R-spodin2 enhances canonical Wnt signaling to maintain the stemness of glioblastoma cells.

BACKGROUND: As newly identified Wnt enhancer, R-spondin gene family members have been linked to various cancers; however, their role in isocitrate dehydrogenase-wildtype subtype of human glioblastoma (GBM) cells remains unknown. METHODS: Human U87 and U251 cell lines were used to perform the experiments. GBM stem-like cells were enriched in stem cell growth media and induced to differentiate using retinoid acid or growth factor deprivation. Wnthigh and Wntlow subpopulations were isolated and evaluated by MTS, sphere formation, transwell migration and xenograft formation assays. RESULTS: R-spondin 2 but not R-spondin 3 potentiates Wnt/ß-catenin signaling in GBM cell lines. While R-spondin 2 does not affect cell growth, it induces the expression of pluripotent stem cell markers in combination with Wnt3A. GBM stem-like cells are endowed with intrinsic high activity of ß-catenin signaling, which can be further intensified by R-spondin 2. In addition, R-spondin2 promotes stem cell self-renewal and suppresses retinoid acid- or growth factor deprivation-induced differentiation, indicating R-spondin 2 maintains stem cell traits in GBM. On the other hand, we identify subpopulations of GBM cells that show distinctive responsiveness to Wnt/ß-catenin signaling. Interestingly, Wnthigh and Wntlow cells display distinctive biologic properties. Moreover, Wnthigh cell-inoculated xenografts exhibit enhanced tumorigenicity and increased expression levels of R-spondin 2 compared to Wntlow cell-inoculated xenografts. CONCLUSION: Our study reveals a novel regulatory mechanisms underlying the over-activation of ß-catenin-mediated signaling in the pathogenesis of GBM.

Disrupted interaction between CFTR and AF-6/afadin aggravates malignant phenotypes of colon cancer.

How mutations or dysfunction of CFTR may increase the risk of malignancies in various tissues remains an open question. Here we report the interaction between CFTR and an adherens junction molecule, AF-6/afadin, and its involvement in the development of colon cancer. We have found that CFTR and AF-6/afadin are co-localized at the cell-cell contacts and physically interact with each other in colon cancer cell lines. Knockdown of CFTR results in reduced epithelial tightness and enhanced malignancies, with increased degradation and reduced stability of AF-6/afadin protein. The enhanced invasive phenotype of CFTR-knockdown cells can be completely reversed by either AF-6/afadin over-expression or ERK inhibitor, indicating the involvement of AF-6/MAPK pathway. More interestingly, the expression levels of CFTR and AF-6/afadin are significantly downregulated in human colon cancer tissues and lower expression of CFTR and/or AF-6/afadin is correlated with poor prognosis of colon cancer patients. The present study has revealed a previously unrecognized interaction between CFTR and AF-6/afadin that is involved in the pathogenesis of colon cancer and indicated the potential of the two as novel markers of metastasis and prognostic predictors for human colon cancer.

Dynamically Regulated CFTR Expression and Its Functional Role in Cutaneous Wound Healing.

The physiological role of cystic fibrosis transmembrane conductance regulator (CFTR) in keratinocytes and skin wound healing is completely unknown. The present study shows that CFTR is expressed in the multiple layers of keratinocytes in mouse epidermis and exhibits a dynamic expression pattern in a dorsal skin wound healing model, with diminishing levels observed from day 3 to day 5 and re-appearing from day 7 to day 10 after wounding. Knockdown of CFTR in cultured human keratinocytes promotes cell migration but inhibits differentiation, while overexpression of CFTR suppresses migration but enhances differentiation, indicating an important role of CFTR in regulating keratinocyte behavior. In addition, we have demonstrated a direct association of CFTR with epithelial junction formation as knockdown of CFTR downregulates the expression of adhesion molecules, such as E-cadherin, ZO-1 and ß-catenin, and disrupts the formation of cell junction, while overexpression of CFTR enhances cell junction formation. More importantly, we have shown that ΔF508cftr-/- mice with defective CFTR exhibit delayed wound healing as compared to wild type mice, indicating that normal function of CFTR is critical for wound repair. Taken together, the present study has revealed a previously undefined role of CFTR in regulating skin wound healing processes, which may have implications in injury repair of other epithelial tissues.

