Parathyroid hormone-related protein enhances insulin-like growth factor-I expression by fetal rat dermal fibroblasts

Interactions between cells of differing embryonic origins comprise a common theme during tissue development and repair. Often, communication between them can be mediated by soluble growth mediators and in some cases is restricted in focus. That is, some cells respond to, but do not produce, mediators expressed by other cells within the tissue. Because keratinocytes respond to but do not express insulin-like growth factor I (IGF-I), another skin cell population, the dermal fibroblast, may supply this factor. However, keratinocytes express, but do not respond to parathyroid hormone related protein (PTHrp), which increases cAMP production by dermal fibroblasts. Based on earlier results where inducers of cAMP increase local IGF-I expression in skeletal tissue, we postulated that PTHrp might induce local IGF-I by dermal fibroblasts and provide a source of this factor for keratinocyte activity. Our studies reveal that IGF-I mRNA and protein levels increase in response to PTHrp in vitro, and that this effect is replicated by inducers of cAMP, but not by activators of protein kinase C. Consequently, these factors appear to comprise a paracrine loop within the skin, permitting focused but restricted IGF-I expression to support skin growth, remodeling, or repair.     

Expression of insulin-like growth factor-I gene in normal and diabetic rat eye

The expression of insulin-like growth factor I (IGF-I) gene was studied in both normal and diabetic rat eyes via in situ hybridization. The results revealed the regional expression of IGF-I gene in the rat eyes. The expression of IGF-I mRNA in the internal nuclear layer and ganglion cell layer is strong, in choroid, retinal pigment epithelium and external nuclear layer is moderate, in sclera is weak in degree and no such mRNA is detected in the cornea. The abundance of IGF-I mRNA in diabetic eye tissues is significantly (P < 0.05 or P < or = 0.01) higher than that in normal eye. These findings suggest that (1) functionally, the eye ball be considered to be an "IGF-I paracrine-autocrine system", and (2) the high expression of IGF-I indicate its initiation of diabetic retinopathy and its promotion of the progression of the lesion.     

High ratios of free to total insulin-like growth factor-I in early infancy

A case of huge desmoid tumor successfully treated by hyperthermoradiotherapy is described. A 23-year-old man with familial adenomatous polyposis was operated upon for a desmoid tumor in the mesenterium involving the right kidney and small intestine in 1988. In 1990, the tumor recurred and could not be resected because of the involvement of the vena cava. The tumor grew larger and larger, and occupied two-thirds of the right lower quadrant. Several therapies using sulindac, tamoxifen, prednisolone, indomethacin, luteinizing hormone-releasing hormone analogue, and ascorbate were all ineffective. Finally, the combination of radiation and hyperthermia was used over a 6-month period. At the end of the hyperthermoradiotherapy, the tumor in the abdominal wall was markedly reduced in size, and the protruded abdominal wall became flat. To our best knowledge, this is the first report of the successful treatment of a huge desmoid tumor by hyperthermoradiotherapy.     

Correlation between longitudinal bone growth, growth hormone, and insulin-like growth factor-I in prepubertal dogs

OBJECTIVE: To determine the association between longitudinal bone growth and concentrations of growth hormone (GH) and insulin-like growth factor-I (IGF-I) in serum from prepubertal dogs. Animals-6 male 14-week-old German Shepherd Dogs. PROCEDURE: Blood was obtained every 30 minutes for 14 consecutive days. Concentrations of GH and IGF-I in serum were determined, using a canine-specific radioimmunoassay and conventional radioimmunoassay after acid-ethanol extraction, respectively. Simultaneous biplanar radiography was performed daily to measure bone growth. Spectral analysis was used to estimate specific features of GH secretion during an extended period. Multiple linear regression with different lag times between independent and dependent variables was used to determine the strongest predictors of bone growth. RESULTS: The power spectra of GH concentrations in serum had a primary peak at a frequency of 0.02 cycles/h or a periodicity of 50 h/cycle. A significant determinant of longitudinal bone growth was a lag time of 1 day in concentration of GH in serum. The relationship between IGF-I concentration in serum and bone growth was not significant. CONCLUSIONS: The primary frequency of GH secretion is outside the time frame of a single day and the concentration of GH in serum is a primary determinant of bone growth. CLINICAL RELEVANCE: A better understanding of the components of bone growth provide discernment to improved diagnosis and treatment of abnormal bone growth.     

