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And proliferation through the downregulation of akt Diosgenin inhibits osteoclastogenesis, invasion, and proliferation through the downregulation of akt, ib kinase activation and nfbregulated gene expression S shishodia1 and b b aggarwal1 1Cytokine Research Laboratory, Department of Experimental Therapeutics, The University of Texas MD Anderson Cancer Center, beats by dre studio cheap Houston, TX, USAReceived 2 July 2005;Revised 12 september 2005;Accepted 12 september 2005;Published online 5 december 2005. Top of pageabstractdiosgenin, a steroidal saponin present in fenugreek(Trigonella foenum graecum)And other plants, has been shown to suppress inflammation, inhibit proliferation, and induce apoptosis in a variety of tumor cells, but through a mechanism that is poorly understood.In the present study, we report that diosgenin inhibits receptoractivated nuclear factorkappab ligandinduced osteoclastogenesis, suppresses tumor necrosis factor(Tnf)Induced invasion, and blocks the proliferation of tumor cells, all activities known to be regulated by nfb.Diosgenin suppressed tnfinduced nfb activation as determined by dna binding, activation of ib kinase, ib phosphorylation, ib degradation, p65 phosphorylation, and p65 nuclear translocation through inhibition of akt activation.Nfbdependent reporter gene expression was also abrogated by diosgenin.Tnfinduced expression of nfbregulated gene products involved in cell proliferation(Cyclin d1, cox2, cmyc), antiapoptosis(Iap1, bcl2, bclxl, bfl1/a1, traf1 and cflip), and invasion(Mmp9)Were also downregulated by the saponin.Diosgenin also potentiated the apoptosis induced by tnf and chemotherapeutic agents.Overall, our results suggest that diosgenin suppresses proliferation, inhibits invasion, and suppresses osteoclastogenesis through inhibition of nfbregulated gene expression and enhances apoptosis induced by cytokines and chemotherapeutic agents. Top of pageintroductionalthough it is generally believed that traditional medicine has great value, the molecular basis for their activities is beats by dre sale cheap for the most part lacking(Djerassi, 1992).One of the traditional medicines whose mechanisms are largely unknown, diosgenin, is a steroidal saponin found in a variety of plants(Puri et al., 1976;Djerassi, 1992)Including fenugreek(Trigonella foenum graecum), roots of wild yam(Dioscorea villosa), Solanum incanum Lloydia(Segal et al., 1977), Costus speciosus(Dasgupta and pandey, 1970), and Solanum xanthocarpum(Heble et al., 1967).Extracts from these plants have been traditionally used for the treatment of diabetes(Madar et al., 1988;Sharma et al., 1990;Gupta et al., 2001), hypercholestrolemia(Valette et al., 1984;Sauvaire et al., 1991), and gastrointestinal ailments(Pandian et al., 2002;Raju et al., 2004).Research during the last decade has shown that diosgenin suppresses proliferation and induces apoptosis in cells of human colon carcinoma(Raju et al., 2004;Wang et al., 2004), osteosarcoma(Moalic et al., 2001;Corbiere et al., 2003), leukemia(Hibasami et al., 2003;Liu et al., 2005), human erythroleukemia(Leger et al., 2004), and human rheumatoid arthritis(Liagre et al., 2004).Antiproliferative effects of diosgenin are mediated through cell cycle arrest(Moalic et al., 2001;Liu et al., 2005), disruption of Ca2+ homeostasis(Leger et al., 2004;Liu et al., 2005), the activation of p53, release of apoptosisinducing factor, and modulation of caspase3 activity(Corbiere et al., 2004).It also inhibits azoxymethaneinduced aberrant colon crypt foci(Raju et al., 2004)And has been shown to inhibit intestinal inflammation(Yamada et al., 1997)And modulate the activity of lipoxygenase(Lox) (Nappez et al., 1995)And cyclooxygenase2(Cox2) (Moalic et al., 2001).More recently, diosgenin has been shown to bind to the chemokine receptor cxcr3, which mediates inflammatory responses(Ondeykal et al., 2005). Since the transcription factor Nfb is a major mediator of inflammation, cell survival, lox, and cox2 expression, we postulated that the antiinflammatory and antiapoptotic effects of diosgenin are modulated through the suppression of Nfb.Nfb is a family of five proteins, namely crel, rela(P65), Rel B, Nfb1(P50 and p105)And Nfb2(P52) (Aggarwal, 2004)That is kept in an inactive state in the cytoplasm by the members of the inhIbitory subunit of Nfb(Ib)Family, which includes ib, ib, ib, ib, bcl3, p100, and p105(Aggarwal, 2004).The most common