Inhibition of the mammalian target of rapamycin blocks epilepsy progression in NS-Pten conditional knockout mice

Purpose: Increased activity of mTOR Complex 1 (mTORC1) has been demonstrated in cortical dysplasia and tuberous sclerosis complex, as well as in animal models of epilepsy. Recent studies in such models revealed that inhibiting mTORC1 with rapamycin effectively suppressed seizure activity. However, seizures can recur after treatment cessation, and continuous rapamycin exposure can adversely affect animal growth and health. Here, we evaluated the efficacy of an intermittent rapamycin treatment protocol on epilepsy progression using neuron subset‐specific‐Pten (NS‐Pten) conditional knockout mice. Methods: NS‐Pten knockouts were treated with a single course of rapamycin during postnatal weeks 4 and 5, or intermittently over a period of 5 months. Epileptiform activity was monitored using video–electroencephalography (EEG) recordings, and mossy fiber sprouting was evaluated using Timm staining. Survival and body weight were assessed in parallel. Key Findings: NS‐Pten knockouts treated with a single course of rapamycin had recurrence of epilepsy 4–7 weeks after treatment ended. In contrast, epileptiform activity remained suppressed, and survival increased if knockout mice received additional rapamycin during weeks 10–11 and 16–17. Aberrant mossy fiber sprouting, present by 4 weeks of age and progressing in parallel with epileptiform activity, was also blocked by rapamycin. Significance: These findings demonstrate that a single course of rapamycin treatment suppresses epileptiform activity and mossy fiber sprouting for several weeks before epilepsy recurs. However, additional intermittent treatments with rapamycin prevented this recurrence and enhanced survival without compromising growth. Therefore, these studies add to the growing body of evidence implicating an important role for mTORC1 signaling in epilepsy.     

Autocrine prolactin induced by the Pten–Akt pathway is required for lactation initiation and provides a direct link between the Akt and Stat5 pathways

Extrapituitary prolactin (Prl) is produced in humans and rodents; however, little is known about its in vivo regulation or physiological function. We now report that autocrine prolactin is required for terminal mammary epithelial differentiation during pregnancy and that its production is regulated by the Pten–PI3K–Akt pathway. Conditional activation of the PI3K–Akt pathway in the mammary glands of virgin mice by either Akt1 expression or Pten deletion rapidly induced terminal mammary epithelial differentiation accompanied by the synthesis of milk despite the absence of lobuloalveolar development. Surprisingly, we found that mammary differentiation was due to the PI3K–Akt-dependent synthesis and secretion of autocrine prolactin and downstream activation of the prolactin receptor (Prlr)–Jak–Stat5 pathway. Consistent with this, Akt-induced mammary differentiation was abrogated in Prl^−/−, Prlr^−/−, and Stat5^−/− mice. Furthermore, cells treated with conditioned medium from mammary glands in which Akt had been activated underwent rapid Stat5 phosphorylation in a manner that was blocked by inhibition of Jak2, treatment with an anti-Prl antibody, or deletion of the prolactin gene. Demonstrating a physiological requirement for autocrine prolactin, mammary glands from lactation-defective Akt1^−/−;Akt2^+/− mice failed to express autocrine prolactin or activate Stat5 during late pregnancy despite normal levels of circulating serum prolactin and pituitary prolactin production. Our findings reveal that PI3K–Akt pathway activation is necessary and sufficient to induce autocrine prolactin production in the mammary gland, Stat5 activation, and terminal mammary epithelial differentiation, even in the absence of the normal developmental program that prepares the mammary gland for lactation. Together, these findings identify a function for autocrine prolactin during normal development and demonstrate its endogenous regulation by the PI3K–Akt pathway.     

Cell type specificity of PI3K signaling in Pdk1- and Pten-deficient brains

Loss of PTEN causes unregulated activation of downstream components of phosphatidylinositol 3-kinase (PI3K) signaling, including PDK1, and disrupts normal nervous system development and homeostasis. We tested the contribution of Pdk1 to the abnormalities induced by Pten deletion in the brain. Conditional deletion of Pdk1 caused microcephaly. Combined deletion of Pdk1 and Pten rescued hypertrophy, but not migration defects of Pten-deficient neurons. Pdk1 inactivation induced strikingly different effects on the regulation of phosphorylated Akt in glia versus neurons. Our results show Pdk1-dependent and Pdk1-independent abnormalities in Pten-deficient brains, and demonstrate cell type specific differences in feedback regulation of the ubiquitous PI3K pathway.     

