (up to 2.3 m in width) of some of the coral colonies when compared to other Cretaceous occurrences, and in the persistence and resilience of the coral populations. The observed coral genera and species (suborders Archeocaeniina, Faviina, Fungiina and Microsolenina) are very common in the time interval between the Barremian and the Early Albian and most of them have been reported from several other localities in the western and central Tethyan realm. In addition, the coral-bearing levels also contain the poorly known and exotic genera Agrostyliastraea and Procladocora. There are no significant differences at species level or in community structure between the Early and Late Aptian faunas investigated. Therefore, the coral communities as well as the environmental conditions controlling them would have been relatively stable during the time intervals when these corals flourished. An important palaeoecological implication is that comparatively low species diversities and the absence of reef frameworks do not necessarily imply unfavourable environmental conditions for coral growth. Furthermore, this study may serve as an example for the analysis of other level-bottom coral communities displaying a loose growth fabric.

Delivery of VEGF using collagen‐coated polycaprolactone scaffolds stimulates angiogenesis

Establishing sufficient vascularization in scaffold remains a challenge for tissue-engineering. To improve angiogenesis, we incorporated vascular endothelial growth factor (VEGF) in collagen-coating over the porous polycaprolactone (PCL) scaffolds. The release kinetics of loaded VEGF from collagen-coated PCL (col-PCL) scaffolds was same as from scaffolds without the collagen. The bioactivity of VEGF delivered by the col-PCL scaffolds was confirmed by human umbilical vein endothelial cell (HUVEC) proliferation and chorioallantoic membrane (CAM) assay. The col-PCL scaffolds were implanted subcutaneously in mice for 7 and 14 days. At day 7, vascularization within scaffolds loaded with VEGF was superior to that in the scaffolds without VEGF. However, the vessel connectivity to host circulatory system was incomplete and restricted to the scaffold edges. At day 14, blood vessels in scaffolds reached density similar to the subcutaneous tissue and were perfusable throughout the implant thickness. Prewashing the scaffolds with saline to remove the unbound growth factor decreased the initial burst release and sustained the VEGF-mediated angiogenesis in vivo. In conclusion, our study demonstrates that physically adsorbed VEGF stimulated angiogenesis in collagen-coated PCL scaffolds.

Inkjet Printing of Growth Factor Concentration Gradients and Combinatorial Arrays Immobilized on Biologically-Relevant Substrates

Current methods for engineering immobilized, 'solid-phase' growth factor patterns have not addressed the need for presentation of the growth factors in a biologically-relevant context. We developed an inkjet printing methodology for creating solid-phase patterns of unmodified growth factors on native biological material substrates. We demonstrate this approach by printing gradients of fluorescently labeled bone morphogenetic protein-2 (BMP-2) and insulin-like growth factor-II (IGF-II) bio-inks on fibrin-coated surfaces. Concentration gradients were created by overprinting individual substrate locations using a dilute bio-ink to modulate the surface concentration of deposited growth factor. Persistence studies using fluorescently-labeled BMP-2 verified that the gradients retained their shape for up to 7 days. Desorption experiments performed with (125)I-BMP-2 and (125)I-IGF-II were used to quantify the surface concentration of growth factor retained on the substrate for up to 10 days in serum containing media after rinsing of the unbound growth factor. The inkjet method is programmable so the gradient shape can be easily modified as demonstrated by printed linear gradients with varying slopes and exponential gradients. In addition, the versatility of this method enabled combinatorial arrays of multiple growth factors to be created by printing overlapping patterns. The overlapping printing method was used to create a combinatorial square pattern array consisting of various surface concentrations of BMP-2 and fibroblast growth factor-2 (FGF-2). C2C12 myogenic precursor cells were seeded on the arrays and alkaline phosphatase staining was performed to determine the effect of FGF-2 and BMP-2 surface concentration on guiding C2C12 cells towards an osteogenic lineage. These results demonstrate the utility of inkjet printing for creating orthogonal growth factor gradients to investigate how combinations of immobilized growth factors influence cell fate.

Topology and phase transitions II. Theorem on a necessary relation

In this second paper, we prove a necessity theorem about the topological origin of phase transitions. We consider physical systems described by smooth microscopic interaction potentials V N ( q ) , among N degrees of freedom, and the associated family of configuration space submanifolds { M v } v ∈ R , with M v = { q ∈ R N | V N ( q ) ⩽ v } . On the basis of an analytic relationship between a suitably weighed sum of the Morse indexes of the manifolds { M v } v ∈ R and thermodynamic entropy, the theorem states that any possible unbound growth with N of one of the following derivatives of the configurational entropy S ( − ) ( v ) = ( 1 / N ) log ∫ M v d N q , that is of | ∂ k S ( − ) ( v ) / ∂ v k | , for k = 3 , 4 , can be entailed only by the weighed sum of Morse indexes. Since the unbound growth with N of one of these derivatives corresponds to the occurrence of a first- or of a second-order phase transition, and since the variation of the Morse indexes of a manifold is in one-to-one correspondence with a change of its topology, the Main Theorem of the present paper states that a phase transition necessarily stems from a topological transition in configuration space. The proof of the theorem given in the present paper cannot be done without Main Theorem of paper I.

