Human growth hormone releasing factor and somatostatin from two pancreatic tumors: isolation and characterization

Peptides with high intrinsic activity to release growth hormone from pituitary cells in tissue cultures were isolated from two different human pancreatic tumors that had caused acromegaly. Homogeneous peptides were obtained after gel filtration and two steps of reverse-phase high-performance liquid chromatography. From one tumor a 44-residue peptide (human pancreas growth hormone releasing factor, hpGRF-44) was isolated, together with two shorter fragments of reduced bioactivity having 40 and 37 amino acid residues (hpGRF-40, hpGRF-37). In contrast, the other tumor contained only one form of GRF which proved to be identical to hpGRF-40. These hpGRFs are indistinguishable from partially purified preparations of hypothalamic growth hormone releasing factor of human, porcine and murine origins with respect to biological activity and are very similar in their physicochemical properties (molecular weight, retention behavior on reverse-phase HPLC, absence of sulfhydryl groups). One of the pancreatic tumors also contained two forms of immunoreactive somatostatin. One form, after isolation and partial microsequencing, was identified as somatostatin-14 with a structure identical to that of the peptide found in other species. The second form has tentatively been identified as somatostatin-28 on the basis of chromatographic behavior.     

Immunoreactive neuronal pathways of growth hormone-releasing hormone (GRH) in the brain and pituitary of the teleost Gadus morhua

Summary Using an antiserum directed against the C-terminus of hGRH(1–44)NH₂ and another recognizing the mid portion to C-terminal of hGRH(1–40)OH, we identify two immunocytochemically distinct GRH-immunoreactive systems in the brain of the codfish, Gadus morhua. The antiserum directed against GRF(1–44)NH₂ stains cell bodies exclusively in the rostral pars distalis. The other antiserum immunoreactive with GRF(1–40)OH reacts with a population of parvocellular and magnocellular neuronal cell bodies in the hypothalamus and with two major axonal pathways which project toward the median eminence and terminate primarily in the pars nervosa. These results indicate the presence of at least two forms of hGRH-like peptides in the teleost which may have different roles in the regulation of pituitary function.     

Plasma growth hormone secretion during constant growth hormone-releasing factor infusion

Synthetic human pancreatic growth hormone-releasing factor (hpGRF-44) was infused intravenously at a constant rate of 2.5 micrograms/min for 180 minutes in 3 normal boys of short stature. Plasma GH levels reached a peak at 60-120 min with a mean value (+/- SEM) of 69.1 +/- 14.3 ng/ml, and then, declined gradually in spite of continuous hpGRF-44 infusion up to 180 minutes. Similarly, constant infusion of hpGRF-44 at a rate of 2.5 micrograms/min in 5 normal but short boys for 90 minutes, together with an iv bolus injection of hpGRF-44 (2 micrograms/kg) administered at 0 and 90 minutes, elicited a prompt rise in plasma GH 15-30 minutes after the first bolus but no significant elevation of GH was observed after the second bolus. In contrast, when two iv bolus injections of hpGRF-44 (2 micrograms/kg) were given in 4 normal boys with short stature at 0 and 90 minutes, respectively, significant elevation of plasma GH was found after each bolus. These results suggest that under constant infusion of GRF the pituitary experiences a down-regulation after the initial peak of GH response, possibly due to desensitization to GRF.     

Growth hormone releasing effect of hpGRF-40 in rats at different time intervals following ablation of the mediobasal hypothalamus

