Hypermethylation of TGF-β1 gene promoter in gastric cancer

AIM:To examine transforming growth factor-β1(TGF-β1)promoter methylation in gastric cancer and to determine if Helicobacter pylori(H.pylori)or interleukin(IL)-1β could induce TGF-β1 hypermethylation in vitro.METHODS:We examined the frequency and extent of TGF-β1 promoter methylation using methylationspecific PCR in the gastric tissues from 47 gastric cancer patients and 39 non-gastric cancer subjects.H.pylori infection was confirmed by a positive result from either a serological test,histological analysis or C13urea breath test.GES-1 and MKN-45 cells co-cultured with H.pylori or treated with IL-1β for 12,24 and 48 h in vitro tested the effects of H.pylori or IL-1β on TGF-1β.RESULTS:Twenty-four/forty-seven(51%)cases of gastric cancer(GC)tissues showed TGF-β1 promoter methylation,15/47(31.9%)cases of matched noncancerous gastric mucosa tissues from the GC patients,and 11/39(28%)case of the normal gastric mucosa tissues from non-GC subjects showed TGF-β1 promoter methylation(51%vs 28%,P<0.05).Significantly higher levels of methylation of TGF-β1 were found in the tumor tissues than in non-tumor tissues from GC patients(0.24±0.06 vs 0.17±0.04,P<0.05)and normal gastric tissues from non-GC subjects(0.24±0.06 vs 0.15±0.03,P<0.05).TGF-β1 methylation was found in 48.3% of H.pylori-positive gastric mucosal tissues whereas only 23.1% of H.pylori-negative gastric mucosal tissues showed TGF-β1 methylation(48.3%vs 23.1%,P<0.05).IL-1β appeared to induce a dose-dependent methylation of TGF-β1 and the strongest methylation was observed in GES-1 cells treated with 2.5 ng/mL of IL-1β for 48 h.Further studies showed that pre-treatment of GES-1 cells with 20ng/mL IL-1RA for 1 h could partially abolish the effect of IL-1β on TGF-β1 methylation.Infection of GES-1cells by H.pylori was not found to induce significant TGF-β1 promoter methylation.CONCLUSION:Our data revealed that TGF-1 promoter is methylated in GC patients.IL-1β may be an important mediator for H.pylori induced gene methylation during GC     

Combinatorial antibody libraries from cancer patients yield ligand-mimetic Arg-Gly-Asp-containing immunoglobulins that inhibit breast cancer metastasis

Combinatorial antibody libraries have the potential to display the entire immunological record of an individual, allowing one to detect and recover any antibody ever made, irrespective of whether it is currently being produced. We have termed this the "fossil record" of an individual's antibody response. To determine whether cancer patients have ever made antibodies with disease-fighting potential, we screened combinatorial antibody libraries from cancer patients for immunoglobulins that can identify metastatic tumor cells. This strategy yielded human antibodies specific for the activated conformation of the adhesion receptor integrin alphavbeta3 that is associated with a metastatic phenotype. In a remarkable example of convergent evolution, two of these antibodies were shown to contain the Arg-Gly-Asp integrin recognition motif of the natural ligand within the third complementarity-determining region of the heavy chain. These antibodies interfered with lung colonization by human breast cancer cells in a mouse model and inhibited existing metastatic disease. Our data imply that, at least at some time, these antibodies were part of a patient's surveillance system against metastatic cells, targeting the activated conformer of integrin alphavbeta3 and disrupting its functions. The ligand-mimetic nature of these antibodies, combined with specificity for a single receptor, is unique in the integrin-ligand repertoire. The convergent evolution of critical sequences in antibodies and other ligands that bind to the same target means that the immune response has sufficient power to find a best chemical solution for the optimization of binding energy, even though antibodies evolve in real time, as compared with billions of years for the natural ligand.     

