The TGR5 receptor mediates bile acid–induced itch and analgesia

Patients with cholestatic disease exhibit pruritus and analgesia, but the mechanisms underlying these symptoms are unknown. We report that bile acids, which are elevated in the circulation and tissues during cholestasis, cause itch and analgesia by activating the GPCR TGR5. TGR5 was detected in peptidergic neurons of mouse dorsal root ganglia and spinal cord that transmit itch and pain, and in dermal macrophages that contain opioids. Bile acids and a TGR5-selective agonist induced hyperexcitability of dorsal root ganglia neurons and stimulated the release of the itch and analgesia transmitters gastrin-releasing peptide and leucine-enkephalin. Intradermal injection of bile acids and a TGR5-selective agonist stimulated scratching behavior by gastrin-releasing peptide– and opioid-dependent mechanisms in mice. Scratching was attenuated in Tgr5-KO mice but exacerbated in Tgr5-Tg mice (overexpressing mouse TGR5), which exhibited spontaneous pruritus. Intraplantar and intrathecal injection of bile acids caused analgesia to mechanical stimulation of the paw by an opioid-dependent mechanism. Both peripheral and central mechanisms of analgesia were absent from Tgr5-KO mice. Thus, bile acids activate TGR5 on sensory nerves, stimulating the release of neuropeptides in the spinal cord that transmit itch and analgesia. These mechanisms could contribute to pruritus and painless jaundice that occur during cholestatic liver diseases.     

Hyperactivation of phosphatidylinositol-3 kinase promotes escape from hormone dependence in estrogen receptor–positive human breast cancer

Many breast cancers exhibit a degree of dependence on estrogen for tumor growth. Although several therapies have been developed to treat individuals with estrogen-dependent breast cancers, some tumors show de novo or acquired resistance, rendering them particularly elusive to current therapeutic strategies. Understanding the mechanisms by which these cancers develop resistance would enable the development of new and effective therapeutics. In order to determine mechanisms of escape from hormone dependence in estrogen receptor–positive (ER-positive) breast cancer, we established 4 human breast cancer cell lines after long-term estrogen deprivation (LTED). LTED cells showed variable changes in ER levels and sensitivity to 17β-estradiol. Proteomic profiling of LTED cells revealed increased phosphorylation of the mammalian target of rapamycin (mTOR) substrates p70S6 kinase and p85S6 kinase as well as the PI3K substrate AKT. Inhibition of PI3K and mTOR induced LTED cell apoptosis and prevented the emergence of hormone-independent cells. Using reverse-phase protein microarrays, we identified a breast tumor protein signature of PI3K pathway activation that predicted poor outcome after adjuvant endocrine therapy in patients. Our data suggest that upon adaptation to hormone deprivation, breast cancer cells rely heavily on PI3K signaling. Our findings also imply that acquired resistance to endocrine therapy in breast cancer may be abrogated by combination therapies targeting both ER and PI3K pathways.     

Tamoxifen regulation of bone growth and endocrine function in the ovariectomized rat: discrimination of responses involving estrogen receptor α/estrogen receptor β, G protein-coupled estrogen receptor, or estrogen-related receptor γ using fulvestrant (ICI 182780)

Tamoxifen is a selective estrogen receptor (ER) modulator, but it is also a deactivating ligand for estrogen-related receptor-γ (ERRγ) and a full agonist for the G protein-coupled estrogen receptor (GPER). Fulvestrant is a selective ER down-regulator that lacks agonist effects on ERα/ERβ, is inactive on ERRγ, but acts as a full agonist on GPER. Fulvestrant effects on tamoxifen actions on uterine and somatic growth, bone, the growth hormone (GH)-insulin-like growth factor I (IGF-I) axis, and pituitary prolactin were analyzed to pharmacologically discriminate tamoxifen effects that may be mediated by ERα/ERβ versus ERRγ versus GPER. Ovariectomized rats received tamoxifen (0.6 mg/kg/daily) plus fulvestrant at 0, 3, 6, or 12 mg/kg/daily for 5 weeks; controls received vehicle or 6 mg/kg fulvestrant daily. Tamoxifen effects to increase uterine weight, decrease serum IGF-I, increase pituitary prolactin, and increase bone mineral density could be fully blocked by fulvestrant, indicating mediation by ERα/ERβ. Tamoxifen effects to decrease pituitary GH, tibia length, and body weight were only partially blocked by fulvestrant, indicating involvement of mechanisms unrelated to ERα/ERβ. Fulvestrant did not inhibit tamoxifen actions to reduce total pituitary protein, again indicating effects not mediated by ERα/ERβ. Tamoxifen actions to reduce serum GH were mimicked rather than inhibited by fulvestrant, pharmacological features consistent with GPER involvement. However, fulvestrant alone increased IGF-I and also blocked tamoxifen-evoked IGF-I decreases; thus fulvestrant effects on serum GH might reflect increased IGF-I feedback inhibition. Fulvestrant alone had no effect on the other parameters. The findings indicate that mechanisms unrelated to ERα/ERβ contribute to tamoxifen effects on body weight, bone growth, and pituitary function.     