Downregulation of CFTR promotes epithelial-to-mesenchymal transition and is associated with poor prognosis of breast cancer.

The epithelial-to-mesenchymal transition (EMT), a process involving the breakdown of cell-cell junctions and loss of epithelial polarity, is closely related to cancer development and metastatic progression. While the cystic fibrosis transmembrane conductance regulator (CFTR), a Cl(-) and HCO3(-) conducting anion channel expressed in a wide variety of epithelial cells, has been implicated in the regulation of epithelial polarity, the exact role of CFTR in the pathogenesis of cancer and its possible involvement in EMT process have not been elucidated. Here we report that interfering with CFTR function either by its specific inhibitor or lentiviral miRNA-mediated knockdown mimics TGF-ß1-induced EMT and enhances cell migration and invasion in MCF-7. Ectopic overexpression of CFTR in a highly metastatic MDA-231 breast cancer cell line downregulates EMT markers and suppresses cell invasion and migration in vitro, as well as metastasis in vivo. The EMT-suppressing effect of CFTR is found to be associated with its ability to inhibit NFκB targeting urokinase-type plasminogen activator (uPA), known to be involved in the regulation of EMT. More importantly, CFTR expression is found significantly downregulated in primary human breast cancer samples, and is closely associated with poor prognosis in different cohorts of breast cancer patients. Taken together, the present study has demonstrated a previously undefined role of CFTR as an EMT suppressor and its potential as a prognostic indicator in breast cancer.

Regulation of miR-101/miR-199a-3p by the epithelial sodium channel during embryo implantation: involvement of CREB phosphorylation.

In our previous study, we have demonstrated that the epithelial sodium channel (ENaC) mediates the embryo-derived signals leading to the activation of CREB and upregulation of cyclooxygenase type 2 (COX2) required for embryo implantation. This study aims to investigate whether microRNAs (miRNAs) are involved in the ENaC-induced upregulation of COX2 during embryo implantation. The results show that the levels of miR-101 and miR-199a-3p, two COX2 targeting miRNAs, are reduced by ENaC activation, and increased by ENaC inhibition or knock-down of ENaC subunit (ENaCα) in human endometrial surface epithelial (HES) cells or in mouse uteri during implantation. Phosphorylation of CREB is induced by the activation of ENaC, and blocked by ENaC inhibition or knockdown in HES cells. Knockdown of ENaCα or CREB in HES cells or in mouse uterus in vivo results in increases in miR-101 and miR-199a-3p, accompanied with decreases in COX2 protein levels and reduction in implantation rate. The downregulation of COX2 caused by knockdown of ENaC or CREB can be recovered by the inhibitors of miR-101 or miR-199a-3p in HES cells. These results reveal a novel molecular mechanism modulating COX2 expression during embryo implantation via ENaC-dependent CREB activation and COX2-targeting miRNAs.

CFTR negatively regulates cyclooxygenase-2-PGE(2) positive feedback loop in inflammation.

Cystic fibrosis (CF) is an autosomal recessive disorder caused by mutations of the cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP-dependent anion channel mostly expressed in epithelia. Accumulating evidence suggests that CF airway epithelia are overwhelmed by excessive inflammatory cytokines and prostaglandins (PGs), which eventually lead to the over-inflammatory condition observed in CF lung disease. However, the exact underlying mechanism remains elusive. In this study, we observed increased cyclooxygenase-2 (COX-2) expression and over-production of prostaglandin E(2) (PGE(2)) in human CF bronchial epithelia cell line (CFBE41o--) with elevated NF-κB activity compared to a wild-type airway epithelial cell line (16HBE14o--). Moreover, we demonstrated that CFTR knockout mice had inherently higher levels of COX-2 and NF-κB activity, supporting the notion that lack of CFTR results in hyper-inflammatory signaling. In addition, we identified a positive feedback loop for production of PGE(2) involving PKA and transcription factor, CREB. More importantly, overexpression of wild-type CFTR significantly suppressed COX-2 expression in CFBE41o- cells, and wild-type CFTR protein expression was significantly increased when 16HBE14o-- cells were challenged with LPS as well as PGE(2), indicating possible involvement of CFTR in negative regulation of COX-2/PGE(2). In conclusion, CFTR is a negative regulator of PGE(2)-mediated inflammatory response, defect of which may result in excessive activation of NF-κB, leading to over production of PGE(2) as seen in inflammatory CF tissues.

