PPARalpha deficiency increases secretion and serum levels of apolipoprotein B-containing lipoproteins.

This study investigates the importance of peroxisome proliferator activated receptor alpha (PPARalpha) for serum apolipoprotein B (apoB) levels and hepatic secretion of apoB-containing lipoproteins. Total serum apoB and VLDL-apoB levels were higher in female PPARalpha-null mice compared with female wild-type mice, but no difference was seen in male mice. Furthermore, hepatic triglyceride secretion rate, determined in vivo after Triton WR1339 injection, was 2.4-fold higher in female PPARalpha-null mice compared with female wild-type mice, but no difference was observed in male mice. However, when fed a high fat diet, male PPARalpha-null mice displayed 2-fold higher serum levels of apoB and LDL cholesterol compared with male wild-type mice, but triglyceride levels were not affected. Hepatic LDL receptor protein levels were not influenced by PPARalpha deficiency, gender, or the fat diet. Hepatocyte cultures from female PPARalpha-null mice (cultured for 4 days in serum free medium) showed 2-fold higher total apoB secretion and increased secretion of apoB-48 VLDL, as well as 2.7-fold larger accumulation of VLDL-triglycerides in the medium compared with wild-type cultures. In conclusion, PPARalpha-deficient female mice, but not males, display high serum apoB associated with VLDL and increased hepatic triglyceride secretion. Moreover, male PPARalpha-null mice show increased susceptibility to high fat diet in terms of serum apoB levels.

Increased resistin expression in the adipose tissue of male prolactin transgenic mice and in male mice with elevated androgen levels.

The aim of this study was to investigate the regulation of resistin, a recently identified adipocyte-secreted peptide, in the adipose tissue of prolactin (PRL)-transgenic (tg) mice using ribonuclease protection assay. The level of resistin mRNA increased 3.5-fold in the adipose tissue of untreated male PRL-tg mice compared to controls. However, there was no difference in resistin expression in the adipose tissue of female PRL-tg mice compared to control mice. PRL-tg male mice have elevated serum testosterone levels and we therefore analyzed the effects of testosterone alone on resistin mRNA expression. Furthermore, the effects of elevated androgen levels on PRL receptor (PRLR) mRNA expression in the adipose tissue were investigated. Resistin mRNA increased 2.6-fold in the adipose tissue of control male mice with elevated serum androgen levels. In addition, PRLR mRNA expression was increased in the adipose tissue of male mice with elevated testosterone. These results suggest testosterone to be a regulator of resistin and PRLR mRNA expression in the adipose tissue of male mice.

Liver-derived IGF-I regulates GH secretion at the pituitary level in mice.

We have reported that liver-specific deletion of IGF-I in mice (LI-IGF-I-/-) results in decreased circulating IGF-I and increased GH levels. In the present study, we determined how elimination of hepatic IGF-I modifies the hypothalamic-pituitary GH axis to enhance GH secretion. The pituitary mRNA levels of GH releasing factor (GHRF) receptor and GH secretagogue (GHS) receptor were increased in LI-IGF-I-/- mice, and in line with this, their GH response to ip injections of GHRF and GHS was increased. Expression of mRNA for pituitary somatostatin receptors, hypothalamic GHRF, somatostatin, and neuropeptide Y was not altered in LI-IGF-I-/- mice, whereas hypothalamic IGF-I expression was increased. Changes in hepatic expression of major urinary protein and the PRL receptor in male LI-IGF-I-/- mice indicated an altered GH release pattern most consistent with enhanced GH trough levels. Liver weight was enhanced in LI-IGF-I-/- mice of both genders. In conclusion, loss of liver-derived IGF-I enhances GH release by increasing expression of pituitary GHRF and GHS receptors. The enhanced GH release in turn affects several liver parameters, in line with the existence of a pituitary-liver axis.

Growth hormone receptor is expressed in human breast cancer.

Several clinical observations and experimental studies indicate that pituitary hormones, including growth hormone, play a role in the development of human breast cancer. We analyzed 48 human breast carcinomas using reverse transcription polymerase chain reaction, immunohistochemistry, and Western blotting techniques to assess growth hormone receptor expression. In 17 of these cases, adjacent normal breast tissue was similarly analyzed. These analyses revealed that growth hormone receptor (GHR) is expressed in human breast cancer and appears to be up-regulated compared to adjacent normal breast tissue. GHR expression correlated inversely with tumor grade and MIB-1 index. Progesterone receptor expression correlated positively with GHR expression. These findings, along with our observation of GHR expression in breast cancer stromal cells and previous reports of local production of growth hormone in breast carcinoma, suggest that GHR-mediated signaling pathways are involved in the development of human breast cancer, possibly via autocrine or paracrine mechanisms.