Growth hormone receptor regulates β cell hyperplasia and glucose-stimulated insulin secretion in obese mice

Insulin, growth hormone (GH), and insulin-like growth factor-1 (IGF-1) play key roles in the regulation of β cell growth and function. Although β cells express the GH receptor, the direct effects of GH on β cells remain largely unknown. Here we have employed a rat insulin II promoter-driven (RIP-driven) Cre recombinase to disrupt the GH receptor in β cells (βGHRKO). βGHRKO mice fed a standard chow diet exhibited impaired glucose-stimulated insulin secretion but had no changes in β cell mass. When challenged with a high-fat diet, βGHRKO mice showed evidence of a β cell secretory defect, with further deterioration of glucose homeostasis indicated by their altered glucose tolerance and blunted glucose-stimulated insulin secretion. Interestingly, βGHRKO mice were impaired in β cell hyperplasia in response to a high-fat diet, with decreased β cell proliferation and overall reduced β cell mass. Therefore, GH receptor plays critical roles in glucose-stimulated insulin secretion and β cell compensation in response to a high-fat diet.     

Neurotrophin receptor p75NTR mediates Huntington’s disease–associated synaptic and memory dysfunction

Learning and memory deficits are early clinical manifestations of Huntington’s disease (HD). These cognitive impairments have been mainly associated with frontostriatal HD pathology; however, compelling evidence provided by several HD murine models suggests that the hippocampus may contribute to synaptic deficits and memory dysfunction in HD. The neurotrophin receptor p75^NTR negatively regulates spine density, which is associated with learning and memory; therefore, we explored whether disturbed p75^NTR function in the hippocampus could contribute to synaptic dysfunction and memory deficits in HD. Here, we determined that levels of p75^NTR are markedly increased in the hippocampus of 2 distinct mouse models of HD and in HD patients. Normalization of p75^NTR levels in HD mutant mice heterozygous for p75^NTR prevented memory and synaptic plasticity deficits and ameliorated dendritic spine abnormalities, likely through normalization of the activity of the GTPase RhoA. Moreover, viral-mediated overexpression of p75^NTR in the hippocampus of WT mice reproduced HD learning and memory deficits, while knockdown of p75^NTR in the hippocampus of HD mice prevented cognitive decline. Together, these findings provide evidence of hippocampus-associated memory deficits in HD and demonstrate that p75^NTR mediates synaptic, learning, and memory dysfunction in HD.     

Divergent roles for thyroid hormone receptor β isoforms in the endocrine axis and auditory system

Thyroid hormone receptors (TRs) modulate various physiological functions in many organ systems. The TR alpha and TR beta isoforms are products of 2 distinct genes, and the beta 1 and beta 2 isoforms are splice variants of the same gene. Whereas TR alpha 1 and TR beta 1 are widely expressed, expression of the TR beta 2 isoform is mainly limited to the pituitary, triiodothyronine-responsive TRH neurons, the developing inner ear, and the retina. Mice with targeted disruption of the entire TR beta locus (TR beta-null) exhibit elevated thyroid hormone levels as a result of abnormal central regulation of thyrotropin, and also develop profound hearing loss. To clarify the contribution of the TR beta 2 isoform to the function of the endocrine and auditory systems in vivo, we have generated mice with targeted disruption of the TR beta 2 isoform. TR beta 2-null mice have preserved expression of the TR alpha and TR beta 1 isoforms. They develop a similar degree of central resistance to thyroid hormone as TR beta-null mice, indicating the important role of TR beta 2 in the regulation of the hypothalamic-pituitary-thyroid axis. Growth hormone gene expression is marginally reduced. In contrast, TR beta 2-null mice exhibit no evidence of hearing impairment, indicating that TR beta 1 and TR beta 2 subserve divergent roles in the regulation of auditory function.     