Lymphocyte CFTR promotes epithelial bicarbonate secretion for bacterial killing.

The expression of cystic fibrosis transmembrane conductance regulator (CFTR) in lymphocytes has been reported for nearly two decades; however, its physiological role remains elusive. Here, we report that co-culture of lymphocytes with lung epithelial cell line, Calu-3, promotes epithelial HCO(3)- production/secretion with up-regulated expression of carbonic anhydrase 2 and 4 (CA-2, CA-4) and enhanced bacterial killing capability. The lymphocyte-enhanced epithelial HCO(3)- secretion and bacterial killing activity was abolished when Calu3 cells were co-cultured with lymphocytes from CFTR knockout mice, or significantly reduced by interfering with E-cadherin, a putative binding partner of CFTR. Bacterial lipopolysaccharide (LPS)-induced E-cadherin and CA-4 expression in the challenged lung was also found to be impaired in CFTR knockout mice compared to that of the wild-type. These results suggest that the interaction between lymphocytes and epithelial cells may induce a previously unsuspected innate host defense mechanism against bacterial infection by stimulating epithelial HCO(3)- production/secretion, which requires CFTR expression in lymphocytes.

Dedifferentiation-reprogrammed mesenchymal stem cells with improved therapeutic potential.

Stem cell transplantation has been shown to improve functional outcome in degenerative and ischemic disorders. However, low in vivo survival and differentiation potential of the transplanted cells limits their overall effectiveness and thus clinical usage. Here we show that, after in vitro induction of neuronal differentiation and dedifferentiation, on withdrawal of extrinsic factors, mesenchymal stem cells (MSCs) derived from bone marrow, which have already committed to neuronal lineage, revert to a primitive cell population (dedifferentiated MSCs) retaining stem cell characteristics but exhibiting a reprogrammed phenotype distinct from their original counterparts. Of therapeutic interest, the dedifferentiated MSCs exhibited enhanced cell survival and higher efficacy in neuronal differentiation compared to unmanipulated MSCs both in vitro and in vivo, with significantly improved cognition function in a neonatal hypoxic-ischemic brain damage rat model. Increased expression of bcl-2 family proteins and microRNA-34a appears to be the important mechanism giving rise to this previously undefined stem cell population that may provide a novel treatment strategy with improved therapeutic efficacy.

Bak Foong Pills induce an analgesic effect by inhibiting nociception via the somatostatin pathway in mice.

Dysmenorrhoea, defined as cramping pain in the lower abdomen occurring before or during menstruation, affects, to varying degrees, up to 90% of women of child-bearing age. We investigated whether BFP (Bak Foong Pills), a traditional Chinese medicine treatment for dysmenorrhoea, possesses analgesic properties. Results showed that BFP was able to significantly reduce pain responses following subchronic treatment for 3 days, but not following acute (1 h) treatment in response to acetic acid-induced writhing in C57/B6 mice. The analgesic effect was not due to inhibition of COX (cyclo-oxygenase) activity, evidenced by the lack of inhibition of prostacyclin and PGE2 (prostaglandin E2) production. Molecular analysis revealed that BFP treatment modulated the expression of a number of genes in the spinal cord of mice subjected to acetic acid writhing. RT-PCR (reverse transcription-PCR) analysis of spinal cord samples showed that both sst4 (somatostatin receptor 4) and sst2 receptor mRNA, but not µOR (µ-opiate receptor) and NK1 (neurokinin-1) receptor mRNA, were down-regulated following BFP treatment, thus implicating somatostatin involvement in BFP-induced analgesia. Administration of c-som (cyclo-somatostatin), a somatostatin antagonist, prior to acetic acid-induced writhing inhibited the analgesic effect. Thus subchronic treatment with BFP has anti-nociceptive qualities mediated via the somatostatin pathway.

Flavonoid Myricetin Modulates GABA(A) Receptor Activity through Activation of Ca(2+) Channels and CaMK-II Pathway.