We have investigated the effect of hypothalamo-pituitary disconnection in the rat on the growth hormone (GH) responsiveness to human pancreatic GH-releasing factor (hpGRF). Adult female rats, sham-operated (sham-op) or bearing a complete mechanical ablation of the mediobasal hypothalamus (MBH-A) were challenged, while under urethane anesthesia, with hpGRF-40 (20,100,500 ng/rat i.v.) at different time intervals after surgery. In sham-op rats only 500 ng/rat of hpGRF-40 stimulated GH release, while in 1-and 7-day MBH-A rats the stimulation also occurred with the lower hpGRF doses and the rise in plasma GH was greater than in sham-op controls. Twenty-one and 42 days after the placing of the lesions the GH response to hpGRF-40 was still present at the 500 ng/rat dose, though it was smaller than in sham-op controls. Evaluation of pituitary GH content demonstrated a progressive and rapid decline starting the first day after the placing of the lesions. These data indicate that GH responsiveness to hpGRF is: 1) enhanced in the anterior pituitary shortly after hypothalamo-pituitary disconnection and, 2) despite a striking reduction of the pituitary GH stores, it is maintained after these lesions.The physiologic growth hormone (GH) releaser in the rat is GH-releasing factor and, recently, a group of peptides has been characterized from human pancreatic tumors (hpGRFs) (1,2) which are potent and specific GH-releasers in both animals (3) and man (4). The availability of these peptides, which show a high degree of homology with the physiologic rat hypothalamic GRF (5), offers the unique opportunity to assess somatotrope responsiveness to GRF molecules in rats with hypothalamo-pituitary disconnection.In this study we have first evaluated the GH pituitary responsiveness to increasing doses of hpGRF-40 in rats following mechanical ablation of the mediobasal hypothalamus (6). These rats, by definition, lack the effect of both central nervous system (CNS) inhibitory (e.g. somatostatin) and stimulatory (e.g. GRF) influences to GH release. With the aim to ascertain how the lack of these two opposing inputs reflects on the secretory capacity of the somatotropes, we also investigated the GH response to hpGRF-40 at different time intervals after the lesioning. In a study in rats with electrolytic lesions of the ventromedial-arcuate region of the hypothalamus Tannenbaum et al (7) had shown persistence of the GH response to huge doses of a hpGRF analog.     

Topographical and ontogenetic study of the neurons producing growth hormone-releasing factor in human hypothalamus

Neurons producing growth hormone-releasing factor have been characterized and analyzed by immunohistochemistry in the hypothalami of human fetuses, neonates, infants and adults, using two antibodies against human pancreatic GRF (hpGRF). One of the antibodies recognized both the hpGRF(1-40)OH and hpGRF(1-44)NH2 in the mid portion (between the 28th and 39th amino acid), the other one specifically recognized the C-terminal end of hpGRF(1-44)NH2. These two antibodies stain a single neuronal system with cell bodies mainly located in the infundibular (arcuate) nucleus, and in the ventromedial and lateralis tuber nuclei. These neurons project to the median eminence where they give numerous endings in contact with portal vessels. These neurons are distinct from those containing LH-RH, somatostatin, CRF or pro-opiocortin. In fetuses, neurons immunoreactive with hpGRF antibodies are first detected at the 29th week. They display a neuroblastic aspect which persists after birth. Immunoreactive fibers are detectable in the median eminence after the 31st week. These results demonstrate that the infundibular nucleus plays a major role in control of GH secretion in man and that secretion of GRF appears late during fetal life; this suggests that the first stages of differentiation and development of GH producing cells in the human fetus do not depend on hypothalamic GRF secretion.     

Pituitary response to growth hormone-releasing factor in rats with functional or anatomical lesions of the central nervous system that inhibit endogenous growth hormone secretion

The pituitary growth hormone (GH) response to the growth hormone-releasing factor, hpGRF-44, was evaluated in male rats with various lesions of the central nervous system. These included an electrical lesion of the ventromedial hypothalamus, a chemical lesion of the arcuate nucleus induced by neonatal treatment with monosodium glutamate, a functional lesion of catecholamine synthesis with alpha-methyl-p-tyrosine or a functional lesion of catecholamine storage with reserpine. The first three lesions appear to partially inhibit normal somatostatin secretion since in every instance hpGRF-44 administration induced a significant increase in plasma GH concentrations. In contrast, reserpine blocked the GH response to hpGRF-44, presumably by stimulating somatostatin secretion. The pituitary GH response to hpGRF-44 in the above described models was enhanced by pretreatment of the rats with antibodies against somatostatin. The pituitary GH response to repeated injections of hpGRF-44 was also evaluated in rats with an anatomical lesion of the arcuate nucleus or a functional lesion of catecholamine synthesis. The maximum GH response did not vary over time to the repeated injections of hpGRF-44 in rats with lesions of the arcuate nucleus; however, interruption of catecholamine synthesis resulted in a significant decrease in the GH response to hpGRF-44 over time.     

