Cartilage oligomeric matrix protein increases in serum after the start of growth hormone treatment in prepubertal children.

Both GH and IGF-I stimulate bone growth, but the molecular mechanisms mediating their effects on the growth plate are not fully understood. We measured gene expression by microarray analysis in primary cultured human chondrocytes treated with either GH or IGF-I. One of the genes found to be up-regulated by both GH and IGF-I was that encoding cartilage oligomeric matrix protein (COMP). This protein is predominantly found in the extracellular matrix of cartilage. Mutations in the COMP gene have been associated with syndromes of short stature. To verify that COMP is regulated by GH in vivo, we measured COMP levels in serum in short children treated with GH. The study included 113 short prepubertal children (14 girls and 99 boys) with a mean (+/- sd) age of 8.84 +/- 2.76 yr, height sd score of -2.74 +/- 0.67, and IGF-I sd score of -1.21 +/- 1.07 at the start of GH administration. Serum levels of COMP were 1.58 +/- 0.28, 1.83 +/- 0.28 (P < 0.0001), 1.91 +/- 0.28 (P < 0.0001), 1.78 +/- 0.28 (P < 0.001), and 1.70 +/- 0.24 (P < 0.05) microg/ml at baseline and after 1 wk and 1, 3, and 12 months, respectively. In conclusion, we have demonstrated that COMP expression is up-regulated by both GH and IGF-I in primary cultured human chondrocytes. Furthermore, serum levels of COMP increase after the start of GH treatment in short children.

Growth hormone treatment prevents the decrease in insulin-like growth factor I gene expression in patients undergoing abdominal surgery.

Acquired GH resistance together with reduced skeletal muscle mass are found in patients with increased protein catabolism due, for example, to sepsis, trauma, or major surgery. Both administration of glutamine-containing parenteral nutrition and GH treatment have been found to diminish this catabolism. The effects of GH are mediated in part by insulin-like growth factor I (IGF-I) that is produced in the liver and locally in GH target tissues. The aim of this study was to investigate the effect of GH treatment on expression of the IGF-I gene and GH receptor (GHR) gene in skeletal muscle after major surgery. A new quantitative RT-PCR-based assay was established to measure IGF-I gene expression. Metabolically healthy patients, without significant preoperative weight loss, who were undergoing elective abdominal surgery were included in the study. Five patients (one woman and four men) were treated with daily injections of GH (0.3 IU/kg.day) in addition to being given total parenteral nutrition including glutamine (0.28 g/kg.day). The control group consisted of eight patients (three women and five men), who were given glutamine-enriched total parenteral nutrition but no GH. A muscle biopsy was taken from the lateral portion of the quadriceps femoris muscle preoperatively (day 0) after induction of anesthesia. A second biopsy was taken under local anesthesia on postoperative day 3. Total ribonucleic acid (RNA) was extracted from the muscle biopsies, and IGF-I messenger RNA (mRNA) and GHR mRNA were measured by competitive quantitative RT-PCR assays. IGF-I mRNA and GHR mRNA levels were related to the expression of a housekeeping gene (cyclophilin). In the control group, IGF-I mRNA levels decreased from 1505 +/- 265 (mean +/- SEM) transcripts/cpm cyclophilin on day 0 to 828 +/- 172 on day 3 (P < 0.05). In contrast, IGF-I mRNA levels did not change in the GH-treated group (1188 +/- 400 transcripts/cpm cyclophilin on day 0 vs. 1089 +/- 342 transcripts/cpm cyclophilin on day 3). No statistically significant changes were seen in GHR expression. We conclude that administration of GH prevents the reduction in IGF-I gene expression in skeletal muscle after abdominal surgery.

No evidence for involvement of the growth hormone/insulin-like growth factor-1 axis in psoriasis.