Androgen regulation of the insulin-like growth factor-I and the estrogen receptor in rat uterus and liver

For the first time testosterone is shown to be an important regulator of the insulin-like growth factor-I (IGF-I) in the rat uterus under in vivo conditions. In this study the regulation of IGF-I and the estrogen receptor (ER) by gonadal steroids in the uterus and liver of female rats was monitored. The ER level was assayed by hormone binding after treatment with testosterone, 5 alpha-dihydrotestosterone or estradiol and specific mRNA species were analyzed by a solution hybridization/RNase protection assay using 35S-labeled RNA probes. Ovariectomized rats restored uterine weight after treatment with testosterone. Uterine IGF-I mRNA was more than 20-fold higher in testosterone treated rats compared to untreated ovariectomized controls after 48 h treatment. The effects of testosterone on ovariectomized animals was followed in a timecourse study. Testosterone administration increased uterine IGF-I mRNA expression during the first 48 h and the maximally induced level was maintained throughout the duration of the experiment (168 h). Since induction of IGF-I mRNA by estrogen is transient, these data indicate that androgen and estrogen increase IGF-I mRNA by different mechanisms. Regulation of IGF-I mRNA by gonadal steroids was also studied in hypophysectomized animals. The rats were given either testosterone, 5 alpha-dihydrotestosterone or estradiol, and uterine IGF-I mRNA was measured after 1 week of treatment. At this timepoint estrogen treated rats showed levels of IGF-I mRNA not significantly different from those of hypophysectomized controls. In contrast testosterone and 5 alpha-dihydrotestosterone increased the IGF-I mRNA level 30 and 40 times, respectively, relative to hypophysectomized control animals. Since 5 alpha-dihydrotestosterone is not convertable to estrogen, the induction by testosterone was considered to be a true androgenic phenomenon.     

Role of growth hormone and insulin-like growth factor-I in mammary development

Mammary glands from 3- to 4-week-old mice were incubated in whole organ culture to determine the effects of GH, PRL, and insulin-like growth factor-I (IGF-I) on lobulo-alveolar development and milk protein expression. Virgin mice were implanted with pellets of estrogen and progesterone (1:1000). After 9 days, abdominal no. 4 glands were removed and place on siliconized lens paper in Waymouths' medium supplemented with insulin (Ins), aldosterone, hydrocortisone, and epidermal growth factor. Concentrations of bovine GH, ovine GH, rat GH, or ovine PRL added to the medium varied from 0-1 micrograms/ml. IGF-I was added to replace either Ins or PRL up to 1 microgram/ml. When glands were incubated with Ins, aldosterone, hydrocortisone, and 250 ng/ml PRL, they exhibited lobulo-alveolar development and expressed the milk protein beta-casein. When GH was substituted for PRL, little lobulo-alveolar development occurred, although beta-casein mRNA was expressed at low levels. Either PRL or GH at 1 microgram/ml induced lobulo-alveolar development and beta-casein mRNA. Addition of epidermal growth factor to whole organ culture with GH or PRL (1 microgram/ml) was equally effective in stimulating lobulo-alveolar development. IGF-I did not substitute for PRL, GH, or insulin in tissue maintenance. It is clear that GH at high concentrations can act directly on mouse mammary tissue to induce both lobulo-alveolar development and casein expression.     

Muscle reinnervation following neonatal nerve crush. Interactive effects of glycosaminoglycans and insulin-like growth factor-I

This study shows that glycosaminoglycans promote muscle reinnervation following neonatal sciatic nerve injury. Such an effect appears to be mediated by insulin-like growth factor-1. The glycosaminoglycan moiety of proteoglycans is a constituent of the basal lamina active on nerve regeneration by means of the interaction with laminin and with several growth factors. We have previously shown that supplementation of glycosaminoglycans affects neuronal degeneration and regeneration. In this study we report that following neonatal lesion of the rat sciatic nerve glycosaminoglycan treatment promoted extensor digitorum longus muscle reinnervation with consequent improvement of muscle morphology. In saline-treated rats, reinnervation was only partial and there was a marked muscle fibre atrophy. In addition glycosaminoglycan treatment of lesioned rats increased insulin-like growth factor-I messenger RNA and protein in the reinnervated muscle, and insulin-like growth factor-I and insulin-like growth factor binding protein-3 plasma levels. Similarly, treatment of nerve lesioned rats with insulin-like growth factor-I promoted muscle reinnervation and prevention of muscle fibre atrophy, higher levels of insulin-like growth factor-I in the reinnervated muscle and of insulin-like growth factor-I and insulin-like growth factor binding proteins in plasma. These data suggest that glycosaminoglycans are potent stimulants of muscle reinnervation and that their effects may be mediated by increased levels of insulin-like growth factor-I.     