Effects of urotensin-Ⅱ on cytokines in early acute liver failure in mice

AIM:To investigate urotensin-Ⅱ(UⅡ) and its effects on tumor necrosis factor(TNF)-α and interleukin(IL)-1β in early acute liver failure(ALF).METHODS:We investigated the time-dependent alteration in UⅡ levels and its effects on TNF-αand IL-1β in liver and blood in the early stage of lipopolysaccharide/D-galactosamine-induced ALF.RESULTS:After lipopolysaccharide/D-galactosamine challenge,UⅡ rose very rapidly and reached a maximal level 0.5 h,and the level remained significantly elevated after 2 h(P 0.05).Six hours after challenge,UⅡ began to degrade,but remained higher than at 0 h(P 0.05).Pretreatment with urantide,an inhibitor of the UⅡ receptor,suppressed the degree of UⅡ increase in liver and blood at 6 h after challenge(P 0.05 vs paired controls).In addition,liver and blood TNF-α increased from 1 to 6 h,and reached a peak at 1 and 2 h,respectively; however,IL-1β did not rise until 6 h after challenge.Urantide pretreatment inhibited the degree of TNF-α and IL-1β increase following downregulation of UⅡ post-challenge(all P 0.05).CONCLUSION:UⅡ plays a role in the pathogenesis and priming of ALF by triggering an inflammatory cascade and driving the early release of cytokines in mice.     

Liver metastasis is the only independent prognostic factor in AFP-producing gastric cancer

AIM:To investigate differences between common gastric cancer andα-fetoprotein(AFP)-producing gastric cancer according to the presence or absence of liver metastasis.METHODS:Between 1997 and 2011,1299 patients underwent gastrectomy for gastric cancer(GC)at our institute and their hospital records were reviewed retrospectively.Patients were immunohistochemically divided into two groups:23 patients(1.8%)with AFPproducing GC and 1276 patients(98.2%)without it.RESULTS:AFP-producing GC patients had a significantly higher incidence of deeper tumors,venous invasion,lymphatic invasion,lymph node metastasis,and liver metastasis and a poorer prognosis(P<0.005)than those without AFP-producing GC.However,multi-variate analysis revealed that AFP-positivity was not an independent prognostic factor.The prognosis of AFPproducing GC was similar to that of AFP-non producing GC according to the presence or absence of liver metastasis.Concerning recurrence,47.8%of patients(11/23)with AFP-producing GC and 20.0%of patients(256/1276)without AFP-producing GC exhibited recurrence.Liver metastasis[90.9%(10/11)]was the most prevalent in AFP-producing GC patients.Multivariate analysis revealed that liver metastasis was the only independent prognostic factor in AFP-producing GC(HR=17.6,95%CI:2.1-147.1;P=0.0081).CONCLUSION:AFP-producing GC is similar to common GC without liver metastasis,which should be specifically targeted in an effort to improve the prognosis of AFP-producing GC patients.     

How main-chains of proteins explore the free-energy landscape in native states

Understanding how a single native protein diffuses on its free-energy landscape is essential to understand protein kinetics and function. The dynamics of a protein is complex, with multiple relaxation times reflecting a hierarchical free-energy landscape. Using all-atom molecular dynamics simulations of an α/β protein (crambin) and a β-sheet polypeptide (BS2) in their “native” states, we demonstrate that the mean-square displacement of dihedral angles, defined by 4 successive C^α atoms, increases as a power law of time, t^α, with an exponent α between 0.08 and 0.39 (α = 1 corresponds to Brownian diffusion), at 300 K. Residues with low exponents are located mainly in well-defined secondary elements and adopt 1 conformational substate. Residues with high exponents are found in loops/turns and chain ends and exist in multiple conformational substates, i.e., they move on multiple-minima free-energy profiles.     