We have examined whether alterations in the growth hormone/insulin-like growth factor-1 axis play a role in the pathogenesis of psoriasis. Serum, urine, full skin biopsies, and suction blister roofs were obtained from patients with psoriasis and from healthy controls. Serum concentrations of insulin-like growth factor-1 and insulin-like growth factor binding protein-3 were measured by radioimmunoassay. Growth hormone-binding protein was measured by ligand-mediated immunofunctional assay. Growth hormone concentration in urine was measured by an immunometric assay, and growth hormone receptor-gene expression was measured by RNase protection assay or by quantitative reverse transcriptase polymerase chain reaction in total RNA isolated from epidermal suction blister roofs. Serum concentrations of insulin-like growth factor-1 (249 +/- 12 micrograms per liter, mean +/- SEM, n = 42, and 277 +/- 21 micrograms per liter, n = 9, for psoriatic patients and controls, respectively), insulin-like growth factor binding protein-3 (3.1 +/- 0.08 mg per liter, n = 42, and 3.3 +/- 0.22 mg per liter, n = 9), growth hormone-binding protein (344 +/- 65 pmol per liter, n = 10, and 311 +/- 83 pmol per liter, n = 9), urinary growth hormone excretion during 24 h (12.8 +/- 2.7 microIU per 24 h, n = 12, and 12.3 +/- 1.6 microIU per 24 h, n = 9), and epidermal growth hormone receptor gene expression [32 +/- 12 x 10(3) mRNA transcripts per microgram total RNA (involved skin), n = 11, and 47 +/- 14 x 10(3) mRNA transcripts per microgram total RNA, n = 9] were similar in patients and controls. For insulin-like growth factor-1 and insulin-like growth factor binding protein-3 the values in psoriatic patients were also similar to those in larger control groups, n = 195 and n = 400, respectively. In addition, we found no evidence of local expression of growth hormone or prolactin in full skin punch biopsies from psoriatic involved skin by reverse transcriptase polymerase chain reaction. In conclusion, our results suggest that alterations in the growth hormone/ insulin-like growth factor-1 axis do not play a major role in the pathogenesis of psoriasis.

Measurement of human growth hormone receptor messenger ribonucleic acid by a quantitative polymerase chain reaction-based assay: demonstration of reduced expression after elective surgery.

Studies of GH receptor (GHR) gene expression in human tissues have been hampered by the limited amount of tissue available for analysis and the low sensitivity of conventional methods. We have developed a quantitative reverse transcriptase-PCR assay for measurement of GHR messenger ribonucleic acid levels in small human tissue biopsies. To compensate for sample to sample variation, an internal RNA standard, which differs from the wild-type GHR transcript by only a few nucleotides, was reverse transcribed and amplified together with the GHR transcripts. PCR was carried out using one biotinylated primer to permit the purification of single stranded PCR products on streptavidin-coated microtiter plates. The ratio between the wild-type and mutated transcripts was determined by two separate minisequence reactions in which a primer, annealed immediately 3' of a variable nucleotide, was extended by a single 3H-labeled nucleotide, complementary to either the wild-type or mutated sequence. The assay range was 0.125-8 x 10(5) transcripts/sample, the mean intraassay coefficient of variation was 8.7%, and the lower limit of detection was 0.125 x 10(5) transcripts/sample. GHR messenger ribonucleic acid levels were detectable in small amounts (10-100 ng) of total RNA extracted from adipose tissue, skeletal muscle, and liver. The GHR gene expression in liver was approximately 10-fold higher than that in skeletal muscle, whereas intermediate levels were found in adipose tissue. In nine patients undergoing elective abdominal surgery, GHR gene expression in skeletal muscle was reduced on day 3 after surgery compared to the baseline level. The decrease in GHR gene expression was accompanied by a decrease in skeletal muscle glutamine. This suggests that the postoperative protein catabolism may be caused at least partly by acquired GH insensitivity due to reduced expression of the GHR gene.

Expression of exon 3-retaining and exon 3-excluding isoforms of the human growth hormone-receptor is regulated in an interindividual, rather than a tissue-specific, manner.

GH has multiple effects on growth and metabolism, and these functions are mediated through binding to specific cell surface receptors. The human GH receptor (GHR) exists in two known isoforms; in one form exon 3 is present (GHR3+), and in the other, exon 3 is absent (GHR3-). Recent reports have suggested that the expression of the two isoforms is tissue specific and/or developmentally regulated. We used a reverse transcription-polymerase chain reaction assay to study the expression pattern of the two isoforms in a variety of tissues from normal subjects and patients with acromegaly. In skeletal muscle from both normal subjects and patients with acromegaly, the GHR3+ transcript was expressed, either alone or together with the shorter (GHR3-) transcript. When multiple tissues from six subjects were tested, the expression of the two isoforms varied among subjects, whereas different tissues from the same subject showed the same expression pattern. These results indicate that the expression of the GHR isoforms is not tissue specific. Instead, the expression of the GHR isoforms appears to be specific for each individual, suggesting that it is under the control of factors that affect all tissues in the body.