The flavonoid myricetin is found in several sedative herbs, for example, the St. John's Wort, but its influence on sedation and its possible mechanism of action are unknown. Using patch-clamp technique on a brain slice preparation, the present study found that myricetin promoted GABAergic activity in the neurons of hypothalamic paraventricular nucleus (PVN) by increasing the decay time and frequency of the inhibitory currents mediated by GABA(A) receptor. This effect of myricetin was not blocked by the GABA(A) receptor benzodiazepine- (BZ-) binding site antagonist flumazenil, but by KN-62, a specific inhibitor of the Ca(2+)/calmodulin-stimulated protein kinase II (CaMK-II). Patch clamp and live Ca(2+) imaging studies found that myricetin could increase Ca(2+) current and intracellular Ca(2+) concentration, respectively, via T- and L-type Ca(2+) channels in rat PVN neurons and hypothalamic primary culture neurons. Immunofluorescence staining showed increased phosphorylation of CaMK-II after myricetin incubation in primary culture of rat hypothalamic neurons, and the myricetin-induced CaMK-II phosphorylation was further confirmed by Western blotting in PC-12 cells. The present results suggest that myricetin enhances GABA(A) receptor activity via calcium channel/CaMK-II dependent mechanism, which is distinctively different from that of most existing BZ-binding site agonists of GABA(A) receptor.

Human pluripotent stem cell-derived ectomesenchymal stromal cells promote more robust functional recovery than umbilical cord-derived mesenchymal stromal cells after hypoxic-ischaemic brain damage.

Aims: Hypoxic-ischaemic encephalopathy (HIE) is one of the most serious complications in neonates and infants. Mesenchymal stromal cell (MSC)-based therapy is emerging as a promising treatment avenue for HIE. However, despite its enormous potential, the clinical application of MSCs is limited by cell heterogeneity, low isolation efficiency and unpredictable effectiveness. In this study, we examined the therapeutic effects and underlying mechanisms of human pluripotent stem cell-derived ectomesenchymal stromal cells (hPSC-EMSCs) in a rat model of HIE. Methods: hPSC-EMSCs were induced from either human embryonic stem cells or induced pluripotent stem cells. Stem cells or the conditioned medium (CM) derived from stem cells were delivered intracranially or intranasally to neonatal rats with HIE. Human umbilical cord-derived MSCs (hUC-MSCs) were used as the therapeutic comparison control and phosphate-buffered saline (PBS) was used as a negative control. Lesion size, apoptosis, neurogenesis, astrogliosis and microgliosis were evaluated. The rotarod test and Morris water maze were used to determine brain functional recovery. The PC-12 cell line, rat primary cortical neurons and neural progenitor cells were used to evaluate neurite outgrowth and the neuroprotective and neurogenesis effects of hPSC-EMSCs/hUC-MSCs. RNA-seq and enzyme-linked immunosorbent assays were used to determine the secretory factors that were differentially expressed between hPSC-EMSCs and hUC-MSCs. The activation and suppression of extracellular signal-regulated kinase (ERK) and cAMP response element-binding protein (CREB) were characterised using western blotting and immunofluorescent staining. Results: hPSC-EMSCs showed a higher neuroprotective potential than hUC-MSCs, as demonstrated by a more significant reduction in lesion size and apoptosis in the rat brain following hypoxia-ischaemia (HI). Compared with PBS treatment, hPSC-EMSCs promoted endogenous neurogenesis and alleviated astrogliosis and microgliosis. hPSC-EMSCs were more effective than hUC-MSCs. hPSC-EMSCs achieved a greater recovery of brain function than hUC-MSCs and PBS in rats with HIE. CM derived from hPSC-EMSCs had neuroprotective and neurorestorative effects in vitro through anti-apoptotic and neurite outgrowth- and neurogenesis-promoting effects. Direct comparisons between hPSC-EMSCs and hUC-MSCs revealed the significant enrichment of a group of secretory factors in hPSC-EMSCs, including nerve growth factor (NGF), platelet-derived growth factor-AA and transforming growth factor-ß2, which are involved in neurogenesis, synaptic transmission and neurotransmitter transport, respectively. Mechanistically, the CM derived from hPSC-EMSCs was found to potentiate NGF-induced neurite outgrowth and the neuronal differentiation of NPCs via the ERK/CREB pathway. Suppression of ERK or CREB abolished CM-potentiated neuritogenesis and neuronal differentiation. Finally, intranasal delivery of the CM derived from hPSC-EMSCs significantly reduced brain lesion size, promoted endogenous neurogenesis, mitigated inflammatory responses and improved functional recovery in rats with HIE. Conclusion: hPSC-EMSCs promote functional recovery after HI through multifaceted neuromodulatory activities via paracrine/trophic mechanisms. We propose the use of hPSC-EMSCs for the treatment of HIE, as they offer an excellent unlimited cellular source of MSCs.