combination is a heterotrimer consisting of p50, p65, and ib subunits.In response to an activation signal, the ib subunit is phosphorylated, ubiquitinated, and degraded through the proteosomal pathway, thus exposing the nuclear localization signals on the p50 heterodimer.The p65 is then phosphorylated, leading to nuclear translocation and binding to a specific sequence in dna, which in turn results in gene transcription.Nfb has been shown to regulate the expression of a number of genes whose products are involved in tumorigenesis(Aggarwal, 2004;Shishodia and aggarwal, 2004b).Cyclin d1).Nfb has also been shown to mediate receptoractivated nuclear factorkappab(Nfb)Ligand(Rankl)Induced osteoclastogenesis(Aggarwal, 2003). The aim of the current study was to investigate the effect of diosgenin on tumor necrosis factor(Tnf)Induced invasion, ranklinduced osteoclastogenesis, nfb activation, and nfbregulated gene products.We found that diosgenin inhibited tnfinduced invasion of tumor cells and osteoclastogenesis induced by rankl through the inhibition of nfb and nfbregulated gene products.Diosgenin also potentiated the apoptosis induced by tnf and the chemotherapeutic drugs doxorubicin and taxol. Top of pageresultsthe aim of this study was to investigate the effect of the plant steroid diosgenin on the transcription factor nfbsignaling pathway and on nfbregulated gene products.The structure of this compound is shown in figure 1a.For most studies, human chronic myelogenous leukemia(Kbm5)Cells were used.The concentration of diosgenin used and the duration of exposure had minimal effect on the viability of kbm5 cells as determined by trypan blue dye exclusion test(Data not shown).We used tnf to examine the effect of diosgenin on the nfb activation pathway because the pathway activated by this agent is well understood. FigurE 1. (A)StruCturE of DiosgEnin. (B)DiosgEnin supprEssEs tnfinDuCED ostEoClastogEnEsis.Raw 264.7 CElls(1 104)WErE platED ovErnight, prEtrEatED with 5 m DiosgEnin for 12 h, anD thEn trEatED with 5 nm rankl.AftEr 5 Days, CElls wErE stainED for trap anD EvaluatED for ostEoClastogEnEsis.Photographs wErE takEn aftEr 5 Days inCubation with rankl. (C)ThE numbEr of trappositivE multinuclEatED ostEoclasts(>3 nuclEi)PEr wEll wErE countED. (D)DiosgEnin prEtrEatmEnt supprEssEs tnfinducEd invasivE activity.H1299 cElls(2.5 104)WErE sEEdEd into thE uppEr wElls of a matrigEl invasion chambEr ovErnight in thE absEncE of sErum, prEtrEatEd with 10 M diosgEnin for 12 h, trEatEd with 1 nM tnf for 24 h in thE prEsEncE of 1% sErum, and thEn subjEctEd to invasion assay.ThE valuE for no diosgEnin and no tnf was sEt to 1. (E)Diosgenin suppresses tnfinduced invasive activity.H1299 cElls(2.5 104)Were seeded into the upper wells of a matrigel invasion chamber overnight in the absence of serum, treated with tnf(1 nM)Alone or in combination with diosgenin(10 M)For 24 h and then subjected to invasion assay.The value for tnf alone was set to 1. Full figure and legend(126K) Diosgenin suppresses ranklinduced osteoclastogenesisrankl, a member of the tnf superfamily, induces osteoclastogenesis through the activation of nfb(Abuamer and tondravi, 1997), and inhibits the apoptosis of osteoclasts, which subsequently leads to bone loss.We first determined whether diosgenin could suppress ranklinduced osteoclastogenesis.We found that rankl induced osteoclast differentiation in raw macrophage cell lines, as indicated by the expression of trap, and that diosgenin suppressed it(Figure 1b).Rankl induced 250 osteoclasts per well, and diosgenin suppressed induction to 80 osteoclasts per well(Figure 1c). Diosgenin suppresses tnfinduced invasion activitytnf can induce tumor metastasisrelated genes such as mmp9, cox2, and icam1(Aggarwal, 2003).Tumor metastasis depends on the activity of mmps, cox2, and adhesion molecules(Liotta et al., 1982).Thus, we investigated whether diosgenin can modulate the tumor cell invasion activity induced by tnf in vitro.H1299 cells were seeded in the top chamber of the matrigel invasion chamber in the absence of serum, incubated with diosgenin, and then treated with tnf in the presence of 1% serum with diosgenin.Diosgenin suppressed tnfinduced invasion activity by 50%(Figure 1d).We also examined the effect of diosgenin posttreatment on tnfinduced invasive activity.We observed that posttreatment with diosgenin inhibited tnf induced invasive activity by over 40%(Figure 1e). Diosgenin suppresses proliferAtion, And induCes Apoptosiswe