Specific binding of synthetic human pancreatic growth hormone releasing factor (1-40-OH) to bovine anterior pituitaries

We have studied the specific binding of a synthetic 40 amino acid, free carboxy terminus analog of human pancreatic growth hormone releasing factor (hp GRF-40-OH) to partially purified homogenates of bovine anterior pituitaries. The binding of hpGRF-40-OH to pituitary receptors at 4 degrees C reached maximal level in 4 hours and remained steady for the next 18 hours. Specific binding increased linearly with the amount of protein present in the assay. 125I-hpGRF-40-OH binding to pituitary homogenates was competitively inhibited by hpGRF-40-OH but not by unrelated hormones. The competition curve and Scatchard analysis suggest the presence of single class of receptors with a Kd congruent to 3nM and binding capacity of approximately 200 fmoles/mg protein. This is the first demonstration of specific receptors for GRF on anterior pituitary cells.     

Growth hormone releasing factor (GRF) immunoreactivity in human and rat gastrointestinal tract and pancreas

Rabbit antisera were raised against a synthetic growth hormone releasing factor, which was originally isolated from a human pancreatic endocrine tumor (hpGRF-44). The antisera obtained showed no significant cross-reactivity with a variety of neurohormonal peptides. In addition to its occurrence in the human, but not in the rat, hypothalamus, hpGRF-44-like immunoreactivity was identified in human gastric antrum and human as well as rat pancreatic islets, using an indirect immunoperoxidase technique. Staining of serial sections and double staining revealed that in the gastric antrum the immunoreactivity was largely confined to gastrin (G) cells, whereas in pancreatic islets polypeptide (pp) cells were reactive. The physiological significance of these findings remains to be established.     

Growth hormone, thyrotropin and prolactin responses to simultaneous administration of human growth hormone-releasing factor and thyrotropin releasing hormone in the bovine

The effects of intravenous injection of synthetic human pancreatic growth hormone-releasing factor-44-NH2 (hpGRF-44) and synthetic thyrotropin releasing hormone (TRH), or hpGRF-44 in combination with TRH on growth hormone (GH), thyrotropin (TSH), and prolactin (PRL) release in dairy female calves (6- and 12-month-old) were studied. When 0.25 microgram of hpGRF-44 per kg of body weight (bw) was injected in combination with TRH (1.0 microgram per kg of bw), the mean plasma GH concentration of the 12-month-old calves rose to a maximum level of 191.5 ng/ml (P less than 0.001) at 15 min from the value of 6.8 ng/ml before injection at 0 min. The maximum level was 3.1 and 6.1 times as high as the peak values obtained after injection of hpGRF-44 (0.25 microgram per kg of bw) and TRH (1.0 microgram per kg of bw), respectively (P less than 0.001). The area under the GH response curve for the 12-month-old calves for 3 hr after injection of hpGRF-44 in combination with TRH was 2.5 times as large as the sum of the areas obtained by hpGRF-44 and TRH injections. In contrast, the mean plasma GH level was unchanged in saline injected calves. The magnitudes of the first and the second plasma GH responses in the 6-month-old calves to two consecutive injections of hpGRF-44 in combination with TRH at a 3-hr interval were very similar. The peak values of plasma GH in the calves after hpGRF-44 injection were 2-4 times as high as those after TRH injection.(ABSTRACT TRUNCATED AT 250 WORDS)     