Relation of estrogen receptor-α gene polymorphism and hormone replacement therapy to fall risk and muscle strength in early postmenopausal women

BACKGROUND. Several factors may increase fracture risk, among them reduced bone mineral density (BMD), increased bone resorption, microarchitectural deterioration of bone, increased fall risk, and decreased muscle strength. We have previously reported that PvuII polymorphism of the estrogen receptor-α (ER α) gene is associated with bone loss rate, fracture risk, and response to hormone replacement therapy (HRT) in early postmenopausal Finnish women. METHOD. We studied the influence of the ERα genotype on fall risk and muscle strength in a 5-year randomized HRT trial of 331 early postmenopausal women (subgroup of the population-based OSTPRE study, Kuopio, Finland). A 5-year postal inquiry in May 1994 included questions on falls during the previous 12 months. Grip strength was measured with dynamometer. The ERα gene polymorphism was analysed using PCR and PvuII restriction enzyme digestion. RESULTS. In all, 97 out of the 331 women reported falls. Half of those (56%) were slip falls, mostly during the winter season. In the HRT group, the ERα genotype was associated with fall risk (P = 0.002, logistic regression). The risk of falls (RR) was higher in women with the PP genotype than in those with the Pp (RR = 5.26, 95% CI 1.98-13.94, P = 0.001) or the pp (RR = 3.84, 95% CI 1.46-10.12, P = 0.007) genotype. When the falls were divided into slip (environment-related) and non-slip (endogenous) falls, the non-slip falls were associated with the genotype (P = 0.004), but the slip falls were not so clearly (P = 0.061). When all falls and non-slip falls were adjusted to the number of chronic health disorders and the variable time-since-menopause, the difference between the genotypes persisted (P = 0.003 and P = 0.010, respectively). In the non-HRT group, the ERα genotype was not associated with fall risk. The baseline or the 5-year grip strength values were not influenced by the ERα genotype. In conclusion, ER α polymorphism is associated with fall risk, especially with non-slip falls, in early postmenopausal Finnish women during the HRT. We have previously reported that, during HRT, women with the P allele have decreased fracture risk and that they may preferentially derive benefit from the positive effect of HRT on BMD. This suggests that the influence of ERα polymorphism may depend on the target tissue (bone versus the nervous system). CONCLUSIONS. In these early postmenopausal, non-osteoporotic and relatively healthy women, the increased fall risk associated with the PP genotype was not associated with increased fracture risk, possibly due to improved bone strength during the HRT although falls generally predispose to fractures.     

Correlation of the expression of gonadotropin releasing hormone and its receptor in human gastric adenocarcinoma cell proliferation and differentiation

AIM:To inestigate the possible correlation of the expression of gonadotropin releasing hormone(GnRH) and its receptor with of proliferating cell nuclear antigen(PCNA) in human gastric adenocarcinoma cell proliferation and differentiation.METHODS:GnRH and its receptor and PCNA were detected in 30 gastric adenocarcinoma by immunohistochemical ABC technique.RESULTS:GnRH and its receptor had the same distribution pattern in gastric adenocarcinoma cells.The positive signal was found mainly in cytoplssm.The content of GnRH and its receptor immunoreative product in high differentiatied adenocarecinoma was the lowest(P&;lt;0.05).PCNA positive signal which was found mainly in nucleus was gradually abated along with the rainsing of the extent of the differentiation.THe differences were significant among the three groups(P&;lt;0.05).In human gastric adenocarcinoma.the expression of GnRH and its receptor was negative correlation with the expression of PCNA(r=0.9).CONCLUSION:GnRH might be involved in the regulation of human gastric adenocarcinoma cell proliferation and direrentiation.     

Neutralization of IL-10 restores the downregulation of IL-18 receptor on natural killer cells and interferon-γ production in septic mice, thus leading to an improved survival

The objective of the study was to investigate the mechanisms of insufficient interferon-γ (IFN-γ) response to interleukin 18 (IL-18) and the treatment for the insufficient response in septic mice. Interleukin 18 stimulation does not restore IFN-γ production by blood mononuclear cells in septic patients but does restore its production in postoperative patients. Although sepsis impairs the IFN-γ response to IL-18, little is known about why the IL-18/IFN-γ-mediated immune response is ineffective in patients with sepsis. A cecal ligation and puncture was made in C57BL/6 mice following a sublethal lipopolysaccharide challenge to examine their IFN-γ response to IL-18, focusing on natural killer (NK) cells and cytokines. We next examined the effect of neutralization of IL-10 on the NK cell and survival in septic mice. Interleukin 18 injection did not restore IFN-γ production in septic (cecal ligation and puncture) mice. Despite an increase in the numbers of liver NK cells, the IL-18 receptor (IL-18R) expression was decreased in the septic mice compared with sham mice. Serum IL-10 levels were positively correlated with the percentage of liver NK cells, but negatively with their IL-18R expression. Neutralization of IL-10 restored the IL-18R expression on liver NK cells and restored the IFN-γ response in the septic mice, improving their survival. Sepsis might impair IL-18R expression on liver and spleen NK cells and impair the IL-18-mediated IFN-γ response. Neutralization of IL-10 may restore this response in septic hosts, thereby improving survival.     