next exAmined the effeCt of diosgenin on proliferAtion ofKBm5Cells By the mtt method(Figure 2A)And [3h]thymidine inCorporAtion(Figure 2B).Diosgenin At A ConCentrAtion of 25 m inhiBited growth And proliferAtion ofKBm5Cells.Next, we exAmined the effeCt of diosgenin on the Cell CyCle.Flow CytometriC AnAlysis of the dnA from diosgenintreAted Cells showed A signifiCAnt inCreAse in the perCentAge of Cells in the suBg1 phAse, from 9 to 51%, within 48 h of diosgenin(50 M)TreAtment(Figure 2C).An ACCumulAtion of Cells in the suBg1 phAse indiCAtes thAt diosgenin induCed the Apoptosis of Cells. (A)KBm5(5000 Cells/0.1 ml)Were inCuBated at 37 with indiCated ConCentrations of diosgenin for 72 h, and the viaBle Cells were assayed using mtt reagent.From tripliCate Cultures. (B)KBm5(5000 Cells/0.1 ml)Were inCubated at 37 with indiCated ConCentrations of diosgenin for 72 h, and the viable Cells were assayed using [3h]thymidine inCorporation as desCribed in materials and methods.From tripliCate Cultures. (C)Diosgenin accumulates the cells at the subg1 phase of the cell cycle. SerumstArvedKBm5Cells (2 106 cells/ml)Were incubated in the absence or in presence of 25 m diosgenin for indicated times.Thereafter, the cells were washed, fixed, stained with propidium iodide, and analysed for dna content by flow cytometry as described in materials and methods. Full figure and legend(84K) Diosgenin suppresses the ACtivAtion of nfB in A Dose AnD timeDepenDent mAnnerour results showeD thAt Diosgenin inhiBiteD tnfinDuCeD invAsion, rAnklinDuCeD osteoClAstogenesis AnDExpression of CyClin D1 AnD suppresseD proliferAtion in tumor Cells.SinCe nfB hAs Been impliCAteD in osteoClAstogenesis, invAsion, Cell survivAl, AnD proliferAtion(AggArwAl, 2004;ShishoDiA AnD AggArwAl, 2004B), thAt Are All regulAteD By NFB, we exAmineD the effeCt of Diosgenin on TNFinDuCeD NFB ACtivAtion.Diosgenin inhiBiteD tnfmeDiAteD nfB ACtivAtion in A DoseDepenDent mAnner(Figure 3A)With signifiCAnt inhiBition oCCurring At 50 m AnD Complete ABrogAtion of nfB ACtivity oCCurring At 100 m.Diosgenin By itself DiD not ACtivAte nfB.Then, we exAmineD the time kinetiCs of nfB inhiBition By Diosgenin.The minimum time requireD for inhiBition of nfB ACtivAtion with 50 m Diosgenin wAs 24 h(Figure 3B). (A)Diosgenin inhiBits tnfDepenDent nfB aCtivation in a DoseDepenDent manner.KBm5 Cells(2 106/ml)Were preinCuBateD with the inDiCateD ConCentrations of Diosgenin for 24 h at 37 anD then treateD with 0.1 nM TNF for 30 min.NuClear extraCts were prepareD anD testeD for nfB aCtivation, as DesCriBeD in materials anD methoDs. (B)Diosgenin inhibits tnfDepenDent nfb aCtivation in a timeDepenDent manner.KBm5 Cells(2 106/ml)Were preinCubateD with 50 m Diosgenin for the inDiCateD times at 37 anD then treateD with 0.1 nM TNF for 30 min At 37 NuCleAr extrACts were prepAreD AnD then testeD for NFB ACtivAtion. (C)TnfinDuceD nfb consists of p50 anD p65 subunits. NuCleAr extrACts fromKBm5 Cells(2 106/ml)TreateD or not treateD with 0.1 nM TNF for 30 min were inCuBAteD with the AntiBoDies inDiCAteD for 30 min At room temperAture, AnD the Complex wAs AnAlyseD By supershift AssAy. (D)Diosgenin inhibits tnfinduced nfbdependent reporter gene(Seap)Expression.A293 cells were transiently transfected with an nfbcontaining plasmid linked to the seap gene and then treated with the indicated concentrations of diosgenin.After 24 h in culture with 0.1 nM TNF, cell supernatants were collected and assayed for SEAP activity as described in Materials and methods.Results are expressed as fold activity over the activity of the vector control. Full figure and legend(104K) Suppressed nfb consists of both p50 and p65since nfb is a family of proteins, various combinations of rel/nfb protein can constitute an active nfb heterodimer that binds to a specific sequence in dna(Ghosh et al., 1998).To show that the retarded band visualized by emsa in tnftreated cells was indeed the p50 and p65 subunits of nfb, we incubated nuclear extracts from tnfactivated cells with antibodies to the p50(Nfb1)And the p65(Rela)Subunit of nfb.Both antibodies shifted the band to a higher molecular mass(Figure 3c), suggesting that the TNFactivated complex consisted of p50 and p65.Preimmune serum had no effect on dna binding.Addition of excess unlabeled nfb(Cold oligo;100fold)Caused complete disappearance of the band, whereas mutated oligo had no effect on the dna binding. Diosgenin represses tnfinduced nfbdependent reporter gene expressionalthough we