Isoform-specific knockdown of long and intermediate prolactin receptors interferes with evolution of B-cell neoplasms.

Prolactin (PRL) is elevated in B-cell-mediated lymphoproliferative diseases and promotes B-cell survival. Whether PRL or PRL receptors drive the evolution of B-cell malignancies is unknown. We measure changes in B cells after knocking down the pro-proliferative, anti-apoptotic long isoform of the PRL receptor (LFPRLR) in vivo in systemic lupus erythematosus (SLE)- and B-cell lymphoma-prone mouse models, and the long plus intermediate isoforms (LF/IFPRLR) in human B-cell malignancies.

A cautionary tale: Alien prolactins may induce lesser, no, or opposite effects to homologous hormone!

Cost and availability have often dictated the use of heterologous/alien prolactins in experiments, particularly in vivo. The assumption has been that what is initiated in the target cell is representative of the homologous hormone since many heterologous mammalian prolactins bind to and activate rodent receptors. Here, we examined gene expression in mouse liver in response to a 7-day treatment with recombinant mouse prolactin (mRecPRL), recombinant ovine prolactin (oRecPRL) and pituitary extract ovine prolactin (oPitPRL).

Tumor cell-derived asymmetric dimethylarginine regulates macrophage functions and polarization.

Background: Asymmetric dimethylarginine (ADMA), which is significantly elevated in the plasma of cancer patients, is formed via intracellular recycling of methylated proteins and serves as a precursor for resynthesis of arginine. However, the cause of ADMA elevation in cancers and its impact on the regulation of tumor immunity is not known.

Methods: Three mouse breast cell lines (normal breast epithelial HC11, breast cancer EMT6 and triple negative breast cancer 4T1) and their equivalent 3D stem cell culture were used to analyze the secretion of ADMA using ELISA and their responses to ADMA.

Bittersweet: relevant amounts of the common sweet food additive, glycerol, accelerate the growth of PC3 human prostate cancer xenografts.

Objective: In a study of potential prostate cancer therapeutics, glycerol was used to increase the density of one solution. Glycerol alone was therefore one of the controls. Tumors of human PC3 castrate-resistant prostate cancer cells were initiated in male nude mice and grown for 12 days.

Microdissection and Dissociation of the Murine Oviduct: Individual Segment Identification and Single Cell Isolation.

Mouse model systems are unmatched for the analysis of disease processes because of their genetic manipulability and the low cost of experimental treatments. However, because of their small body size, some structures, such as the oviduct with a diameter of 200-400 μm, have proven to be relatively difficult to study except by immunohistochemistry. Recently, immunohistochemical studies have uncovered more complex differences in oviduct segments than were previously recognized; thus, the oviduct is divided into four functional segments with different ratios of seven distinct epithelial cell types.

Prolactin enhances T regulatory cell promotion of breast cancer through the long form prolactin receptor.

Previous work has shown systemic knockdown of the long form prolactin receptor (LFPRLR) in vivo markedly reduced metastasis in mouse models of breast cancer, but whether this translated to prolonged survival was unknown. Here we show that LFPRLR knockdown in the highly metastatic, immunocompetent 4T1 model prolonged survival and reduced recruitment of T regulatory cells (Tregs) to the tumor through effects on the production of CCL17. For the Tregs still recruited to the primary tumor, LFPRLR knockdown both directly and indirectly reduced their ability to promote tumor parenchymal epithelial to mesenchymal transition.

Sex Differences in the Immune System Become Evident in the Perinatal Period in the Four Core Genotypes Mouse.

We have used the four core genotypes (FCG) mouse model, which allows a distinction between effects of gonadal secretions and chromosomal complement, to determine when sex differences in the immune system first appear and what influences their development. Using splenic T cell number as a measure that could be applied to neonates with as yet immature immune responses, we found no differences among the four genotypes at postnatal day 1, but by day 7, clear sex differences were observed. These sex differences were unexpectedly independent of chromosomal complement and similar in degree to gonadectomized FCG adults: both neonatal and gonadectomized adult females (XX and XY) showed 2-fold the number of CD4+ and 7-fold the number of CD8+ T cells their male (XX and XY) counterparts.