Striking the balance between PTEN and PDK1: it all depends on the cell context

The phosphatidyl-inosital-3 kinase (PI3K) signaling pathway is critical for normal brain development and function and is commonly hyperactivated in brain cancer. The PTEN (phosphatase and tensin homolog deleted on chromosome 10) tumor suppressor protein and phosphate-depended kinase 1 (PDK-1) are critical regulators of this pathway. In the July 15, 2009, issue of Genes & Development, Chalhoub and colleagues (pp. 1619–1624) demonstrate PDK1-dependent and PDK1-independent effects of conditional PTEN deletion in the brain, and they identify cell type-specific differences in feedback regulation of the PI3K pathway. These studies provide important insights as to how neurons and glia may differentially regulate PI3K signaling, yielding intriguing clues about targeting PTEN-deficient brain cancers.     

Effective clearance of intracellular Leishmania major in vivo requires Pten in macrophages

Leishmaniases are a major international public health problem, and macrophages are crucial for host resistance to this parasite. To determine if phosphatase and tensin homologue deleted on chromosome ten (Pten), a negative regulator of the PI3K pathway, plays a role in macrophage-mediated resistance to Leishmania, we generated C57BL/6 mice lacking Pten specifically in macrophages (LysMCrePten(flox/flox) mice). Examination of lesions resulting from Leishmania major infection showed that LysMCrePten(flox/flox) mice were more susceptible to the parasite than wild-type (WT) mice in the early phase of the infection, but were eventually able to eliminate the pathogen. In vitro Pten-deficient macrophages showed a reduced ability to kill parasites in response to IFN-gamma treatment, possibly because the mutant cells exhibited decreased TNF secretion that correlated with reductions in inducible nitric oxide synthase expression and nitric oxide production. In response to various TLR ligands, Pten-deficient macrophages produced less TNF and IL-12 but more IL-10 than WT cells. However, analysis of cells in the lymph nodes draining L. major inoculation sites indicated that both LysMCrePten(flox/flox) and WT mice developed normal Th1 responses following L. major infection, in line with the ability of LysMCrePten(flox/flox) mice to eventually eliminate the parasite. Our results indicate that the efficient clearance of intracellular parasites requires Pten in macrophages.     

Identification of cellular and genetic drivers of breast cancer heterogeneity in genetically engineered mouse tumour models

The heterogeneous nature of mammary tumours may arise from different initiating genetic lesions occurring in distinct cells of origin. Here, we generated mice in which Brca2, Pten and p53 were depleted in either basal mammary epithelial cells or luminal oestrogen receptor (ER)‐negative cells. Basal cell‐origin tumours displayed similar histological phenotypes, regardless of the depleted gene. In contrast, luminal ER‐negative cells gave rise to diverse phenotypes, depending on the initiating lesions, including both ER‐negative and, strikingly, ER‐positive invasive ductal carcinomas. Molecular profiling demonstrated that luminal ER‐negative cell‐origin tumours resembled a range of the molecular subtypes of human breast cancer, including basal‐like, luminal B and ‘normal‐like’. Furthermore, a subset of these tumours resembled the ‘claudin‐low’ tumour subtype. These findings demonstrate that not only do mammary tumour phenotypes depend on the interactions between cell of origin and driver genetic aberrations, but also multiple mammary tumour subtypes, including both ER‐positive and ‐negative disease, can originate from a single epithelial cell type. This is a fundamental advance in our understanding of tumour aetiology. © 2014 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.     

Autocrine prolactin: an emerging market for homegrown (prolactin) despite the imports

Prolactin (PRL) is a peptide hormone that is produced by the pituitary gland and is known to regulate lactogenic differentiation. There is a significant body of evidence that points to autocrine production of prolactin and activation of an autocrine/paracrine signaling pathway to regulate cell proliferation and migration and inhibition of cell death. This perspective highlights the recent study in the October 1, 2012, issue of Genes & Development by Chen and colleagues (pp. 2154–2168) that describes a mechanism for autocrine prolactin production and places the finding in the context of a role for prolactin in breast development and cancer.     