VITREOUS LEVELS OF UNBOUND BEVACIZUMAB AND UNBOUND VASCULAR ENDOTHELIAL GROWTH FACTOR IN TWO PATIENTS

Vitreous levels of unbound bevacizumab (Avastin) and unbound vascular endothelial growth factor (VEGF) were determined in two patients. Patient 1 underwent repair of an 8-day-old rhegmatogenous retinal detachment 4 weeks after a single intravitreal bevacizumab injection, and Patient 2 underwent vitreous biopsy for endophthalmitis 48 hours after a combined bevacizumab and triamcinolone injection. The samples of vitreous fluid were analyzed for unbound bevacizumab and unbound VEGF levels using microsphere immunoassays targeted for bevacizumab and VEGF. In Patient 1, the unbound bevacizumab level was 0.16% of the loading dose (or 500,000 pg/mL) and the unbound VEGF concentration was <41 pg/mL 4 weeks after the bevacizumab injection. In Patient 2, the unbound bevacizumab level was 53% of the loading dose (or 166,000,000 pg/mL) at 48 hours, with an unbound VEGF level of <41 pg/mL. A single dose of intravitreal bevacizumab is likely to provide complete intravitreal VEGF blockade for a minimum of 4 weeks, with an intravitreal bevacizumab half-life of approximately 3 days.

The Mucin Muc4 Potentiates Neuregulin Signaling by Increasing the Cell-surface Populations of ErbB2 and ErbB3

Mucins provide a protective barrier for epithelial surfaces, and their overexpression in tumors has been implicated in malignancy. We have previously demonstrated that Muc4, a transmembrane mucin that promotes tumor growth and metastasis, physically interacts with the ErbB2 receptor tyrosine kinase and augments receptor tyrosine phosphorylation in response to the neuregulin-1beta (NRG1beta) growth factor. In the present study we demonstrate that Muc4 expression in A375 human melanoma cells, as well as MCF7 and T47D human breast cancer cells, enhances NRG1beta signaling through the phosphatidylinositol 3-kinase pathway. In examining the mechanism underlying Muc4-potentiated ErbB2 signaling, we found that Muc4 expression markedly augments NRG1beta binding to A375 cells without altering the total quantity of receptors expressed by the cells. Cell-surface protein biotinylation experiments and immunofluorescence studies suggest that Muc4 induces the relocalization of the ErbB2 and ErbB3 receptors from intracellular compartments to the plasma membrane. Moreover, Muc4 interferes with the accumulation of surface receptors within internal compartments following NRG1beta treatment by suppressing the efficiency of receptor internalization. These observations suggest that transmembrane mucins can modulate receptor tyrosine kinase signaling by influencing receptor localization and trafficking and contribute to our understanding of the mechanisms by which mucins contribute to tumor growth and progression.

Multi-sensor data fusion for land vehicle attitude estimation using a fuzzy expert system

In Inertial Navigation Systems (INS), the attitude estimated from gyro measurements by the Kalman filter is subject to an unbound error growth during the stand-alone mode, especially for land vehicle applications using low-cost sensors. To improve the attitude estimation of a land vehicle, this paper applies a fuzzy expert system to assist in multi-sensor data fusion from MEMS accelerometers, MEMS gyroscopes and a digital compass based on their complementary motion detection characteristics. Field test results have shown that drift-free and smooth attitude estimation can be achieved and will lead to a significant performance improvement for velocity and position estimation.

Hard Disk Drive Capacity at High Magnetic Areal Density

Increases in recording areal density have propelled the disk drive industry in the last dozen years. Several studies and research programs strive to accomplish an areal density of 1 Tb/in/sup 2/. At present, the future of the hard disk drive technology appears predicated on monotonic and unbound areal density growth. The system consequences of using large areal densities are the subject of this paper. It is argued that the limited signal-to-noise ratio (SNR) predicted at 1 Tb/in/sup 2/, as well as the extreme mechanical demands on the system, will cause the user capacity not to grow linearly with areal density, as has been the case in the past. This is due, among other reasons, to the increased servo overhead, the increased error-correction code overhead, and the mechanical requirements that compel the reduction of the size of the mechanism. Without casting in doubt the feasibility of magnetic bit recording at 1 Tb/in/sup 2/, scaling laws nevertheless show that there may be limited rewards for the technical efforts directed toward accomplishing 1 Tb/in/sup 2/. Unless the system SNR can be maintained at about 18 dB, areal density gains will not result in proportional user-capacity gains.