Differential gene expression of chemokines in KRAS and BRAF mutated colorectal cell lines:Role of cytokines

AIM:To study KRAS/BRAF mutations in colorectalcancer(CRC)that influences the efficacy of treatment.To develop strategies for overcoming combination of treatment.METHODS:Five colonic cell-lines were investigated:DLD-1 with KRAS(G13D)mutation,HT 29 and Colo205 with BRAF(V600E)mutation as well as the wild type(Wt)cell-lines Caco2 and Colo-320.DLD-1(KRAS),HT-29(BRAF)and Caco2(Wt)cell lines were treated with cytokines(TNFα50 ng,IL-1β1 ng and IFNγ50ng)and harvested at different time points(1-24 h).KRAS inhibition was performed by the siRNA-approach.Two colorectal cancer cells DLD-1 and Caco2 were used for KRAS inhibition.About 70%confluency were confirmed before transfection with small interferring RNA(siRNA)oligonucleotides.All the synthetic siRNA sequences were designed in our laboratory.Total RNA and protein was isolated from the cells for RT-PCR and Western blotting.Densitometry of the Western blotting was analyzed with the Image J software(NIH).Results are shown as mean±SD.RESULTS:RT-PCR analysis in non-stimulated cells showed a low basal expression of TNFαand IL-1βin the DLD-1 KRAS-mutated cell-line,compared to Caco2wild type.No detection was found for IL-6 and IFNγin any of the studied cell lines.In contrast,pro-angiogenic chemokines(CXCL1,CXCL8)showed a high constitutive expression in the mutated cell-lines DLD-1(KRAS),HT-29 and Colo205(BRAF),compared to wild type(Caco2).The anti-angiogenic chemokine(CXCL10)showed a high basal expression in wild-type,compared to mutated cell-lines.KRAS down-regulation by siRNA showed a significant decrease in CXCL1 and CXCL10gene expression in the DLD-1(KRAS)cell-line in comparison to wild type(Caco2)at 72 h after KRAS silencing.In contrast,the specific KRAS inhibition resulted in an up-regulation of CXCL1 and CXCL10.The results of our study show a higher expression of pro-angiogenic chemokines at basal level in mutated cell-lines,which was further increased by cytokine treatment.CONCLUSION:To summarize,basal chemokine gene expression for pro-angiogenic chemokines was high in mutated as compared to wild type cell-lines.This reflects the likely existence of a different microenvironment in tumours consistent of wild type or mutated cells.This may help to rationalize the choice of molecular targets for suitable therapeutic investigation in clinical studies.     

Elevated cysteine-rich protein 61 (CCN1) promotes skin aging via upregulation of IL-1β in chronically sun-exposed human skin

Chronic exposure of human skin to solar ultraviolet (UV) irradiation causes premature skin aging, which is characterized by reduced type I collagen production and increased fragmentation of the dermal collagenous extracellular matrix. This imbalance of collagen homeostasis is mediated, in part, by elevated expression of the matricellular protein cysteine-rich protein 61 (CCN1), in dermal fibroblasts, the primary collagen producing cell type in human skin. Here, we report that the actions of CCN1 are mediated by induction of interleukin 1β (IL-1β). CCN1 and IL-1β are strikingly induced by acute UV irradiation, and constitutively elevated in sun-exposed prematurely aged human skin. Elevated CCN1 rapidly induces IL-1β, inhibits type I collagen production, and upregulates matrix metalloproteinase-1, which degrades collagen fibrils. Blockade of IL-1β actions by IL-1 receptor antagonist largely prevents the deleterious effects of CCN1 on collagen homeostasis. Furthermore, knockdown of CCN1 significantly reduces induction of IL-1β by UV irradiation, and thereby partially prevents collagen loss. These data demonstrate that elevated CCN1promotes inflammaging and collagen loss via induction of IL-1β and thereby contributes to the pathophysiology of premature aging in chronically sun-exposed human skin.     