Tumour suppressor gene function in carcinoma‐associated fibroblasts: from tumour cells via EMT and back again?

Recent reports indicate that inactivation of the RB, TP53 or PTEN tumour suppressor genes is detected in tumour stroma of oropharyngeal, breast and other human cancers. Mouse models have validated the tumour-promoting effects of deleting Rb, Pten or p53 in fibroblasts that converts them from normal fibroblasts to carcinoma associated fibroblasts (CAFs). The tumour-promoting activity of CAFs in these contexts was associated with increased paracrine signaling to tumour cells through production of specific growth factors, chemokines and MMPs by CAFs. The conversion of NOFs into CAFs through acquisition of specific mutations, such as loss of tumour suppressors, or deregulated expression of microRNAs or key epigenetic events, can clearly occur independently of genetic and epigenetic changes in tumour cells but an alternative source of CAFs that is being reconsidered is that CAFs derive from the tumour cells by EMT. Recent mouse models employing lineage-tracing techniques have suggested that this can take place in vivo and the extent to which this is relevant more broadly is discussed. Copyright © 2013 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Pten associates with important gene regulatory network to fine-tune Müller glia-mediated zebrafish retina regeneration

Unlike mammals, zebrafish possess a remarkable ability to regenerate damaged retina after an acute injury. Retina regeneration in zebrafish involves the induction of Müller glia-derived progenitor cells (MGPCs) exhibiting stem cell-like characteristics, which are capable of restoring all retinal cell-types. The induction of MGPC through Müller glia-reprograming involves several cellular, genetic and biochemical events soon after a retinal injury. Despite the knowledge on the importance of Phosphatase and tensin homolog (Pten), which is a dual-specificity phosphatase and tumor suppressor in the maintaining of cellular homeostasis, its importance during retina regeneration remains unknown. Here, we explored the importance of Pten during zebrafish retina regeneration. The Pten gets downregulated upon retinal injury and is absent from the MGPCs, which is essential to trigger Akt-mediated cellular proliferation essential for retina regeneration. We found that the downregulation of Pten in the post-injury retina accelerates MGPCs formation, while its overexpression restricts the regenerative response. We observed that Pten regulates the proliferation of MGPCs not only through Akt pathway but also by Mmp9/Notch signaling. Mmp9-activity is essential to induce the proliferation of MGPCs in the absence of Pten. Lastly, we show that expression of Pten is fine-tuned through Mycb/histone deacetylase1 and Tgf-β signaling. The present study emphasizes on the stringent regulation of Pten and its crucial involvement during the zebrafish retina regeneration.     

非小细胞肺癌组织中PTEN.CYCLIND1表达的意义

目的 研究抑癌基因10q丢失的与张力蛋白同源的磷酸酶基因(PTEN)在非小细胞肺癌 (NSCLC)组织中的表达水平与NSCLC的发生和发展的关系。方法 采用免疫组化方法分别测定58 例NSCLC和10例肺部非恶性病变病理切片中PTEN、CYCLIND1的蛋白表达,并对58例NSCLC组织 中PTEN、CYCIND1的表达与患者的年龄、性别、吸烟情况、淋巴结转移情况及生存时间进行统计 分析。结果 PTEN的表达在肺鳞癌中阳性率为13/26、肺腺癌中为15/32;中高分化组肺癌中阳 性率20/33、低分化组中8/25;淋巴结转移组阳性率为7/32、无转移组阳性率为21/26。同时检测 CYCLIND1在NSCLC中的表达情况。10例肺部非恶性病变标本中,PTEN表达全部阳性、CYCLIND1全 部阴性。结论. PTEN的失表达在NSCLC发生、发展和转移中扮演着重要角色,其失表达与NSCLC 的分化程度(P<0.05)、淋巴结转移(P<0.05)、临床分期(P<0.05)有关,PTEN、CYCLIND1协同在 NSCLC的发生、发展中起重要作用。