In agreement with these scientific studies, the lactational and adolescent rats on this examine have an irreversible impairment in hippocampal ERK1 2 and CREB following developmental publicity to ID and hypothyroidism. This really is mainly because that, in the course of these devel opmental intervals, the mom is definitely the only supply of iodine for the fetus and neonate. Thus, maternal ID leads to maternal and filial hypothyroxinemia. Taking together, developmental intervals through pregnancy and lactation are critical during the brain advancement. Thyroid hormone is well known to regulate morphologi cal and biochemical alterations in brain all through significant intervals of brain improvement. This regulation is usually carried out by genomic, posttranslational, and direct actions on neural substrates. For that reason, thyroid hormone is usually a funda mental component that regulates standard advancement of the CNS.
Logically, thyroid hormone insufficiency all through early brain growth is detrimental to synaptic trans mission in hippocampus and also to a cellular and you can look here behavioral model of learning and memory. In need, our pre vious research has plainly demonstrated that ID exposure lowers the youngsters IQs in China. Experimentally, we’ve also proven the ID can cause an inhibition of LTP during the rat CA1 region. It really is popular that MAPK mediates neuronal metabo lism, differentiation and proliferation. Also, the persisting alteration in developmental brain consists of alterations in myelination and migration patterns of neu rons. The present examine even further confirmed the hypothesis that developmental ID and hypothyroidism induce irreversible impairment of two important proteins expression while in the CA1 and CA3 areas, exactly where the two ERK1 2 and CREB expression was significantly reduced in either complete protein degree or phorsphorylated fraction.
In agreement with our findings, Gilbert and colleagues observed the irreversibly CNS impairments in rats induced by developmental hypothyroidism from GD6 to PN30. This review supplies us a clue to examine the mechanism by which thyroid hormone regu lates brain advancement during lactational and adolescent periods. As a member of MAPK, ERK1 2 is actually a important memory knowing it relevant protein inside the generation of discovering and memory. Thyroid hormone activates G protein coupled receptors, leads to ERK1 2 phosphorylation, and in flip enhances the transcription of some proteins that are essential in memory. It’s been proven that hypothyroidism reduces the basal p ERK1 two in hippocampal CA1 area with out affecting t ERK1 two. While in the DG region, on the other hand, hypothyroidism had no effect on t ERK1 two and p ERK1 two. Inside the lactational and adolescent rats, right here we show that t ERK1 2 and p ERK1 2 in iodine deficient, 15 ppm PTU therapy and five ppm PTU treat ment groups in CA1 and CA3 areas had been substantially reduced than controls.

The monoclonal anti entire body to phospho ERK was obtained from Cell Signaling Technologies. The rabbit anti ERK, utilised as controls for equal loading, was obtained from Santa Cruz Biotechnology. Goat anti rabbit and mouse IgG with a FITC conjugate have been obtained from Sigma. PD98059 was pur chased from Calbiochem and manufactured up with dimethyl sulfoxide at 1 mM stock answer. Pyrrolidien dithiocarbamate was obtained from Sigma and dissolved in PBS. NE PER Nuclear and Cyto plasmic Extraction Reagents was obtained from Pierce. Sodium fluorescein was bought from Amersco. Cell culture The human RPE D407 cell line was generously presented by Dr Guo Zhongmin. Cells were cultured in DMEM with high glucose. containing 10% FBS, penicil lin and streptomycin. The medium was transformed each and every two days, and cells have been subcul tured by trypsinization every single four days at a split of 1.5.
Tat protein planning and treatment The 86 amino acid isoform in the Tat protein was obtained in the Nationwide Institutes of Wellbeing AIDS over at this website Reagent System. Prasugrel It had been reconstituted in phosphate buffered saline containing 1 mg ml bovine serum albumin and 0. 1 mM dithiothreitol and deaerated by bubbling with helium. The protein was stored at 80 C within the dark in advance of use. The specificity of Tat mediated results was assessed by treating cells with heat inactivated Tat ready by incubating the protein at over 85 C for thirty min. Mainly because Tat binds strongly to serum proteins, all experiments have been carried out in serum no cost media. D407 cells remained healthier and viable underneath these experimental circumstances. The Tat therapy during the present study concerned exposing D407 cells exposure to a hundred nM Tat for 24 hours, which has regularly been used in preceding in vitro scientific studies. Cell viability assay Cells have been grown in 96 well plates at a density of one ? 104 cells well.
Immediately after the indicated treatments, MTT was added at 5 mg ml to every properly for four hours, after which the culture medium was removed and 1501 of DMSO was extra to every single well. The absorbance was measured at 490 nm utilizing a multifunctional microplate reader. Measurement of TER Transparent Millicell CM filters have been coated with 501 of the rat tail collagen I ethanol mixture and left to dry prior to cells had been subcul tured. D407 cells were seeded at a density of 104 cells filter over the filters was supported by 24 very well culture plates. The volumes to the apical and basolateral side have been 4001 and 6001, respec tively. The fluid strain was precisely the same from the two cham bers. The cultures have been incubated in a humidified ambiance. The medium was modified over the fol lowing day, and subsequently transformed just about every 2nd day for the duration with the experiment. Phase contrast micro scopy exposed that cells reached confluence at day 3, then serum concentration with the culture medium was reduced to 1%.