Increases in Free, Unbound Insulin-like Growth Factor I Enhance Insulin Responsiveness in Human Hepatoma G2 Cells in Culture

Insulin-like growth factor-binding protein (IGFBP)-1 binds to insulin-like growth factor (IGF)-I and -II with high affinity and has been shown to modulate IGF-I actions in vivo and in vitro. The synthesis of IGFBP-1 is suppressed by insulin, and administration of IGFBP-1 to rats results in impaired glucose metabolism. A synthetic peptide (bp1-01) has been shown to have a high affinity and specificity for human IGFBP-1 and to inhibit IGF-I binding. The current studies were undertaken to determine if, after incubation of bp1-01 with IGF-I.IGFBP-1 complexes, anabolic and insulin-like effects of IGF-I could be detected in human hepatoma (HepG2) cell cultures and to determine the receptor subtype(s) through which these effects were mediated. Incubation of HepG2 cells with bp1-01 (200 nm) increased IGF-I-stimulated protein synthesis by 44% and glycogen synthesis by 170% compared with stimulation by IGF-I alone. Incubation with bp1-01 also enhanced IGF-I-stimulated tyrosine phosphorylation of the IGF-I/insulin hybrid receptor and insulin receptor substrate 1. Exposure of the cells to bp1-01 alone enhanced glycogen synthesis and phosphorylation of IGF-I/insulin hybrid receptors. This was not a direct effect of bp1-01 because it did not bind to the receptor and did not activate tyrosine kinase activity in the presence of an anti-IGF-I receptor antibody. The addition of bp1-01 (200 nm) plus insulin to HepG2 cell culture medium resulted in increased tyrosine phosphorylation of the hybrid receptor, insulin receptor substrate 1, and the glycogen synthesis response compared with the effects of insulin alone. This enhancement of hybrid receptor phosphorylation and glycogen synthesis by bp1-01 peptide was diminished by preincubation with an inhibitory antibody for the alpha subunit of IGF-I receptor (alphaIR3). bp1-01 stimulated the hybrid receptor phosphorylation response to IGF-I, and this effect was inhibited by prior incubation of the cells with alphaIR3. In conclusion, bp1-01 competes with IGF-I for binding to IGFBP-1, which leads to release of free IGF-I from IGF-I.IGFBP-1 complexes. This released IGF-I stimulates biologic actions that are mediated predominantly through the IGF-I/insulin hybrid receptor.

Unbound vascular endothelial growth factor and its receptors in breast, human milk, and newborn intestine

Human milk, rich in cytokines, may contain the potent permeability- and angiogenesis-promoting agent vascular endothelial growth factor (VEGF). We wanted to study whether free or bound VEGF is present in human milk and whether it and its receptors (VEGFR-1 and -2) are expressed in lactating breast or newborn intestinal tissue. The study had a longitudinal design with collection of human milk from healthy (n = 32) and diabetic (n = 5) women at 2, 7, and 30 d postpartum. Milk was analyzed for VEGF by enzyme-linked immunosorbent assay along with plasma samples collected 2 d postpartum. Immunohistochemistry was used to localize VEGF and its receptors in lactating breast and newborn intestine. Gel filtration with radiolabeled VEGF was performed to study whether human milk contains VEGF binding proteins. Human milk VEGF concentrations in healthy (76 +/- 19 microg/L, x +/- SD) and diabetic (75 +/- 25 microg/L) women did not differ at 2, 7 (23 +/- 7 and 27 +/- 8 microg/L, respectively), or 30 d (14 +/- 5 and 17 +/- 7 microg/L, respectively) postpartum. VEGF was undetectable in all but 3 plasma samples. Human milk was free of VEGF binding proteins. VEGFR-1 and -2 immunoreactivity was seen in the glandular epithelial cells of the newborn intestine and lactating breast, whereas VEGF was present only in breast glandular epithelium. The high concentrations of VEGF in human milk, especially colostrum, are not affected by maternal diabetes and may play a role in newborn nutrition.