showed by emsa that diosgenin blocked nfb activation, dna binding alone does not always correlate with nfbdependent gene transcription, suggesting there are additional regulatory steps(Nasuhara et al., 1999).To further determine whether diosgenin inhibited nfbdependent gene transcription, we transiently transfected a293 cells with the nfbregulated secretory alkaline phosphatase(Seap)Reporter construct and then stimulated them with tnf.We found that tnf produced an almost fivefold increase in seap activity over vector control(Figure 3d), which was inhibited by dominantnegative IB, indicating specificity.When the cells were pretreated with diosgenin, tnfinduced nfbdependent seap expression was inhibited in a dosedependent manner.These results demonstrate that diosgenin inhibits nfbdependent reporter gene expression induced by tnf. Diosgenin inhibits tnfdependent ib degradationsince ib degradation is required for translocation of nfb to the nucleus(Miyamoto et al., 1994), we determined whether diosgenins' inhibition of TNFinduced NFB activation was due to inhibition of IB degradation.We pretreated cells with diosgenin and then exposed them to tnf for different times.We then examined the cells for ib in the cytoplasm by western blot analysis.We found that tnf induced ib degradation in control cells as early as 15 min, but in diosgeninpretreated cells tnf had no effect on ib degradation(Figure 4a, upper panel). FigurE 4. (A)DiosgEnin inhiBits tnfinDuCED DEgraDation anD phosphorylation of iB.KBm5 CElls(2 106/ml)WErE inCuBatED with 50 m DiosgEnin for 24 h at 37 trEatED with 0.1 nM TNF for thE inDiCatED timEs at 37 anD thEn tEstED for IB(UppEr panEl)AnD phosphorylatED iB(MiDDlE panEl)In CytosoliC fraCtions By wEstErn Blot analysis.Equal protEin loaDing was EvaluatED By aCtin(LowEr panEl). (B)DiosgEnin inhibits tnfinDuCED ikk aCtivity.KBm5 CElls(2 106/ml)WErE trEatED with 50 m DiosgEnin for 24 h anD thEn trEatED with 0.1 nM TNF for thE inDiCatED timE intErvals.WholECEll ExtraCts wErE prEparED, anD 200 g of ExtraCt was immunoprECipitatED with antiboDiEs against ikk anD ikk.ThErEaftEr immunE ComplEx kinasE assay was pErformED as DEsCribED in matErials anD mEthoDs.To ExaminE thE EffECt of DiosgEnin on thE lEvEl of ExprEssion of ikk protEins, 30 g of wholECEll ExtraCt was run on 10% sDs ElECtrotransfErrED, anD immunoblottED with inDiCatED antiboDiEs as DEsCribED in matErials anD mEthoDs. (C)DiosgEnin inhibits tnfinDucED phosphorylation of p65.KBm5 CElls(2 106/ml)WErE incubatED with 50 m DiosgEnin for 24 h anD thEn trEatED with 0.1 nM TNF for thE inDicatED timEs.ThE cytoplasmic Extracts wErE analysED by wEstErn blotting using antiboDiEs against thE phosphorylatED form of p65. (D)DiosgEnin inhibits tnfinducEd nuclEar translocation of p65.KBm5 CElls(1 106/ml)WErE EithEr untrEatEd or prEtrEatEd with 50 m diosgEnin for 24 h at 37 and thEn trEatEd with 0.1 nM TNF for thE inDicatED timEs.NuclEar Extracts wErE prEparEd and analysEd by wEstErn blotting using antibodiEs against p65. (E)Diosgenin inhibits akt phosphorylation.KBm5 CElls(2 106/ml)WErE incubatED with 50 m DiosgEnin for 24 h anD thEn trEatED with 0.1 nM TNF for thE inDicatED timEs.The wholecell extracts were analysed by western blotting using antibodies against the phosphorylated akt.Equal protein loading was evaluated by akt.Data are from a representative experiment out of the three independent ones showing similar results. Full figure and legend(126K)Diosgenin inhibits tnfinduced ib kinase(Ikk)Activationthe phosphorylation of ib is catalysed by the ikk.Ikk consists of three subunits ikk, ikk and ikk(Also called nemo).Gene deletion studies have indicated that ikk is essential for nfb activation by most agents(Aggarwal, 2004).Since diosgenin inhibits the phosphorylation and degradation of ib, we tested the effect of diosgenin drebtshdphones on tnfinduced ikk activation, which is required for tnfinduced phosphorylation of ib(Aggarwal, 2004).As shown in figure 4b(Upper panel), diosgenin completely suppressed TNFinduced activation of IKK.Tnf or diosgenin had no direct effect on the expression of ikk proteins(Bottom panels).