Rapid activation of nuclear factor κB by 17β-estradiol and selective estrogen receptor modulators: pathways mediating cellular protection

17β-Estradiol (E2) treatment activates a set of protective response that has been found to protect cells from injury and more importantly to significantly abate the injuries associated with trauma-hemorrhage in vivo. Rapid NF-κB activation has been found to be an important signaling step in E2-mediated protection in cell culture, in vivo ischemia, and trauma-hemorrhage. In the current study, we investigated the signaling cascades linking E2 signaling with NF-κB activation and the protective response and compared them with the effects of two selective estrogen receptor modulators (SERMs), raloxifene and tamoxifen. Two candidate pathways, mitogen-activated protein kinases and phosphatidylinositol-3-kinase (PI3K) were studied. Selective inhibitors were used to identify each pathway's contribution to NF-κB activation. Treatment of human coronary artery endothelial cells with E2 activated PI3K/Akt, p38, and JNK, all of which activated ERK1/2 followed by NF-κB activation. The combined activation of Akt, p38, and JNK was essential to activate NF-κB. The two SERMs activated PI3K and p38, which then phosphorylated ERK1/2 and activated NF-κB independent of the JNK pathway. Nuclear factor κB activation by these compounds protected cells from hypoxia/reoxygenation injury. However, E2, unlike either SERM, led to modest increases in apoptosis through the JNK pathway. Selective estrogen receptor modulator treatment led to increased expression of the protective proteins, Mn superoxide dismutase, and endothelial nitric oxide synthase, which was not seen with E2. These results provide new insight into the pathways activating NF-κB by E2 and SERMs and demonstrate that SERMs may have greater protective benefits than E2 in adult endothelial cells and potentially in vivo, as well.     

Effects of osteoporotic cytokines in ovary-intact and ovariectomised rats with induced hyperthyroidism; is skeletal responsiveness to thyroid hormone altered in estrogen deficiency?

This experimental study was designed to examine the effects of hyperthyroidism on osteoporotic cytokines such as interleukin (IL)-1beta, IL-6 and tumor necrosis factor (TNF)-alpha in the physiological concentrations and in the deficiency of estrogen. We investigated the effects of thyroid hormones on cytokines and bone metabolism in L-thyroxine induced ovary-intact and ovariectomised rats, as levels of cytokines were increased in hyperthyroidism. The rats were divided into three groups. In the first group, L-thyroxine-induced hyperthyroid rats were ovariectomised (OVX), while the OVX rats were administered L-thyroxine in the second group. The third group received sham-operation. Blood samples taken from the tail vein of rats were analyzed for plasma T3, T4, TSH and serum IL-1beta, IL-6, TNFalpha, calcium (Ca), phosphorous (P), parathyroid hormone (PTH), alkaline phosphatase (ALP), bone-specific alkaline phosphatase (b-ALP). L-thyroxine administration increased the cytokines, ALP and b-ALP and decreased PTH, while there was no change in Ca and P. However, the ovariectomy of these rats did not change the levels of cytokines, Ca, P, PTH, ALP, and b-ALP. In ovariectomised rats, the cytokines, ALP and b-ALP increased but not Ca and P conversely, PTH decreased. L-thyroxine administration to ovariectomised rats did not change the levels of cytokines, Ca, P, PTH, ALP and b-ALP. In sham-operated rats there was no change in any of the parameters compared with initial values. Thyroid hormones may not be effective on bone metabolism in estrogen deficiency.     

Association of estrogen receptor α gene polymorphisms with BMD and their affect on estradiol levels in pre- and postmenopausal women in south Indian population from Andhra Pradesh

BackgroundOsteoporosis is a multifactorial disorder with a strong genetic component and ESR1 is suggested as a candidate gene for osteoporosis. Therefore the present study is aimed to investigate the role of ESR1 gene polymorphisms and its influence on estradiol levels and BMD in osteoporotic women of Indian ethnicity.MethodsFour-hundred twenty-seven osteoporotic women and 460 age matched controls were included in the study. ESR1 gene polymorphism was assessed by PCR-RFLP method. Serum estradiol was measured by ELISA.ResultsThe frequency of pp and xx genotypes as well as p and x alleles was significantly high in pre- and postmenopausal osteoporotics when compared to controls (p < 0.001). They had low BMD and estradiol levels in comparison with PP and XX genotype individuals (p < 0.05).ConclusionThe ESR1 gene is associated with low bone mass and low estradiol levels in all our study subjects. It is likely that the allele exerts its influence on the bone in early adulthood leading to an increased risk of osteoporosis later in life.