Human embryonic stem cell-derived neural crest model unveils CD55 as a cancer stem cell regulator for therapeutic targeting in MYCN-amplified neuroblastoma.

BACKGROUND: Neuroblastoma (NB) is a common childhood malignant tumor of neural crest (NC) origin with remarkable heterogeneity in outcomes. Amplification of the oncogene MYCN is strongly associated with highly malignant behaviour and poor prognosis. METHODS: This study aims to use a human embryonic stem cell (hESC)-derived NC model to identify novel downstream effectors of MYCN that can be potentially used as prognostic marker and/or therapeutic target. RESULTS: We show that MYCN-driven NB derived from human neural crest cells (hNCCs) recapitulate the pathological and molecular features of MYCN-amplified neuroblastoma (MNA-NB). By using this platform, we identify a group of 14 surface protein-encoding genes that are associated with MYCN expression level in MNA-NB. Among these genes, high CD55 expression is correlated with poor survival in MNA-NB but not in non-MNA-NB. Furthermore, CD55 promotes tumorigenesis, tumor growth, and cancer stemness in MNA-NB cell lines (MNA-NBL) through regulating the JNK pathway. Mechanistically, MYCN binds to both canonical and noncanonical E-boxes on the promoter of CD55 to regulate its transcriptional expression. Finally, neutralizing antibody targeting CD55 significantly attenuates cancer stemness, suppresses tumor growth, and improves survival exclusively in MNA-NBL-inoculated mice. CONCLUSION: MYCN shapes CD55 into a cancer stem cell regulator which represents a prognostic marker and therapeutic target of MNA-NB. The hESC-derived NC model serves as a valuable platform for investigating NB initiation and progression and developing potential therapeutic targets.

An epididymis-specific beta-defensin is important for the initiation of sperm maturation.

Although the role of the epididymis, a male accessory sex organ, in sperm maturation has been established for nearly four decades, the maturation process itself has not been linked to a specific molecule of epididymal origin. Here we show that Bin1b, a rat epididymis-specific beta-defensin with antimicrobial activity, can bind to the sperm head in different regions of the epididymis with varied binding patterns. In addition, Bin1b-expressing cells, either of epididymal origin or from a Bin1b-transfected cell line, can induce progressive sperm motility in immotile immature sperm. This induction of motility is mediated by the Bin1b-induced uptake of Ca(2+), a mechanism that has a less prominent role in maintaining motility in mature sperm. In vivo antisense experiments show that suppressed expression of Bin1b results in reduced binding of Bin1b to caput sperm and in considerable attenuation of sperm motility and progressive movement. Thus, beta-defensin is important for the acquisition of sperm motility and the initiation of sperm maturation.

Involvement of CFTR in uterine bicarbonate secretion and the fertilizing capacity of sperm.

Cystic fibrosis transmembrane conductance regulator (CFTR) is a cAMP-activated chloride channel expressed in a wide variety of epithelial cells, mutations of which are responsible for the hallmark defective chloride secretion observed in cystic fibrosis (CF). Although CFTR has been implicated in bicarbonate secretion, its ability to directly mediate bicarbonate secretion of any physiological significance has not been shown. We demonstrate here that endometrial epithelial cells possess a CFTR-mediated bicarbonate transport mechanism. Co-culture of sperm with endometrial cells treated with antisense oligonucleotide against CFTR, or with bicarbonate secretion-defective CF epithelial cells, resulted in lower sperm capacitation and egg-fertilizing ability. These results are consistent with a critical role of CFTR in controlling uterine bicarbonate secretion and the fertilizing capacity of sperm, providing a link between defective CFTR and lower female fertility in CF.