Investigation of the role of prolactin in the development and function of the lacrimal and harderian glands using genetically modified mice.

PURPOSE: To determine whether prolactin receptor is essential for normal development and function of the lacrimal gland and whether hyperprolactinemia can alter lacrimal development. METHODS: Lacrimal gland morphology and function were examined in two genetic mouse models of prolactin action: a prolactin receptor knockout model that is devoid of prolactin action and a transgenic model of hyperprolactinemia. RESULTS: Image analysis of lacrimal and Harderian gland sections was used to quantify glandular morphology. In females, lacrimal acinar area decreased by 30% and acinar cell density increased by 25% over control subjects in prolactin transgenic animals, but prolactin receptor knockout mice showed no changes. In males, transgenic animals showed no changes, but prolactin receptor knockout mice showed a 5% reduction in acinar area and an 11% increase in acinar cell density, which was lost after castration. The morphology of the Harderian glands underwent parallel changes but to a lesser degree. A complete loss of porphyrin accretions was seen in the Harderian glands of male and female knockout animals. No differences in tear protein levels were seen in knockout animals by two-dimensional gels. Enzyme-linked immunosorbent assay (ELISA) and Western blot analysis showed that the level of secretory component and IgA in knockout mouse tears remained unchanged. There was no change in the predisposition of the 129 mouse strain to conjunctivitis in the knockout animals. CONCLUSIONS: Prolactin plays a small role in establishing the sexual dimorphism of male lacrimal glands. In females, hyperprolactinemia causes a hyperfemale morphology, suggesting a role in dry eye syndromes. Prolactin is required for porphyrin secretion by the Harderian gland but plays no essential role in the secretory immune function of the lacrimal gland.

Prolactin (PRL) receptor gene expression in mouse adipose tissue: increases during lactation and in PRL-transgenic mice.

There are indications that PRL may exert important metabolic actions on adipose tissue in different species. However, with the exception of birds, the receptor has not been identified in white adipose tissue. The present study was designed to examine the possible expression and regulation of the PRL receptor (PRLR) in mouse adipose tissue. The long PRLR messenger RNA (mRNA) splice form (L-PRLR) and two short splice forms (S2- and S3-PRLR) were detected in mouse adipose tissue by RT-PCR. Furthermore, L-PRLR mRNA was detected by ribonuclease protection assay. Immunoreactive PRLR with a relative molecular mass of 95,000 was revealed by immunoblotting. Furthermore, L-PRLR mRNA expression was demonstrated in primary isolated adipocytes. In mouse adipose tissue, the level of L-PRLR mRNA expression increased 2.3-fold during lactation compared with those in virgin and pregnant mice. In contrast, in the liver the expression of L-PRLR increased 3.4-fold during pregnancy compared with those in virgin and lactating mice. When comparing the levels of L-PRLR expression in virgin female and male mice, no difference was detected in adipose tissue. However, in virgin female liver the expression was 4.5-fold higher than that in male liver. As PRL up-regulates its own receptor in some tissues, we analyzed L-PRLR expression in PRL-transgenic female and male mice. In PRL-transgenic mice L-PRLR expression was significantly increased in both adipose tissue (1.4-fold in females and 2.4-fold in males) and liver (1.9-fold in females and 2.7-fold in males) compared with that in control mice. Furthermore, in female PRL-transgenic mice retroperitoneal adipose tissue was decreased in weight compared with that in control mice. However, no difference was detected when comparing the masses of parametrial adipose tissue. Our results suggest a direct role for PRL, mediated by PRLR, in modulating physiological events in adipose tissue.

The role of prolactin and growth hormone in breast cancer.