Low doses of insulin-like growth factor-I improve nitrogen retention and food efficiency in rats with early cirrhosis

N-Methyl-D-aspartate receptors are thought to be involved in synaptic signaling within the hypothalamo-neurohypophysial system, but the extent and nature of their involvement has not been determined. In this study, in the rat, we evaluated the effect of hyperosmotic stimulation on the N-methyl-D-aspartate receptor subunit, NR1, which confers function to N-methyl-D-aspartate receptor heteromers. Co-localization of immunoreactivity for NR1 and vasopressin- or oxytocin-associated neurophysin in magnocellular neurons of the supraoptic and paraventricular hypothalamic nuclei was accomplished using double-label immunohistochemistry. Our results show that vasopressin- and oxytocin-neurophysin-positive populations contained detectable levels of NR1 labeling. Using NR1 labeling as a measure of N-methyl-D-aspartate receptor density, we examined the effect of dehydration in these nuclei. Using computer-assisted densitometry, we found significantly greater NR1 labeling densities in the magnocellular regions of both the supraoptic and paraventricular nuclei of saline-treated rats than of control rats. This increase was not due to methodological factors, since no changes in NR1 labeling density were found in a nearby nucleus, the nucleus reuniens. Western blot analysis showed similar selective increases in NR1 labeling in homogenates from the supraoptic nucleus, paraventricular nucleus and in some cases from the anterior hypothalamic area. In both immunohistochemical and western blotting experiments we did not observe a dehydration-induced increase in NR1 in other brain areas examined. Our results showing an up-regulation of NR1-containing N-methyl-D-aspartate receptors during dehydration suggest that these receptors are involved in the regulation of body water and may represent an adaptive physiological response following activation of the hypothalamo-neurohypophysial axis. In addition, these results suggest that the functional expression of N-methyl-D-aspartate receptors is dynamic and may be modified according to the physiological state of the animal.     

Effects of 9-ene-tetrahydrocannabinol on expression of beta-type transforming growth factors, insulin-like growth factor-I and c-myc genes in the mouse uterus

Effects of cannabinoid on expression of beta-type transforming growth factors (TGF-beta 1, -beta 2 and -beta 3), insulin-like growth factor-I (IGF-I) and c-myc genes in the uteri of adult ovariectomized mice were examined using Northern blot hybridization. Mice were exposed to 9-ene-tetrahydrocannabinol (THC) alone or in combination with an injection of estradiol-17 beta (E2) and/or progesterone (P4), and uteri were analyzed at various times thereafter. TGF-beta isoform messenger RNAs (mRNAs) persisted in ovariectomized uteri and their levels were not altered after THC treatment, whereas an injection of E2 caused a modest increase in TGF-beta 1 and -beta 3 mRNA levels at 24 h. Imposition of THC treatment advanced the stimulatory effects of E2 by changing the timing for the peak of TGF-beta 3 mRNA levels to 12 h. In comparison, E2 treatment substantially elevated the levels of TGF-beta 2 mRNA at 6 h, and THC potentiated this E2 response without affecting the timing for the response. Imposition of P4 treatment did not antagonize any of these responses. P4 treatment alone or with THC had insignificant effects on mRNA levels for these TGF-beta isoforms. Uterine levels of IGF-I and c-myc mRNAs were low in ovariectomized mice and THC did not alter these mRNA levels. In contrast, E2 treatment induced a rapid, but transient, increase in IGF-I and c-myc mRNAs, and THC antagonized the rapid c-myc mRNA response and altered the timing of the IGF-I mRNA response. P4 treatment alone also caused the transient induction of these mRNAs, but THC failed to antagonize these effects. An injection of P4 plus E2 resulted in further modest increases in IGF-I and c-myc mRNA levels as compared to E2 or P4 treatment alone. However, THC did not antagonize these transient stimulatory effects of the combined ovarian steroids. The data suggest that THC should not be classified as estrogenic or antiestrogenic. However, this compound can modulate (potentiate, antagonize and/or alter timing) the effects of ovarian steroids on uterine gene expression.     