Prolactin Attenuates Neuroinflammation in LPS-Activated SIM-A9 Microglial Cells by Inhibiting NF-κB Pathways Via ERK1/2.

Prolactin (PRL) is a pleiotropic hormone with multiple functions in several tissues and organs, including the brain. PRL decreases lesion-induced microgliosis and modifies gene expression related to microglial functions in the hippocampus, thereby providing a possible mechanism through which it might participate in neuroimmune modulatory responses and prevent neuronal cell damage. However, the direct contribution of microglial cells to PRL-mediated neuroprotection is still unclear and no studies have yet documented whether PRL can directly activate cellular pathways in microglial cells.

S179D Prolactin Sensitizes Human PC3 Prostate Cancer Xenografts to Anti-tumor Effects of Well-Tolerated Doses of Calcitriol.

Calcitriol has been shown to have multiple anti-prostate cancer effects both and in xenograft models, and associations between low levels of calcitriol and more aggressive forms of prostate cancer have been observed clinically. However, the concentrations of calcitriol required to have a substantive anti-cancer effect are toxic. In previous work, we had observed that the selective prolactin receptor modulator, S179D PRL, sensitized prostate cancer cells to physiological concentrations of calcitriol through an ability to increase expression of the vitamin D receptor.

Sex Differences in Mouse Popliteal Lymph Nodes.

Females have more robust immune responses than males, well-illustrated by the degree of inflammation elicited during delayed-type hypersensitivity (DTH) responses. Here, we have investigated underlying sex differences that may contribute to differential footpad DTH responses using wildtype and four core genotypes (FCG) mice and popliteal lymphnode cellularity and gene expression. DTH responses in XX and XY FCG females showed no role for almost all genes expressed on sex chromosomes.

Epitope-specific affinity maturation improved stability of potent protease inhibitory antibodies.

Targeting effectual epitopes is essential for therapeutic antibodies to accomplish their desired biological functions. This study developed a competitive dual color fluorescence-activated cell sorting (FACS) to maturate a matrix metalloprotease 14 (MMP-14) inhibitory antibody. Epitope-specific screening was achieved by selection on MMP-14 during competition with N-terminal domain of tissue inhibitor of metalloproteinase-2 (TIMP-2) (nTIMP-2), a native inhibitor of MMP-14 binding strongly to its catalytic cleft.

Use of a novel camelid-inspired human antibody demonstrates the importance of MMP-14 to cancer stem cell function in the metastatic process.

Matrix metalloproteinases (MMPs) are considered excellent targets for cancer therapy because of their important roles in multiple aspects of tumor growth and metastatic spread. However, not all MMPs, or even all activities of specific MMPs, promote cancer. Therefore, there is a need for highly specific inhibitors.

Lactation-Based Maternal Educational Immunity Crosses MHC Class I Barriers and Can Impart Th1 Immunity to Th2-Biased Recipients.

We have previously demonstrated lactational transfer of T cell-based immunity from dam to foster pup. In the short term, a significant part of transferred immunity is passive cellular immunity. However, as time progresses, this is replaced by what we have described as maternal educational immunity such that by young adulthood, all immune cells responding to a foster dam immunogen are the product of the foster pup's thymus.

Involvement of miR-106b in tumorigenic actions of both prolactin and estradiol.

Prolactin promotes a variety of cancers by an array of different mechanisms. Here, we have investigated prolactin's inhibitory effect on expression of the cell cycle-regulating protein, p21. Using a miRNA array, we identified a number of miRNAs upregulated by prolactin treatment, but one in particular that was strongly induced by prolactin and predicted to bind to the 3'UTR of p21 mRNA, miR-106b.

Role of Prolactin in Promotion of Immune Cell Migration into the Mammary Gland.

Immune cells in the mammary gland play a number of important roles, including protection against infection during lactation and, after passing into milk, modulation of offspring immunity. However, little is known about the mechanism of recruitment of immune cells to the lactating gland in the absence of infection. Given the importance of prolactin to other aspects of lactation, we hypothesized it would also play a role in immune cell recruitment.