Production of antisera to growth hormone-releasing factor: usefulness in radioimmunoassay and passive immunization

We have produced two antisera (R-1 & R-2) to human growth hormone-releasing factor (GRF) [1-44] NH2. Both antisera can be used for human GRF radioimmunoassay (RIA) at a final dilution of 1:50000. The antiserum R-2 was specific for the C-terminal amidated sequence of human GRF-44 and selectively recognized GRF [1-44] NH2 but not GRF [1-44] OH or GRF [1-40] OH. The antiserum R-1 also significantly bound 125I-rat GRF [1-43] OH at a final dilution of 1:5000 and enabled us to establish RIA for rat GRF. In both RIA systems, intra- and inter-assay coefficients of variation at 50% inhibition were 8 and 12%, respectively. A median effective dose was 90-120 pg in human GRF RIA and 250-300 pg in rat GRF RIA. Utilizing the RIA, we demonstrated that the hypothalamic GRF content in rats which received monosodium glutamate during the neonatal period was less than 20% of that of controls. However, the hypothalamic GRF content was not altered in rats made hypothyroid by methimazole administration, another condition known to greatly impair GH secretion. An iv administration of the antiserum R-1 significantly suppressed GH release following the injection of antisomatostatin serum. Thus, these antisera can be a useful tool in examining the physiological and/or pathophysiological roles of GRF in human and rat.     

Growth hormone response to human pancreatic growth hormone releasing factor in cattle

The effects of a growth hormone releasing factor, human pancreatic growth hormone releasing factor-44 (hpGRF-44), on growth hormone (GH) secretion in calves, heifers and cows were studied. A single intravenous (iv) injection of 0.1, 0.25, 0.5 or 1.0 microgram of synthetic hpGRF-44 per kg of body weight (bw) in calves significantly elevated the circulating GH level within 2-5 min, while no increase in plasma GH was observed in saline injected control calves. The plasma GH level increased proportionally to the log dose of hpGRF-44, and reached a peak at 5-10 min (p less than 0.01). Subcutaneous injection of hpGRF-44 also elevated the plasma GH level, but the peak value at 15 min was 37% of that of iv injection (p less than 0.05). Intravenous injection of 0.25 microgram of hpGRF-44 per kg of bw to female calves, heifers, and cows significantly elevated mean the GH levels from 8.5, 2.3, and 1.6 ng/ml at 0 time to peak values of 97, 26, and 11.6 ng/ml, respectively (p less than 0.01). The plasma GH response and basal level in calves were significantly higher than those of heifers or cows (p less than 0.025). The plasma GH response to hpGRF-44 as well as the basal level decreased with advancing age. The plasma GH response to hpGRF-44 and basal GH in male calves were significantly greater than those in female calves (p less than 0.001). These results indicate that synthetic hpGRF-44 is a potent secretogogue for bovine GH, and suggest its usefulness in the assessment of GH secretion and reserve in cattle.     

The dual nature of the reaction between porcine elastase and human plasma alpha1 proteinase inhibitor.

Only a portion of human plasma α₁ proteinase inhibitor (α₁PI) forms a 1:1 complex with porcine elastase; the other portion is inactivated via proteolysis. High temperature (37°) and high salt (2 M) enhance complex formation. The complex is unstable, but no significant liberation of active elastase could be demonstrated. Probably the same two major products of ～50,000 and ～4,000 daltons are formed from α₁PI via proteolysis and via disintegration of the complex. Iodination of α₁PI or oxidation with chloramine-T prevents complex formation with elastase but not with trypsin. Iodinated elastase, however, forms a complex with α₁PI.     