Histomorphometric, physical, and mechanical effects of spaceflight and insulin-like growth factor-I on rat long bones

Pituitary adenylate cyclase activating peptide and vasoactive intestinal peptide belong to the same neuropeptide family. Both peptides are present in nerve fibers in the gastric wall and are thought to be involved in the regulation of inflammatory processes. Experimental ulcers were induced in the rat gastric mucosa by local application of acetic acid. During the healing process we examined the PACAP and VIP innervation by means of immunohistochemistry and in situ hybridization. The ulcer area was examined from day 1 to day 15 after ulcer induction. There was a marked depletion of PACAP in nerve fibers at the ulcer margin from day 1 and onwards. On day 10 and day 15, PACAP-immunoreactive nerve fibers could again be visualized at the ulcer margin. In contrast, VIP immunoreactive nerve fibers were present at the ulcer margin at all time points studied. From day 10 following ulcer induction PACAP- and VIP- immunoreactive nerve fibers were increased in frequency in the smooth muscle beneath the ulcer. An upregulation of VIP and PACAP mRNA was also demonstrated in the myenteric ganglia adjacent to ulcer. The present results indicate that neuronal PACAP and VIP react differently to the inflammation at the ulcer margin but similarly in the smooth muscle during the ulcer healing.     

Blockage of insulin-like growth factor-I receptor inhibits the growth of Ewing's sarcoma in athymic mice

Innovative, more effective treatment modalities are needed for Ewing's sarcoma (ES), a neoplasm with a disappointingly low survival rate despite the use of aggressive multimodal therapeutic approaches. We have previously shown (K. Scotlandi et al, Cancer Res., 56: 4570-4574, 1996) the existence and the pathogenetic relevance of an autocrine loop that is mediated by the insulin-like growth factor-I receptor (IGF-IR) and is crucial for the survival and proliferation of ES cells in vitro. In this study, we report that the IGF-IR-blocking monoclonal antibody alphaIR3 may also significantly inhibit ES cell growth in vivo. In particular, in almost one-half of the animals tested, after s.c. inoculation with TC-71 ES cells, the blockage of IGF-IR by alphaIR3 induced a complete regression of tumors that developed, which suggests that IGF-IR is valuable as a specific target for novel therapeutic strategies. In addition, suramin, a drug that can interfere with growth factor binding with their receptors, inhibited the tumorigenic and the metastatic ability of TC-71 cells and, therefore, is a promising agent to be combined with conventional cytotoxic drugs for the design of more effective therapeutic regimens.     

Refeeding increases hepatic insulin-like growth factor-I (IGF-I) gene expression and plasma IGF-I concentration in fasted chicks

This study compared radiologic and ultrasonographic methods of evaluation of patella position. The radiologic examination was based on the evaluation of Insall and Salvati's index (I-S index), whereas the ultrasonographic examination was based on the determination of analogous coefficient called the patellar tendon-patellar coefficient (T-P coefficient). The total number of examined knee joints was 55 in 30 patients (13 children, aged 7-16 years and 17 adults aged 17-39 years) with knee pain. Considerable differences of the evaluated parameters were observed in the group of examined children: I-S index, 1.50; T-P coefficient, 1.20; and small differences in the group of adults: 1.17 and 1.32, respectively. Those differences resulted from difficulties with interpretation of the apparent radiologic picture of the knee joint with the patella incompletely ossified. The ultrasonographic picture in both children and adults is a real picture, and the possibilities of its interpretation are independent of the degree of patellar ossification.     