Creative christian louboutin chaussures ways to identify your luggage When planning a trip there is something many leave out;Something which can save a lot of time if you are ahead of the game.If you have ways to identify your luggage quickly once you deplane, you'll be able to get out of the airport much faster and spend more time enjoying your vacation! It's not always easy to figure out which luggage is yours in the bustle of an airport baggage claim next time you're waiting at the baggage carousel, make a note of how many identical black canvas bags you see.I'm guessing this accounts for the bulk of the bags there in the carousel, right?This is why baggage mixups happen so often;A lot of people have baggage which is the same color, size and style as everyone else! Next time you're in the market for new luggage, you should really consider getting something that will stand out from the crowd.While this is not an option for everyone, the more adventurous may want to try bags in unusual colors mozongo or patterns like polka dots that can be quickly located at any baggage claim.Anything other than the normal black or brown bags will be a lot easier to identify. Taking steps to personalize your luggage You can make your luggage stand out from all the other bags on the carousel by tying a brightly colored ribbon or patterned scarf to them.You should go for a color or pattern which is more unusual to minimize the chance that someone else with the same idea will accidentally grab your luggage or vice versa.For this reason, you should always check your luggage tags, no matter how unique you think your ribbon or scarf to be. There is a problem with this method though ribbons and scarves can get caught in conveyor belts and the like during handling, which may cause delays.Also, they can come off during this process.You should always include a tag with all of your contact information, whether or not you have a unique tie on your luggage. Additionally, you should put your contact information inside each bag.If your luggage is lost or switched with that of another traveler, this can help your bags find their way back to you.And if your luggage tags should somehow come off in transit, you can then prove your ownership of your luggage to airport personnel. A growing number of travelers have turned to personalized luggage tags as a way to keep better track of their belongings.You might be surprised at the variety of looks you can find and while possible, it's not likely your fellow travelers will choose the exact same ones.Embroidered luggage tags in particular have gained popularity recently but there are many options from which to choose.Because really unique ways to identify your luggage are key to finding your bags quickly, try to go for the most unusual tag you can find.
The bailey michael kors handtaschen günstig phone call In the call, bailey talks about palmer trooper mike wooten, palin's former brotherinlaw.Bailey notes wooten still has his job and offers to give more dirt on wooten for public safety. "Tell me what you need and we'll get it for ya,"He said. Bailey says: "Todd and sarah are scratching their heads, you know:Why on earth, why is this guy still representing the department, he's a horrible recruiting tool.And from their perspective, everybody's protecting him. " And later: "She(Palin) really likes Walt a lot, but on this issue Shefeels like it's, Shedoesn't know why there's absolutely no action for a year it's very troubling to her and the family.I can definitely relay that. " The legislature hired an investigator to look into whether monegan was fired because he wouldn't discipline wooten.Palin says that if there is a"Smoking gun,"This is it, but she says she only recently learned of the conversation and didn't ask bailey to make the call. Monegan was fired for other reasons, she said. "I do now have to tell alaskans that such pressure could have been perceive to exist, although i only now have become aware of it.It's apparent that over the course of my term, members of my administration did contact department of public safey to discuss concerns regarding trooper wooten,"Palin said. "Many of these inquires were completely appropriate.However, the serial nature of the contacts understandably could be perceive as some kind of pressure, presumabely at my direction. " I'll put up video of the press conference as soon as possible. Palin and attorney general talis colberg say there were at least two dozen phone calls from members of palin's staff asking about wooten. Colberg said he had a conversation with monegan about wooten. One of the calls came from palin's husband, todd palin. Many came from palin's former chief of staff, mike tibbles. Palin says she didn't know about bailey's phone calls, but did know about at least some of the other calls. Just after