Ontogenetic appearance of immunoreactive GRF-containing neurons in the rat hypothalamus

Summary Ontogenetic development of GRF-containing neurons in the rat hypothalamus was studied employing antisera which were generated against hpGRF (1–44)NH₂ and rhGRF(1–43)OH: anti-hpGRF-C and -rhGRF sera recognize the species-specific C-terminal portions of the peptides, and anti-hpGRF-MC and -N sera recognize hpGRF(27–44)NH₂ and the N-terminal portion of hpGRF(1–44)NH₂, respectively. The anti-hpGRF-C and-rhGRF sera stained different neuronal cell bodies, which were localized in distinct hypothalamic areas. The former serum did not stain the axonal terminals in the median eminence, but the latter stained them strongly. The antihpGRF-MC and -N sera stained neuronal cell bodies, some of which corresponded to those immunolabelled with antihpGRF-C or -rhGRF serum. The anti-rhGRF serum first demonstrated immunoreactive perikarya in the ventral-lateral border of the arcuate nucleus of 19.5-day-old fetuses that had received an intraventricular colchicine administration 24 h previously. The immunoreactive fibers were recognized first in the external layer of the median eminence of untreated fetuses on day 19.5 of gestation, and then they increased in amount with development. No immunore-active fibers, however, were found in the median eminence of colchicine-treated animals during the fetal period. It is concluded that in rats GRF may be synthesized in the perikarya on day 18.5 of gestation and conveyed to the median eminence without delay via axonal flow.     

Ultrashort cationic lipopeptides and lipopeptoids: Evaluation and mechanistic insights against epithelial cancer cells

Peptides present an attractive scaffold for the development of new anticancer lead agents due to their accessibility and ease of modification. Synthetic ultrashort cationic lipopeptides, with four amino acids or less conjugated to a fatty acid, were developed to retain the biological activity of longer peptides in a smaller molecular size. Herein, we report the activity of amphiphilic lipotripeptides, lipotripeptoids and lipotetrapeptides against breast (MDA-MB-231, JIMT-1), prostate (DU145) and pancreas (MiaPaCa2) epithelial cancer cell lines. The lipotripeptide C16-KKK-NH₂ and lipotetrapeptide C16-P_CatP_HexP_HexP_Cat-NH₂ were identified to possess anticancer activity. The latter lipotetrapeptide possess a short polyproline scaffold consisting of only two L-4R-aminoproline (P_Cat) and two L-4R-hexyloxyproline (P_Hex). However, all the prepared lipotripeptoids lack anticancer activity. The amphiphilic C16-P_CatP_HexP_HexP_Cat-NH₂ exhibited similar anticancer potency to the surfactant benzethonium chloride while superior activity was observed in comparison to myristylamine. Mechanistic studies revealed that the peptides do not lyse ovine erythrocytes nor epithelial cancer cells, thus ruling out necrosis as the mechanism of cell death. Surprisingly, the two lipopeptides exhibit different mechanisms of action that result in cancer cell death. The lipotripeptide C16-KKK-NH₂ was found to induce caspase-mediated apoptosis while C16-P_CatP_HexP_HexP_Cat-NH₂ kills tumor cells independent of caspases.     

Monoclonal antibodies against simian virus 40 nuclear large T tumour antigen: epitope mapping, papova virus cross-reaction and cell surface staining.

Thirty six cloned hybridomas have been isolated which produce monoclonal antibodies directed against simian virus 40 (SV40) large T tumour antigen. They have been shown to recognize at least six different epitopes along the T antigen polypeptide according to their reaction with the various truncated forms of T antigen expressed by adenovirus-SV 40 hybrid viruses. Sixteen antibodies cross-react with cells infected by the closely related human BK virus. Only two antibodies, PAb1604 and PAb1614, directed against different epitopes of the SV40 T antigen, cross-react with polyoma large T tumour antigen which has a more limited amino acid sequence homology. This cross-reaction is rarely seen with polyclonal antibodies. Monoclonal antibody PAb1620 gave nuclear immunofluorescence only with murine cells transformed by SV40 and was found to react with a complex of T-antigen and 53 000-dalton host-coded protein. All the monoclonal antibodies react with nuclear T antigen and all but four antibodies stained the surface of SV40-transformed cells. These were four of the five antibodies directed against the central third of the T antigen. Thus the monoclonal antibodies show that cell surface T antigen differs from nuclear T antigen, either in accessibility or structure.     