Regulatory mechanism underlying cyclic changes in mouse uterine bicarbonate secretion: role of estrogen.

Our previous study has demonstrated that bicarbonate in the uterine fluid plays an indispensable role in sperm capacitation. However, the cellular mechanisms underlying the formation of bicarbonate-rich uterine fluid and the regulatory mechanism remained largely unknown. In this study, the expression profiles of bicarbonate transport/production proteins, the cystic fibrosis transmembrane conductance regulator (CFTR), SLC26A6, carbonic anhydrase 2 (CAR2, CA2) and CAR12 (CA12), throughout the estrous cycle, were examined in the mouse uterus by western blot. The results showed that the maximum expression levels of the proteins examined were observed at estrus. Luminal surface pH measurements showed that the resting uterine surface pH at estrus was significantly higher than that at diestrus, which could be reduced significantly by CFTR blocker, diphenylamine-2,2'-dicarboxylic acid, SLC26A6 inhibitor, 4',4'-diisothiocyanostilbene-2',2'-disulfonic acid, and CA inhibitor, acetazolamide. In ovariectomized mice and primary culture of endometrial epithelial cells, estrogen could upregulate CFTR, SLC26A6, CAR2, and CAR12 expression with a corresponding increase in the bicarbonate-dependent short-circuit current (I(sc)) and endometrial surface pH. The present results have demonstrated dynamic changes in uterine bicarbonate secretion and expression of the proteins involved in bicarbonate secretion during the estrous cycle and suggested a novel role of estrogen in regulating uterine bicarbonate transport, which may be important for successful reproduction.

Involvement of calpain-I and microRNA34 in kanamycin-induced apoptosis of inner ear cells.

Inner ear cells, including hair cells, spiral ganglion cells, stria vascularis cells and supporting cells on the basilar membrane, play a major role in transducing hearing signals and regulating inner ear homoeostasis. However, their functions are often damaged by antibiotic-induced ototoxicity. Apoptosis is probably involved in inner ear cell injury following aminoglycoside treatment. Calpain, a calcium-dependent protease, is essential for mediating and promoting cell death. We have therefore investigated the involvement of calpain in the molecular mechanism underlying ototoxicity induced by the antibiotic kanamycin in mice. Kanamycin (750 mg/kg) mainly induced cell death of cochlear cells, including stria vascularis cells, supporting cells and spiral ganglion cells, but not hair cells within the organ of Corti. Cell death due to apoptosis occurred in a time-dependent manner with concomitant up-regulation of calpain expression. Furthermore, the expression levels of two microRNAs, mir34a and mir34c, were altered in a dose-dependent manner in cochlear cells. These novel findings demonstrated the involvement of both calpain and microRNAs in antibiotic-induced ototoxicity.

Switching from bone marrow-derived neurons to epithelial cells through dedifferentiation and translineage redifferentiation.

While the ability of stem cells to switch lineages has been suggested, the route(s) through which this may happen is unclear. To date, the best characterized adult stem cell population considered to possess transdifferentiation capacity is BM-MSCs (bone marrow mesenchymal stem cells). We investigated whether BM-MSCs that had terminally differentiated into the neural or epithelial lineage could be induced to transdifferentiate into the other phenotype in vitro. Our results reveal that neuronal phenotypic cells derived from adult rat bone marrow cells can be switched to epithelial phenotypic cells, or vice versa, by culture manipulation allowing the differentiated cells to go through, first, dedifferentiation and then redifferentiation to another phenotype. Direct transdifferentiation from differentiated neuronal or epithelial phenotype to the other differentiated phenotype cannot be observed even when appropriate culture conditions are provided. Thus, dedifferentiation appears to be a prerequisite for changing fate and differentiating into a different lineage from a differentiated cell population.

CFTR is required for cellular entry and internalization of Chlamydia trachomatis.