The Conformation-dependent Interaction of α2-Macroglobulin with Vascular Endothelial Growth Factor: A NOVEL MECHANISM OF α2-MACROGLOBULIN/GROWTH FACTOR BINDING

The Conformation-dependent Interaction of α2-Macroglobulin with Vascular Endothelial Growth Factor: A NOVEL MECHANISM OF α2-MACROGLOBULIN/GROWTH FACTOR BINDING

Insulin-like growth factors I and II are unable to form and maintain their native disulfides under in vivo redox conditions

Insulin-like growth factor (IGF) I does not quantitatively form its three native disulfide bonds in the presence of 10 mM reduced and 1 mM oxidized glutathione in vitro [Hober, S. et al. (1992) Biochemistry 31, 1749–1756]. In this paper, we show (i) that both IGF-I and IGF-II are unable to form and maintain their native disulfide bonds at redox conditions that are similar to the situation in the secretory vesicles in vivo and (ii) that the presence of protein disulfide isomerase does not overcome this problem. The results indicate that the previously described thermodynamic disulfide exchange folding problem of IGF-I in vitro is also present in vivo. Speculatively, we suggest that the thermodynamic disulfide exchange properties of IGF-I and II are biologically significant for inactivation of the unbound growth factors by disulfide exchange reactions to generate variants destined for rapid clearance.

Serum free insulin-like growth factor I (IGF-I), total IGF-I, and IGF-binding protein-3 concentrations in normal children and children with growth hormone deficiency.

To evaluate the role of serum free or unbound insulin-like growth factor I (IGF-I) on bone growth, we measured serum free IGF-I levels in 354 healthy children and adults (193 males and 161 females, aged 0-40 yr) and in 21 prepubertal GH-deficient (GHD) children (complete GHD, n = 5; partial GHD, n = 16) using a recently developed immunoradiometric assay. We obtained the following results. 1) In the normal children, the serum free IGF-I levels were low in infancy (<1 yr of age; males, 0.71 +/- 0.26 microg/L, mean +/- SD; females, 1.05 +/- 0.49 microg/L), increased during puberty (males, 5.84 +/- 2.18 microg/L; females, 5.80 +/- 1.49 microg/L), and declined thereafter. 2) Free IGF-I in the serum occupied about 0.95-2.02% of the total IGF-I values, with the highest ratio occurring in infancy (males, 1.77 +/- 0.60%; females, 2.02 +/- 0.87%). 3) The SD scores of serum free IGF-I in the 21 GHD children ranged from -3.30 to 0.30, and the 5 complete GHD children had free IGF-I values more than -2 SD below those of age-matched normal subjects. 4) There was a significant correlation between the SD scores of free IGF-I and those of total IGF-I (r = 0.715; P < 0.0005) in the GHD children. 5) In the 16 partial GHD children receiving GH treatment, the serum free IGF-I levels were elevated to 209% of pretreatment levels after 1 month of GH treatment and remained high during GH therapy. The GH-induced increase in the serum free IGF-I levels was significantly higher than those of the total IGF-I and IGF binding protein-3 levels. 6) The percent increase in the serum free IGF-I level after 1 month of GH treatment showed a significant positive correlation with that of the GH-induced improvement in the percent increase in the height velocity during 1 yr of GH therapy (r = 0.526; P < 0.05). These results show that free IGF-I in the serum has an essential role in bone formation because the higher free IGF-I levels were observed when the growth rate accelerated. The measurement of serum free IGF-I may become a useful tool for both diagnosing GH deficiency and predicting growth responses to long term GH therapy.

Multifactorial control of the elimination kinetics of unbound (free) growth hormone (GH) in the human: regulation by age, adiposity, renal function, and steady state concentrations of GH in plasma.

To evaluate the principal determinants of the MCR and plasma t1/2 of unbound (free) GH in man, we performed steady state infusions of 3 doses of recombinant human GH during pharmacological suppression (iv octreotide) of endogenous GH secretion in 24 healthy adults and 12 patients (6 adults and 6 children) with chronic renal failure (CRF). Free plasma GH was calculated from total plasma GH (measured by immunoradiometric assay) and GH-binding protein activity (radioligand assay). The MCR of free GH was determined from free plasma GH and the rate of recombinant human GH infusion. The t1/2 of free plasma GH, and the concentration and the in vivo dissociation constant (Kd) of GH-binding protein (GHBP) were estimated by dynamic modeling of the postinfusion total plasma GH concentration decay curves. The MCR of free GH decreased and the plasma GH t1/2 increased significantly with increasing plasma GH concentrations. The MCR of free GH over its physiological concentration range was positively correlated with the body mass index as a measure of relative obesity and negatively related to age, but only at supraphysiological GH concentrations. In the adult patients with CRF, the MCR of free GH was decreased at each infusion rate by 25-38%, and the t1/2 was increased by 80-170%. Children with CRF showed a significantly lower MCR and higher t1/2 of plasma free GH than adult patients. Modeling and direct measurements of the off-rate of GH from its high affinity GHBP indicated normal dissociation rate constants but decreased molar concentrations of the GHBP in uremic plasma. We conclude that the rate of elimination of free GH from plasma in man is controlled by 1) plasma total free GH concentrations, 2) relative obesity, and 3) renal function within the physiological GH concentration range, whereas 4) age is a negative predictor of MCR only at supraphysiological GH concentrations.