A mastoparan-derived peptide has broad-spectrum antiviral activity against enveloped viruses

Broad-spectrum antiviral drugs are urgently needed to treat individuals infected with new and re-emerging viruses, or with viruses that have developed resistance to antiviral therapies. Mammalian natural host defense peptides (mNHP) are short, usually cationic, peptides that have direct antimicrobial activity, and which in some instances activate cell-mediated antiviral immune responses. Although mNHP have potent activity in vitro, efficacy trials in vivo of exogenously provided mNHP have been largely disappointing, and no mNHP are currently licensed for human use. Mastoparan is an invertebrate host defense peptide that penetrates lipid bilayers, and we reasoned that a mastoparan analog might interact with the lipid component of virus membranes and thereby reduce infectivity of enveloped viruses. Our objective was to determine whether mastoparan-derived peptide MP7-NH₂ could inactivate viruses of multiple types, and whether it could stimulate cell-mediated antiviral activity. We found that MP7-NH₂ potently inactivated a range of enveloped viruses. Consistent with our proposed mechanism of action, MP7-NH₂ was not efficacious against a non-enveloped virus. Pre-treatment of cells with MP7-NH₂ did not reduce the amount of virus recovered after infection, which suggested that the primary mechanism of action in vitro was direct inactivation of virus by MP7-NH₂. These results demonstrate for the first time that a mastoparan derivative has broad-spectrum antiviral activity in vitro and suggest that further investigation of the antiviral properties of mastoparan peptides in vivo is warranted.     

Effects of glycine-extended and serine13-phosphorylated forms of peptide YY on food intake in rats

The gut hormone peptide YY(3-36)-amide [PYY(3-36)-NH₂] is significantly more potent than PYY(1-36)-NH₂ in reducing food intake in rats and humans. Other Gly-extended and Ser¹³-phosphorylated PYY forms have been detected or predicted based upon known cellular processes of PYY synthesis and modification. Here we compared the effects of 3-h IV infusion of PYY(1-36)-NH₂, PYY(3-36)-NH₂, PYY(1-36)-Gly-OH, PYY(3-36)-Gly-OH, Ser¹³(PO₃)-PYY(1-36)-NH₂, Ser¹³(PO₃)-PYY(3-36)-NH₂, Ser¹³(PO₃)-PYY(1-36)-Gly-OH, and Ser¹³(PO₃)-PYY(3-36)-Gly-OH during the early dark period on food intake in freely feeding rats. PYY(3-36)-NH₂ and Ser¹³(PO₃)-PYY(3-36)-NH₂ reduced food intake similarly at 50 pmol/kg/min, while only PYY(3-36)-NH₂ reduced food intake at 15 pmol/kg/min. PYY(1-36)-NH₂ and Ser¹³(PO₃)-PYY(1-36)-NH₂ reduced food intake similarly at 50 and 150 pmol/kg/min. In contrast, PYY(1-36)-Gly-OH, PYY(3-36)-Gly-OH, Ser¹³(PO₃)-PYY(3-36)-Gly-OH, and Ser¹³(PO₃)-PYY(1-36)-Gly-OH had no effect on food intake at doses of 50 or 150 pmol/kg/min. Taken together, these results indicate that (i) PYY(3-36)-NH₂ is significantly more potent than PYY(1-36)-NH₂ in reducing food intake, (ii) Gly-extended forms of PYY are significantly less potent than non-extended forms, and (iii) Ser¹³-phosphorylation of PYY(3-36)-NH₂ decreases the anorexigenic potency PYY(3-36)-NH₂, but not PYY(1-36)-NH₂. Thus, PYY(3-36)-NH₂ appears to be the most potent PYY form for reducing food intake in rats.     