Lactobacilli, bifidobacteria and E. coli nissle induce pro- and anti-inflammatory cytokines in peripheral blood mononuclear cells

INTRODUCTION There is no doubt that the relationship between intestinal micro ora and immune system is complex. The host has to distinguish between pathogenic bacteria and harmless commensal and must react in an adequate and not self- destructive manner[1…     

Theta frequency stimulation up-regulates the synaptic strength of the pathway from CA1 to subiculum region of hippocampus

The subiculum (SB) is the principal target of the axons of the CA1 pyramidal cells and serves as the final relay in the trisynaptic loop between the entorhinal cortex and the hippocampus. We have examined synaptic plasticity in the synaptic pathway between the CA1 pyramidal cells and the SB in hippocampal slices and compared it under the same experimental condition with the synaptic plasticity in Shaffer collateral pathway (CA3-CA1). We find that the frequency response curve of synaptic strength induced by prolonged low-frequency stimulation (1-5 Hz) is systematically up-shifted from Shaffer collateral to the CA1-SB pathway. The up-regulation of synaptic strength is mediated by the activity-dependent modulation by beta-adrenergic transmission. Because the CA3-CA1 and the CA1-SB synaptic pathways are in series and the beta-adrenergic modulation is region-specific, this modulation seems to be involved in the selective control of signal transmission between the different regions of hippocampus.     

Antiangiogenic agents increase breast cancer stem cells via the generation of tumor hypoxia

Antiangiogenic therapy has been thought to hold significant potential for the treatment of cancer. However, the efficacy of such treatments, especially in breast cancer patients, has been called into question, as recent clinical trials reveal only limited effectiveness of antiangiogenic agents in prolonging patient survival. New research using preclinical models further suggests that antiangiogenic agents actually increase invasive and metastatic properties of breast cancer cells. We demonstrate that by generating intratumoral hypoxia in human breast cancer xenografts, the antiangiogenic agents sunitinib and bevacizumab increase the population of cancer stem cells. In vitro studies revealed that hypoxia-driven stem/progenitor cell enrichment is primarily mediated by hypoxia-inducible factor 1α. We further show that the Akt/β-catenin cancer stem cell regulatory pathway is activated in breast cancer cells under hypoxic conditions in vitro and in sunitinib-treated mouse xenografts. These studies demonstrate that hypoxia-driven cancer stem cell stimulation limits the effectiveness of antiangiogenic agents, and suggest that to improve patient outcome, these agents might have to be combined with cancer stem cell-targeting drugs.     

Inflammasome activation in decompensated liver cirrhosis

Inflammation participates in the pathogenesis ofmany liver diseases, including liver cirrhosis. Certain inflammatory citokines, such as interleukin(IL)-1β and IL-18, are produced after the activation of a multiprotein complex known as the inflammasome. Activation of the inflammasome has been documented in several liver diseases, but its role in the development and progression of liver cirrhosis or the complications associated with this disease is still largely unknown. We have recently studied the impact of the inflammasome in the sterile inflammatory response that takes place in the ascitic fluid of patients with decompensated cirrhosis, providing evidence that activation of the absent in melanoma 2(AIM2) inflammasome is an important response in these patients. Ascitic fluid-derived macrophages were able to mount a very robust AIM2-mediated response even in the absence of a priming signal, which is usually required for the full activation of all the inflammasomes. In addition, high level of inflammasome activation in these patients was associated with a higher degree of liver disease and an increased incidence of spontaneous bacterial peritonitis. These results may help explain the exacerbated inflammatory response that usually occurs in patients with decompensated cirrhosis in the absence of detectable infections. Thus, inflammasomes should be considered as possible therapeutic targets in sterile inflammatory complications in patients with cirrhosis.     

Caspase-1 inhibition alleviates acute renal injury in rats with severe acute pancreatitis