past 1 week, TGF beta1, 2, and three play a critical function in hyper trophic scar formation. Furthermore, knockout mice have revealed their importance in regulating in flammation and tissue restore. However, there exists no report about newly designed TGF B2 knock down transgenic mouse lines and also the sys temic distributions of TGF B2 in Tg mice. The surveys of TGF B2 distributions in newly developed TGF B2 knock down transgenic mouse lines supplied some cru cial details to investigate the role of TGF B2 below physiological and pathological issue. In summary, this research created Tg mice lines with TGF B2 down regulation and the systemic morphologic in formation that can be utilized in even more investigate. Our final results showed that TGF B2 proteins have been widespread in a number of tissues, mainly in nervous methods, intestines and epider mis.
These final results indicated that TGF B2 may play mul tiple different biologic roles according towards the distinctive cell sorts. Additionally, the existing effects created four geno kinds TGF B2 Tg mice of expressional down regulated by distinctive folds, which provided a variety of genotypes Tg mice sources for distinct research. Conclusion read full report Our examine established new transgenic mice lines with ex tensive down regulation of TGF B2. We also provided the down regulated charges and systemic distributions of TGF B2 protein in 4 phenotypic transgenic mice. The results showed that TGF B2 knockdown mice like Founder 66 might be designated since the target lines for more analysis. Approaches Animal generation Animal use and care had been in accordance with all the animal care recommendations, which conformed to the Guide for that Care and Use of Laboratory Animals published by the US National Institutes of Wellness.
TGF B2 knock down transgenic mice with C57BL 6J genetic background had been created by our collaborators during the Institute VX222 of Laboratory Animal Science. The generation in the transgenic mice was described as follows. Briefly, at least 3 silence expression web-sites of TGF B2 had been built by application supplied by Invitrogen Corporation, USA. Then we picked predesigned brief hairpin RNA that target mouse TGF B2 gene. The reconstruction plasmid was made and pur chased from Invitrogen Organization. The constructed recom binant plasmid was transferred into 293T cells. The transformants were screened and recognized by polymers chain reaction detections and restriction analysis. The protocol of PCR is described as follows. The transgene was then isolated from your cloning plasmid and purified by Avr II digestion, followed by diluted to a final concentration of five ng uL. The final transgenic fragment was microinjected into fertilized mouse eggs. De tection for your transgenic fragment is described as follows. Transgenic mice have been mated with nontransgenic element ners to sustain heterozygozity in the transgene or with transgenic partners to make homozygous transgenic offspring.

The usage of herbal treatments depending on spices like a type of complementary and choice medicine is doc umented while in the population especially amongst cancer pa tients. The spice based herbal medicines along with the constituents happen to be reported to inhibit the prolifera tion of cancer cells immediately. In vitro research indicate that herbs, spices, and their bioactive elements can in hibit, and from time to time induce pathways that regulate cell division, cell proliferation, detoxification, moreover to your inflammatory and immune response. For in stance, ursolic acid, a bioactive component in some herbs and spices, suppressed TNF induced expression of genes regulated by NF kB which are associated with tumor initiation, promo tion, and metastasis. In Cameroon, many scientific studies happen to be carried around the cytotoxic exercise of some spices on unique cell lines.
Dorstenia psilurus is actually a plant inhibitor chk inhibitor from extensively utilized in Cameroon, Africa and Madasgascar for different functions. Plant medication from this genus have shown a broader acceptability amongst some indigenous populations. The roots of D. psilurus are used in Cameroon as spices inside the traditional meal known as Nkui and Nah poh. It’s also claimed to protect well being, stop ageing, to im show the wellbeing and freshness of younger mothers right after a number of pregnancies. The chemical composition with regards to macronutrients and micronutrients of roots extract of D. psilurus continues to be reported. A decoction of leaves of D. psilurus is utilized in Cameroon to deal with rheumatism, snake bites, headache and stomach problems, arthralgia, cardiovascular issues, diuretic, tonic, stimulant, anal gesic, inflammatory ailments and cancers. Phyto chemical analysis of root extract of this plant demonstrated the presence of prenylated flavonoids, stearyl p coumarate, stearylferulate, benzofuran derivatives, Dorsilurins C, D and E and Dorsilurins.
The methanol extract of D. psilurus is reported to get anti inflammatory home. Some biological actions of root extract of D. psilurus such as the scavenging house on DPPH rad ical. anti amylase, anti lipase and antioxidant pursuits and hypertensive effects, glucosidase inhibitors professional perty. extra resources antibacterial activity and cytotoxicity action on MiaPaCa 2. CCRF CEM, CEM ADR5000 cells have already been demonstrated. On the other hand, there are no research around the cytotoxicity or apop tosis inducing properties on the roots extracts of D. psilurus on human promyelocytic leukemia and prostate cancer cell lines. For that reason this research aimed to de termine the cytotoxic with the methanol extract of D. psilurus on two cell lines and investigate its toxicological mechanism over the most sensitive cells. Solutions Plant material and extraction The roots of Dorstenia psilurus had been col lected at Komako while in the West Area of Cameroon and recognized by Mr Victor NANA, of the Nationwide Herbarium of Cameroon, in December 2010.