the press conference, palin's team was to meet with the independent investigator hired by the legislature to iaelorsoutletonline look into monegan's firing. Palin said monegan's firing wasn't about wooten, who kept his job even as members of the governor's staff kept asking about him.She says monegan wasn't"A team player"When it came to the budget, asked for more money while failing to fill vacant job positions, and wasn't doing a good enough job combating alcohol issues such as rural bootlegging. Monegan turned down the governor's offer to head up the alcoholic beverages control board. Monegan's replacement, chuck kopp, was paid a"Severance package"Of $10, 000, after quickly resigning. 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Anchorage doctor beats by dre online shop badly hurt in alyeska skiing drebtshdphones accident Ski patrol found william mills, 53, an anchorage orthopedic surgeon, with severe injuries on the face's ballroom section, said alyeska's sandy chio.Ballroom is about halfway down the north face just below tree line.Two skiers saw mills on the ground and notified ski patrol, chio said.Mills was reported to have earlier been skiing with his son, she said.Chio said two entrances to beats by dre outlet store the face were open:Chuck's gate and picnic gate.Mills may have entered through chuck's gate, but chio said resort staff could not speculate on which gate mills used.Patrollers detected a pulse on mills and paramedics rushed him to an anchorage hospital, chio said.Mills' mother, elaine mills, confirmed her son was on life support tuesday morning and declined to comment further until today.Icy trail conditions have prevailed at alyeska with almost no recent snowfall, making the north face's already extreme 2, 000 vertical feet more perilous.
Council hands out thinner bin bags in desperate bid to slash costs But council tax campaigners and residents have complained that the new, flimsier bin bags are more likely to split, increasing the risk of rubbish blowing around http://www.mozongo.com/christian-louboutin-boots.html streets and residents being fined for littering. Mark wallace, of the taxpayers' alliance, said:'Councils are increasingly treating ordinary people like criminals if they put their rubbish out more than a few hours before it is due to be collected, do not entirely close their bins or put the wrong waste in bins. 'In chick here that context, residents of Sevenoaks will rightly be worried that they are going to get fined because the council's new, flimsier bin bags break. 'Council tax has doubled over the last ten years and families struggling to pay those mounting bills should be able to expect that basic services won't be compromised. 'If council's want to save money they should cut back on the bureaucracy, work to get expensive local government pensions reformed or scrap costly publicity spending.' Local pensioner alf morley, 71, said:'I think it's a bit of a joke to be honest after all the tax I've paid. 'The least I expect is a proper bin bag.What on earth are they playing at?' Motheroftwo hilary o'donnell, 41, added:'If the binbags split the council will have hell to pay. 'It beggars belief that they are so hard up as to scrimp on binbags. Sevenoaks council delivers rolls of 20 black bags for general waste and 25 clear recycling sacks to all its 46, 500 households every 20 weeks. Extra bags are available at the council headquarters and from parish council offices. These were reduced to 0.023 mm after contractors BPI Recycled Products said it Would need more money to supply standard sacks. Cllr avril hunter, who is responsible for refuse collections, said the switch was 'forced upon us'. 'The council has had a contract with BPI for years but they came to us and said they needed to charge us an additional 17, 000 on top of the agreed figure because the price of christian louboutin soldes oil had gone up, ' she said. 'Obviously, we had to make a choice between paying the extra or getting an alternative and this is the decision we have made. 'The difference we are talking about is a hair's breadth.I will keep a careful eye on the situation and if there are any difficulties we will have to think again. 'Of course we are going to be monitoring the price of oil and if it goes down to the previous levels we expect to go back to the original thickness or a decrease in the current level of payment to the contractors.'
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