AIM:To assess the effect of inhibition of caspase-1 on acute renal injury in rats with severe acute pancreatitis(SAP).METHODS:Forty-two Sprague-Dawley rats were randomly divided into three groups:healthy controls(HC,n=6),SAP rats treated with saline(SAP-S,n=18),or SAP rats treated with a caspase-1/interleukin(IL)-1β-converting-enzyme(ICE)inhibitor(SAP-I-ICE,n=18).SAP was induced by retrograde infusion of 5%sodium taurocholate into the bile-pancreatic duct.HC rats were subjected to identical treatment and surgical procedures without sodium taurocholate.Rats received an intraperitoneal injection of isotonic saline(SAP-S)or the inhibitor(SAP-ICE-I)at 2 and 12 h after induction of acute pancreatitis.Surviving rats were sacrificed at different time points after SAP induction;all samples were obtained and stored for subsequent analyses.The levels of blood urea nitrogen(BUN)and creatinine(Cr)were measured using automatic methods,and serum IL-1βconcentrations were measured by an enzymelinked immunosorbent assay.Intrarenal expression of IL-1β,IL-18 and caspase-1 mRNAs was detected by RT-PCR.IL-1βprotein expression and the pathologic changes in kidney tissues were observed by microscopy after immunohistochemical or hematoxylin and eosin staining,respectively.RESULTS:The serum levels of BUN and Cr in the SAP-S group were 12.48±2.30 mmol/L and 82.83±13.89μmol/L at 6 h,23.53±2.58 mmol/L and 123.67±17.67μmol/L at 12 h,and 23.60±3.33 mmol/L and125.33±21.09μmol/L at 18 h,respectively.All were significantly increased compared to HC rats(P<0.01for all).Levels in SAP-ICE-I rats were significantly decreased compared to SAP-S rats both at 12 and 18 h(P<0.01 for all).Serum IL-1βlevels in the SAP-S group were 276.77±44.92 pg/mL at 6 h,308.99±34.95pg/mL at 12 h,and 311.60±46.51 pg/mL at 18 h;all significantly higher than those in the HC and SAP-ICE-I groups(P<0.01 for all).Intrarenal expression of IL-1βmRNA was weak in HC rats,but increased significantly in SAP-S rats(P<0.01).ICE inhibition significantly decreased the expression of IL-1βand IL-18 mRNAs(P<0.05 for all vs SAP-S),whereas caspase-1 mRNA expression was not significantly different.Weak IL-1βimmunostaining was observed in HC animals,and marked staining was found in the SAP-S group mainly in renal tubular epithelial cells.IL-1βimmunostaining was significantly descended in SAP-ICE-I rats compared to SAP-S rats(P<0.05).Caspase-1 inhibition had no effect on the severity of kidney tissue destruction.CONCLUSION:The expression of caspase-1-activated cytokines IL-1βand IL-18 plays a pivotal role in acute renal injury in rats with experimental SAP.Caspase-1inhibition improves renal function effectively.     

microRNA 17/20 inhibits cellular invasion and tumor metastasis in breast cancer by heterotypic signaling

microRNAs are thought to regulate tumor progression and invasion via direct interaction with target genes within cells. Here the microRNA17/20 cluster is shown to govern cellular migration and invasion of nearby cells via heterotypic secreted signals. microRNA17/20 abundance is reduced in highly invasive breast cancer cell lines and node-positive breast cancer specimens. Cell-conditioned medium from microRNA17/20–overexpressing noninvasive breast cancer cell MCF7 was sufficient to inhibit MDA-MB-231 cell migration and invasion through inhibiting secretion of a subset of cytokines, and suppressing plasminogen activation via inhibition of the secreted plasminogen activators (cytokeratin 8 and α-enolase). microRNA17/20 directly repressed IL-8 by targeting its 3′ UTR, and inhibited cytokeratin 8 via the cell cycle control protein cyclin D1. At variance with prior studies, these results demonstrated a unique mechanism of how the altered microRNA17/20 expression regulates cellular secretion and tumor microenvironment to control migration and invasion of neighboring cells in breast cancer. These findings not only reveal an antiinvasive function of miR-17/20 in breast cancer, but also identify a heterotypic secreted signal that mediates the microRNA regulation of tumor metastasis.     