Growth hormone, insulin-like growth factor-I and insulin-like growth factor binding protein-3 are regulated differently in small-for-gestational-age and appropriate-for-gestational-age neonates

BACKGROUND: TBP-associated factors contain a variety of structural motifs and their related in vivo significance has remained unclear. We have attempted to identify specific biological phenomena linked to a particular domain of a TAF by analysing domain-exchanged chimeric mutants between Schizosaccharomyces pombe (Sp) and Saccharomyces cerevisiae (Sc) counterparts. RESULTS: Contrary to the case of TBP, Sp TAF containing the WD40 repeat cannot be exchanged for its Sc counterpart, despite their highly conserved primary structures. This 'species-specific' function locates in the N-terminal region. The C-terminal region, largely consisting of the WD40 repeat, is exchangeable for the corresponding region of its Sc counterpart. Growth of the strain harbouring this C-terminal chimeric mutant is temperature-sensitive. The chimeric gene product did not disappear at a restrictive temperature, a finding which strongly suggests that the growth defect is caused by an aberration in the interactions through the WD40 repeat structural motif. With temperature elevation, the chimeric mutants underwent drastic morphological changes due to a defect in cytokinesis. CONCLUSIONS: The WD40 repeat of TAF is primarily involved in reactions which might regulate cytokinesis in Sp.     

Increased expression of platelet-derived growth factor beta receptor gene in hypoxic rat lungs

The changes of platelet-derived growth factor (PDGF) beta receptor gene expression in hypoxic rats lungs was examined. Northern blots analysis revealed that normal lungs expression PDGF-beta receptor mRNA, with the longer of hypoxia the level of the mRNA increased rapidly. It reached a maximum at day 4, and was 1.34 fold as compared with the control (P < 0.05). Immunohistochemistry investigation showed that PDGF-beta receptor mainly distributed on smooth muscle cells and endothelial cells of middle and small arterial trees in rat lung. With hypoxia, the distribution of PDGF-beta receptors did not change, but it was more intense and reached a maximum at day 7, and was 2.40 fold as compared with the control (P < 0.05). The results suggested that increased expression of PDGF receptor gene may play a role in hypoxic pulmonary vascular remodeling.     

Insulin-like growth factor-I expression in normal and diseased endometrium

While the role of steroid hormones in the regulation of endometrial proliferation and differentiation is well established, the effects of growth factors and their receptors in normal and neoplastic endometrium remain a matter of debate. Previous studies have documented the positive effects of insulin-like growth factor-I (IGF-I) on epithelial cell proliferation and the active production of this growth factor in endometrial tissues. In view of decreased expression of transforming growth factor-beta1 (TGF-beta1), an antagonist of IGF-I, in endometrial carcinoma, we investigated the expression of IGF-I, at both the mRNA and protein levels, and the immunoreactivity for type I IGF-I receptor in 30 formalin-fixed, paraffin-embedded tissue samples of normal and neoplastic endometrium, in order to possibly clarify the role of IGF-I in endometrial proliferation and differentiation. Our results demonstrate a reduced expression of IGF-I mRNA in endometrial carcinomas compared with non-neoplastic tissues, despite equivalent immunohistochemical expression of IGF-I and IGF-I receptor. Our data suggest that IGF-I and its corresponding receptor may not be directly involved in endometrial cancer cell proliferation and differentiation in vivo, though other components of the IGF-I system (e.g., IGF binding proteins) may affect endometrial malignant transformation and tumor progression.     

Interplay of the proto-oncogene proteins CrkL and CrkII in insulin-like growth factor-I receptor-mediated signal transduction

The closely related proto-oncogene proteins CrkII and CrkL consist of one SH2 and two SH3 domains and share 60% overall homology with the highest identity within their functional domains. In this study we show that CrkL and CrkII may play overlapping but different roles in insulin-like growth factor (IGF)-I receptor-mediated signal transduction. While both proteins are substrates involved in IGF-I receptor signaling, they apparently demonstrate important different properties and different biological responses. Evidence supporting this hypothesis includes (a) the oncogenic potential of CrkL versus the absence of this potential in CrkII overexpressing cell lines, (b) the inhibition of IGF-I-dependent cell cycle progression by overexpression of CrkII, and (c) the differential regulation of the phosphorylation status of selective proteins in CrkII and CrkL overexpressing cell lines. In addition we demonstrate the specific association of CrkL and CrkII with the newly characterized IRS-4 protein, again in a differential manner. Whereas CrkL strongly interacts with IRS-4 via its SH2 and N-terminal SH3 domains, CrkII interacts only via its SH2 domain, possibly explaining the unstable nature of IRS-4-CrkII association. The results obtained allow us to propose a unique mechanism of CrkL and CrkII tyrosine phosphorylation in response to IGF-I stimulation. Thus these highly homologous proteins apparently possess structural features that allow for the differential association of each protein with different effector molecules, thereby activating different signaling pathways and resulting in unique biological roles of these proteins.