Chlamydia trachomatis is an obligate intracellular Gram-negative pathogen affecting over 600 million people worldwide with 92 million new cases occurring globally each year. C. trachomatis enter the cells and replicate to infect different tissues/organs, giving rise to a spectrum of pathological conditions; however, the exact mechanism or receptor(s) for their entry is not well understood. Here we report that CFTR (cystic fibrosis transmembrane conductance regulator), an apical epithelial anion channel, is required for cellular entry and internalization of C. trachomatis. Human epithelial cell lines expressing functional CFTR internalized more C. trachomatis than the cells expressing mutant Delta508 CFTR. The in vitro cellular uptake of C. trachomatis can be blocked by CFTR inhibitors or antibody, and the in vivo cellular uptake of C. trachomatis in CFTR mutant (CFTR(-/-)) mice was significantly less compared with that in the wild-type. Direct interaction between CFTR and C. trachomatis LPS (lipopolysaccharide) is demonstrated by their immune-co-localization and co-immunoprecipitation. Despite an increase in CFTR expression observed upon C. trachomatis LPS challenge, a reduction in its ion channel activity is observed, consistent with the notion that CFTR functions as a receptor for cellular entry and internationization of C. trachomatis, with compromised ion-channel function. These findings, for the first time, demonstrate that CFTR functions as a cell-surface receptor for epithelial cell entry, and internalization of C. trachomatis and these findings may lead to the development of new treatment strategies to curtail the spread of chlamydial infections.

Functional crosstalk between mTORC1/p70S6K pathway and heterochromatin organization in stress-induced senescence of MSCs.

BACKGROUND: Stem cell senescence has been proposed as one of the major drivers of aging, and MSC senescence contributes to aging-related diseases. Activation of mTORC1 pathway and heterochromatin organization have been characterized as two characteristics of senescent cells; however, whether mTORC1 pathway interacts with heterochromatin organization and contributes to MSC senescence remains unknown. In this study, we investigated the interaction between heterochromatin organization and mTORC1/p70S6K pathway in stress-induced MSC senescence. METHODS: The stress-induced senescence models were established in human umbilical cord-derived MSCs by doxorubicin (Dox) or H2O2. Cellular senescence was evaluated by ß-Gal activity, upregulation of cell cycle suppressor genes, and expression of SASP. Activation of heterochromatin organization and mTORC1 pathway was determined by Western blot and immunofluorescent staining. A D-galactose (D-Gal)-induced aging model was established in rats to evaluate the crosstalk between heterochromatin and mTORC1 pathway in vivo. RESULTS: We found that heterochromatin organization was provoked at the early stage of Dox- or H2O2-induced senescence. Disruption of heterochromatin organization led to robust DNA damage response and exacerbated cellular senescence. Suppression of mTORC1/p70S6K pathway by either rapamycin or p70S6K knockdown promoted heterochromatin organization and ameliorated Dox- or H2O2-induced DNA damage and senescence. In contrast, direct activation of mTORC1 by MHY1485 impaired heterochromatin organization and aggravated stress-induced senescence. Moreover, concomitant activation of mTORC1 pathway and heterochromatin organization was found in D-galactose-induced osteoporosis model in rats. Rapamycin alleviated cellular senescence and promoted heterochromatin organization in BMSCs derived from D-galactose-treated rats. CONCLUSIONS: Altogether, our study indicates the existence of a complex interplay between the mTORC1/p70S6K pathway and the heterochromatin organization during stress-induced MSC senescence, with important implications for the understanding of aging as well as for its prevention and treatment.

A suppressor of multiple extracellular matrix-degrading proteases and cancer metastasis.

Cancer metastasis remains the most poorly understood process in cancer biology. It involves the degradation of extracellular matrix (ECM) proteins by a series of 'tumour-associated' proteases. Here we report the identification of a novel protease suppressor, NYD-SP8, which is located on human chromosome 19q13.2. NYD-SP8 encodes a 27 kD GPI-anchored cell surface protein, which shows structural homology to urokinase plasminogen activator receptor (uPAR). Co-immunoprecipitation experiments showed that NYD-SP8 binds to uPA/uPAR complexes and interfere with active uPA production. Overexpression of NYD-SP8 results in reducing activities of the three major classes of proteases known to be involved in ECM degradation, including uPA, matrix metalloproteinases (MMPs) and cathepsin B, leading to suppression of both in vitro and in vivo cancer cell invasion and metastasis. These data demonstrate an important role of NYD-SP8 in regulating ECM degradation, providing a novel mechanism that modulates urokinase signalling in the suppression of cancer progression.