Pin1 overexpression in colorectal cancer and its correlation with aberrant β-catenin expression

AIM: To investigate clinical significance of Pin1 and beta-catenin expression in colorectal cancers and to demonstrate the relationship of their expression. METHODS: The role of Pin1 and beta-catenin protein in colorectal tumorigenesis and their clinicopathologic significance were analyzed by immunohistochemistry, and the correlation between Pin1 and beta-catenin protein expressions was also studied in 124 patients with colorectal cancer who were surgically treated. RESULTS: Normal colonic epithelium either failed to express or showed focal and weak expression of Pin1 and beta-catenin. Overexpression of Pin1 and beta-catenin protein was found in 23 (18.54%) and 50 (40.3%) of 124 colorectal cancers, respectively. Overexpression of both proteins was not related to the lymph node metastasis, tumor stage and survival period after excision. Survival analysis results indicated that tumor stage was a valuable predictor of survival. Interestingly, a significant correlation was found between Pin1 and beta-catenin protein expression. CONCLUSION: Overexpression of Pin1 and beta-catenin may be closely related with the development and/or progression of colorectal carcinoma and further supports that Pin1 overexpression might contribute to the upregulation of beta-catenin.     

Helicobacter pylori tumor necrosis factor-α inducing protein promotes cytokine expression via nuclear factor-κB

AIM:To study the effects of Helicobacter pylori(H. pylori)tumor necrosis factor-α(TNF)inducing protein (Tip-α)on cytokine expression and its mechanism. METHODS:We cloned Tip-αfrom the H.pylori strain 26695,transformed Escherichia coli with an expression plasmid,and then confirmed the expression product by Western blotting.Using different concentrations of Tip-αthat affected SGC7901 and GES-1 cells at different times,we assessed cytokine levels using enzyme-linked immunosorbent assay.We blocked SGC7901 cells with pyrrolidine dithiocarbamate(PDTC),a specific inhibitor of nuclear factorκB(NF-κB).We then detected interleukin(IL)-1βand TNF-αlevels in SGC7901 cells. RESULTS:Western blot analysis using an anti-Tip-α antibody revealed a 23-kDa protein,which indicated that recombinant Tip-αprotein was recombined successfully.The levels of IL-1β,IL-8 and TNF-αwere sig-nificantly higher following Tip-αinterference,whether GES-1 cells or SGC-7901 cells were used(P<0.05).However,the levels of cytokines(including IL-1β,IL-8 and TNF-α)secreted by SGC-7901 cells were greater than those secreted by GES-1 cells following treatment with Tip-αat the same concentration and for the same duration(P<0.05).After blocking NF-κB with PDTC, the cells(GES-1 cells and SGC-7901 cells)underwent interference with Tip-α.We found that IL-1βand TNF-αlevels were significantly decreased compared to cells that only underwent Tip-αinterference(P<0.05). CONCLUSION:Tip-αplays an important role in cyto-kine expression through NF-κB.     

Interplay among side chain sequence, backbone composition, and residue rigidification in polypeptide folding and assembly

The extent to which polypeptide conformation depends on side-chain composition and sequence has been widely studied, but less is known about the importance of maintaining an alpha-amino acid backbone. Here, we examine a series of peptides with backbones that feature different repeating patterns of alpha- and beta-amino acid residues but an invariant side-chain sequence. In the pure alpha-backbone, this sequence corresponds to the previously studied peptide GCN4-pLI, which forms a very stable four-helix bundle quaternary structure. Physical characterization in solution and crystallographic structure determination show that a variety of alpha/beta-peptide backbones can adopt sequence-encoded quaternary structures similar to that of the alpha prototype. There is a loss in helix bundle stability upon beta-residue incorporation; however, stability of the quaternary structure is not a simple function of beta-residue content. We find that cyclically constrained beta-amino acid residues can stabilize the folds of alpha/beta-peptide GCN4-pLI analogues and restore quaternary structure formation to backbones that are predominantly unfolded in the absence of cyclic residues. Our results show a surprising degree of plasticity in terms of the backbone compositions that can manifest the structural information encoded in a sequence of amino acid side chains. These findings offer a framework for the design of nonnatural oligomers that mimic the structural and functional properties of proteins.     