Monoclonal antibodies against two separate alloantigenic sites of HLA-1340

Two monoclonal antibodies (MB40.2 and MB40.3) which are highly specific for HLA-B40 and HLA-B7 were made. They appear to be directed against two separate alloantigenic sites of these HLA molecules. Semiquantitative analysis of the kinetics of antibody binding show that MB40.2 recognizes a site which shows a degree of cross-reactivity with B7 and is specific to some B40 molecules. This antibody also distinguishes between different molecules typed as B40. In contrast, MB40.3 recognizes an antigenic determinant which is less variable between B7 and B40 and more closely approximates a public antigen or common antigenic site. This study suggests that the introduction of monoclonal antibodies into MHC serology not only permits but demands a quantitative analysis of these complex systems of homologous but highly polymorphic molecules.Abbreviations used in this paper MHC major histocompatibility complex - IgG immunoglobulin G - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis - PBS phosphate-buffered saline - BSA bovine serum albumin - FCS fetal calf serum - RAM rabbit anti-mouse IgG F(ab)₂ fragments For convenience the terms HLA antigen and H-2 antigen will refer only to the ₂-microglobulin-associated, class I molecules coded for by the major histocompatibility complex of man and the mouse, respectively.     

His‐tag binding by antibody C706 mimics β‐amyloid recognition

Alzheimer's disease is a progressive neurodegenerative disease characterized by extracellular deposits of β‐amyloid (Aβ) plaques. Aggregation of the Aβ₄₂ peptide leading to plaque formation is believed to play a central role in Alzheimer's disease pathogenesis. Anti‐Aβ monoclonal antibodies can reduce amyloid plaques and could possibly be used for immunotherapy. We have developed a monoclonal antibody C706, which recognizes the human Aβ peptide. Here we report the crystal structure of the antibody Fab fragment at 1.7 Å resolution. The structure was determined in two crystal forms, P2₁ and C2. Although the Fab was crystallized in the presence of Aβ₁₆, no peptide was observed in the crystals. The antigen‐binding site is blocked by the hexahistidine tag of another Fab molecule in both crystal forms. The poly‐His peptide in an extended conformation occupies a crevice between the light and heavy chains of the variable domain. Two consecutive histidines (His4–His5) stack against tryptophan residues in the central pocket of the antigen‐binding surface. In addition, they form hydrogen bonds to the acidic residues at the bottom of the pocket. The mode of his‐tag binding by C706 resembles the Aβ recognition by antibodies PFA1 and WO2. All three antibodies recognize the same immunodominant B‐cell epitope of Aβ. By similarity, residues Phe–Arg–His of Aβ would be a major portion of the C706 epitope. Copyright © 2010 John Wiley & Sons, Ltd.     

Plasma growth hormone responses to repetitive administrations of growth hormone releasing factor in patients with pituitary dwarfism

Plasma growth hormone (GH) responses to the repetitive administrations of synthetic human pancreatic growth hormone releasing factor (hpGRF-44) were studied in 15 patients with GH deficiency (11 diagnosed as idiopathic and 4 diagnosed as secondary to hypothalamo-pituitary tumor). hpGRF-44 was administered by single iv bolus (2 micrograms/kg), repetitive im (100 micrograms, twice a day), and/or repetitive iv infusion (2.5 micrograms/min for 90 min, once a day) for three to six consecutive days. Three of the eleven idiopathic GH deficient patients had plasma GH responses to both single iv bolus injection and repetitive administrations by im, or iv infusion of hpGRF. In four of the remaining eight, who had not had peak plasma GH levels above 5 ng/ml to a single iv bolus of the peptide, repetitive administrations of hpGRF-44 by im injection and/or iv infusion induced GH responses to the peptide. In the four patients with secondary GH deficiency, three had plasma GH response to hpGRF administration but one patient, who had indications of pituitary disorder, did not show any plasma GH response to either single iv injection or repetitive administrations of hpGRF-44. These data show that repetitive administrations of hpGRF-44 can induce plasma GH responses in some GH deficient patients who do not respond to a single iv bolus of the peptide.