This review will focus on the role for prolactin (PRL) and growth hormone (GH) in mammary tumor formation. Much attention has previously been focused on circulating levels of GH/PRL in relation to mammary tumor formation. We will review data demonstrating that these ligands also could be produced locally in different organs, including the mammary gland and mammary tumors, and suggest that this local production may be of importance for pathological conditions. We will also discuss mechanisms for crosstalk between steroids and GH/PRL. A crosstalk between GH- and PRL response is possible at multiple levels. In the human, GH can activate both the prolactin receptor (PRLR) and the growth hormone receptor (GHR). We have demonstrated that activation of the PRLR, but not the GHR, is inducing mammary tumors in transgenic mice. Furthermore, the elevated levels of insulin-like growth factor 1 (IGF-I) seen in the GHR activating transgenic mice is not sufficient for tumor induction. The induced tumors express functionally active prolactin that could be of importance for the tumor formation. Paracrine/aurocrine stimulation by PRL may be more important than PRL transported via the circulation. In women, the role for stimulation of the PRLR and/or the GHR in mammary tumor formation has not been proven, although experiments from primates suggest that the PRLR could be of importance.

Transgenic mice overexpressing the prolactin gene develop dramatic enlargement of the prostate gland.

An altered endocrine status of elderly men has been hypothesized to be important for development of prostate hyperplasia. The present study addresses the question whether increased PRL expression is of importance for development of prostate hyperplasia in mice. Three lines of PRL transgenic mice were generated having serum levels of PRL of approximately 15 ng/ml, 100 ng/ml, and 250 ng/ml, respectively. These mice developed dramatic enlargement of the prostate gland, approximately 20 times the normal prostate weight and they had a 4- to 5-fold increased DNA content. Histologically, the prostate glands in the transgenic mice were distended from secretion, and the amount of interstitial tissue was increased. The levels oftestosterone and IGF-I were increased in the PRL transgenic animals. In mice overexpressing the bovine GH gene, displaying elevated IGF-I levels, the prostate gland was slightly larger compared with normal mice, indicating that the effect of PRL was not primarily mediated through elevated plasma IGF-I levels. The present study suggests that PRL is an important factor in the development of prostate hyperplasia acting directly on the prostate gland or via increased plasma levels of testosterone.

Activation of the prolactin receptor but not the growth hormone receptor is important for induction of mammary tumors in transgenic mice.

Transgenic mice overexpressing the human growth hormone gene develop mammary carcinomas. Since human growth hormone gene can activate both the growth hormone receptor (GHR) and the prolactin (PRL) receptor (PRLR), it is not clear which receptor system is responsible for the malignant transformation. To clarify the receptor specificity, we created transgenic mice with two different genes: (a) transgenic mice overexpressing the bovine growth hormone (bGH) gene having high levels of bGH only activating the GHR and also high serum levels of IGF-I; and (b) transgenic mice overexpressing the rat PRL (rPRL) gene that have elevated levels of PRL (one line 150 ng/ml and one line 13 ng/ml) only binding to the PRLR and with normal IGF-I levels. When analyzed histologically, all of the PRL transgenic female mice developed mammary carcinomas at 11-15 mo of age. Only normal mammary tissue was observed among the bGH transgenic animals and the controls. Cell lines established from a tumor produced rPRL and expressed PRLR. In organ culture experiments, an auto/paracrine effect of rPRL was demonstrated. In conclusion, activation of the PRLR is sufficient for induction of mammary carcinomas in mice, while activation of the GHR is not sufficient for mammary tumor formation.

High frequency of mammary adenocarcinomas in metallothionein promoter-human growth hormone transgenic mice created from two different strains of mice.

Transgenic mice were developed by injecting a mouse metallothionein promoter-human growth hormone (Mt-hGH) gene fragment into the pronucleus of C57Bl x DBA/2J-f2 or C57Bl x CBA-f2 one cell embryos. Six founder animals with the C57Bl x DBA genetic background grew 1.3-2.2 times larger than littermate controls and had higher levels of hGH in plasma (4.6-279 mU/l). Three of the four female transgenic founders developed malignant papillar adenocarcinomas of mammary origin at 27-43 weeks of age. One male transgenic founder was successfully mated and two of three female transgenic offsprings developed mammary tumors. To examine if the tumor induction was dependent on the strain of mice used the experiments were repeated using animals with different genetic background. Fourteen female hGH transgenic mice from five founder animals were generated using C57Bl x CBA-f2 mice. Thirteen of the animals had elevated levels of hGH in plasma (7-1960 mU/l) and grew larger than control animals. Nine of the animals developed mammary adenocarcinomas. Four of the hGH expressing animals did not demonstrate macroscopic tumor formation but have not yet been analyzed histologically. The present study suggests that markedly elevated endogenous levels of GH cause mammary carcinoma in hGH transgenic mice. The present animal model might prove useful for studying molecular mechanisms involved in the development of hormonally induced mammary tumors.