A unique mechanism of beta-blocker action: carvedilol stimulates beta-arrestin signaling

For many years, beta-adrenergic receptor antagonists (beta-blockers or betaAR antagonists) have provided significant morbidity and mortality benefits in patients who have sustained acute myocardial infarction. More recently, beta-adrenergic receptor antagonists have been found to provide survival benefits in patients suffering from heart failure, although the efficacy of different beta-blockers varies widely in this condition. One drug, carvedilol, a nonsubtype-selective betaAR antagonist, has proven particularly effective in the treatment of heart failure, although the mechanism(s) responsible for this are controversial. Here, we report that among 16 clinically relevant betaAR antagonists, carvedilol displays a unique profile of in vitro signaling characteristics. We observed that in beta2 adrenergic receptor (beta2AR)-expressing HEK-293 cells, carvedilol has inverse efficacy for stimulating G(s)-dependent adenylyl cyclase but, nonetheless, stimulates (i) phosphorylation of the receptor's cytoplasmic tail on previously documented G protein-coupled receptor kinase sites; (ii) recruitment of beta-arrestin to the beta2AR; (iii) receptor internalization; and (iv) activation of extracellular regulated kinase 1/2 (ERK 1/2), which is maintained in the G protein-uncoupled mutant beta2AR(T68F,Y132G,Y219A) (beta2AR(TYY)) and abolished by beta-arrestin2 siRNA. Taken together, these data indicate that carvedilol is able to stabilize a receptor conformation which, although uncoupled from G(s), is nonetheless able to stimulate beta-arrestin-mediated signaling. We hypothesize that such signaling may contribute to the special efficacy of carvedilol in the treatment of heart failure and may serve as a prototype for a new generation of therapeutic beta2AR ligands.     

Liver receptor homolog 1 contributes to intestinal tumor formation through effects on cell cycle and inflammation

Liver receptor homolog 1 (LRH-1) is an orphan nuclear receptor that synergizes with beta-catenin/T cell factor 4 signaling to stimulate intestinal crypt cell renewal. We evaluated here the impact of haploinsufficiency of LRH-1 on intestinal tumorigenesis by using two independent mouse models of human colon tumorigenesis. Haploinsufficiency of LRH-1 blunts intestinal tumorigenesis in the ApcMin/+ mice, a genetic model of intestinal cancer. Likewise, Lrh-1+/- mice are protected against the formation of aberrant crypt foci in the colon of mice exposed to the carcinogen azoxymethane. LRH-1 gene expression is reduced in tumors that express elevated levels of the proinflammatory cytokine TNF-alpha. Reciprocally, decreased LRH-1 expression in Lrh-1+/- mice attenuates TNF-alpha expression. Compared with normal human colon, expression and subcellular localization of LRH-1 is significantly altered in neoplastic colon. In combination, these data suggest a role of LRH-1 in the initiation of intestinal tumorigenesis both by affecting cell cycle control as well as through its impact on inflammatory pathways.     

Sca-1 expression identifies stem cells in the proximal region of prostatic ducts with high capacity to reconstitute prostatic tissue

We previously showed that prostatic stem cells are concentrated in the proximal regions of prostatic ducts. We now report that these stem cells can be purified from isolated proximal duct regions by virtue of their high expression of the cell surface protein stem cell antigen 1 (Sca-1). In an in vivo prostate reconstitution assay, the purified Sca-1-expressing cell population isolated from the proximal region of ducts was more effective in generating prostatic tissue than a comparable population of Sca-1-depleted cells (203.0 +/- 83.1 mg vs. 11.9 +/- 9.2 mg) or a population of Sca-1-expressing cells isolated from the remaining regions of ducts (transit-amplifying cells) (31.9 +/- 24.1 mg). Almost all of the proliferative capacity of the proximal duct Sca-1-expressing cell population resides within the fraction of cells that express high levels of Sca-1 (top one-third), with the proximal region of prostatic ducts containing 7.2-fold more Sca-1(high) cells than the remaining regions. More than 60% of the high-expressing cells coexpress alpha6 integrin and the anti-apoptotic factor Bcl-2, markers that are also characteristic of stem cells of other origins. Further stratification of the phenotype of the stem cells may enable the development of rational therapies for treating prostate cancer and benign prostatic hyperplasia.