Maternal Milk T Cells Drive Development of Transgenerational Th1 Immunity in Offspring Thymus.

Using multiple murine foster-nursing protocols, thereby eliminating placental transfer and allowing a distinction between dam- and pup-derived cells, we show that foster nursing by an immunized dam results in development of CD8(+) T cells in nonimmunized foster pups that are specific for Ags against which the foster dam was immunized (Mycobacterium tuberculosis or Candida albicans). We have dubbed this process "maternal educational immunity" to distinguish it from passive cellular immunity. Of the variety of maternal immune cells present in milk, only T cells were detected in pup tissues.

Prolactin inhibits a major tumor-suppressive function of wild type BRCA1.

Even though mutations in the tumor suppressor, BRCA1, markedly increase the risk of breast and ovarian cancer, most breast and ovarian cancers express wild type BRCA1. An important question is therefore how the tumor-suppressive function of normal BRCA1 is overcome during development of most cancers. Because prolactin promotes these and other cancers, we investigated the hypothesis that prolactin interferes with the ability of BRCA1 to inhibit the cell cycle.

Anti-metastatic outcome of isoform-specific prolactin receptor targeting in breast cancer.

Controversy exists concerning the role of the long prolactin receptor (PRLR) in the progression of breast cancer. By targeting pre-mRNA splicing, we succeeded in knocking down only the long PRLR in vivo, leaving the short forms unaffected. Using two orthotopic and highly-metastatic models of breast cancer, one of which was syngeneic (mouse 4T1) to allow assessment of tumor-immune interactions and one of which was endocrinologically humanized (human BT-474) to activate human PRLRs, we examined the effect of long PRLR knockdown on disease progression.

Antitumor activity of a polypyridyl chelating ligand: in vitro and in vivo inhibition of glioma.

Glioblastoma multiforme is an extremely aggressive and invasive form of central nervous system tumor commonly treated with the chemotherapeutic drug Temozolomide. Unfortunately, even with treatment, the median survival time is less than 12 months. 2,9-Di-sec-butyl-1,10-phenanthroline (SBP), a phenanthroline-based ligand originally developed to deliver gold-based anticancer drugs, has recently been shown to have significant antitumor activity in its own right.

Blockade of estrogen-stimulated proliferation by a constitutively-active prolactin receptor having lower expression in invasive ductal carcinoma.

A comprehensive understanding of prolactin's (PRL's) role in breast cancer is complicated by disparate roles for alternatively-spliced PRL receptors (PRLR) and crosstalk between PRL and estrogen signaling. Among PRLRs, the short form 1b (SF1b) inhibits PRL-stimulated cell proliferation. In addition to ligand-dependent PRLRs, constitutively-active varieties, missing the S2 region of the extracellular domain (ΔS2), naturally occur.

Expression of a constitutively active prolactin receptor causes histone trimethylation of the p53 gene in breast cancer.

Background: Prolactin (PRL) is a pituitary polypeptide hormone characterized by multiple biological actions including stimulation of growth in the prostate and formation of secretory alveoli and stimulation of milk protein gene expression in the mammary gland. PRL exerts its effect by dimerizing its receptor (PRLR) on the plasma membrane and regulating gene expression through the JAK-Stat signal pathway. We have previously described a natural variant of the PRLR in which the S2 subdomain of the extracellular domain is missing (Delta S2).

An N-terminal splice variant of human Stat5a that interacts with different transcription factors is the dominant form expressed in invasive ductal carcinoma.

We have identified a new variant of human Stat5a, found at higher ratios to full-length Stat5a in invasive ductal carcinoma versus contiguous normal tissue. The variant, missing exon 5, inhibits p21 and Bax production and increases cell number. After prolactin stimulation, only full-length Stat5a interacts with the vitamin D and retinoid X receptors, whereas only Δ5 Stat5a interacts with activating protein 1-2 and specificity protein 1.