In the next area, we show how non symmetrical prototype models of heterogeneous differentiation amongst true lines of CD4 T cells can be studied within this unifying framework despite their varied attributes. Mathematical versions based upon the theoretical framework may be made use of to understand experimental success and make testable predictions In this area we discuss three prototype models for studying heterogeneous differentiation of CD4 T cells. The initial two versions are aimed to clarify some curiosity ing biological phenomena that weren’t studied previ ously with mathematical modeling. The third a single is often a simplified version of our earlier model.but we now have created it additional accessible by using the framework presented right here. On account of their limited scope, none of those versions are meant to supply a extensive comprehending on the corresponding biological techniques.
Rather, our intention is usually to illustrate how to use the mod eling framework to clarify observed heterogeneous dif ferentiation and make testable predictions. Prototype Model one. Heterogeneous differentiation of TH1 and TH2 cells Prior mathematical models effectively described the dynamic conduct along with the underlying molecular con trol program of your reciprocal differentiation of selleck chemical Ivacaftor TH1 and TH2 cells.Nevertheless, heterogeneous differenti ation of TH1 and TH2 cells and its underlying molecular controls were not studied with these versions. Yamashita et al. found that the heterogeneous differenti ation of TH1 and TH2 cells is usually obtained with anti genic stimulations. Comparable observations had been obtained by Hosken et al. and Messi et al. We now have developed a mathematical model, depending on the influence dia gram in Figure 2A, to describe heterogeneous differenti ation of TH1 and TH2 cells. The parameter values ARN-509 to the model are listed in Additional file 1.
Table S2. Figure 6A displays the bidirectional two parameter bi furcation diagram, and Figure 6B demonstrates the simulation results because the heterogeneity score with respect to your two single constructive phenotypes. Our simulation results recommend that exogenous polarizing signals, i. e. IL four and IL 12, will not be adequate to set off differentiation. They has to be accompanied by a sufficiently substantial dose of antigenic stimulant to set off the differenti ation to the corresponding phenotypes. This conclu sion is in agreement with former experimental success.High strength of TCR signal alone or with intermediate degree of IL four was ample to induce the differentiation of two single positive phe notypes. With rising strengths of TCR signal, our simulations display a spectrum of heterogeneous popula tions with expanding percentages of TH2 cells and de creasing percentage of TH1 cells.

Just about every dataset was saved as an Excel file. Every single proteomic dataset with identifier was input in to the IPA examination procedure with all the Core evaluation platform. For your pituitary adenoma protein mapping data and nitroprotein information, their Swiss Prot accession numbers in the Excel format were input for the IPA data upload workflow. To the pituitary adenoma comparative proteomic data, the Swiss Prot Accession numbers and also the corresponding fold transform information while in the Excel format had been input on the IPA information upload workflow. The IPA sys tem will automatically search the matched Gene. mole cules, and can generate a two dimensional table style format to show which protein was mapped within the program for following step analysis, and to display the unmapped pro teins. The unmapped proteins Swiss Prot accession variety will probably be converted for the corresponding gene identify by looking the ExPASy proteomics server.
All Swiss Prot accession numbers in combination with gene names were input on the IPA information upload workflow to make the ultimate mapped record for following stage evaluation. The dataset, such as mapped IDs.was saved, and immediately produced 5 subdatasets, which include the All IDs.Unmapped IDs.Mapped IDs.Network eligible IDs.and Functions. EVP 4593 Pathways.Record eligible IDs. To the duplicate IDs for the identical pro tein. gene, the identifier with the highest fold adjust was utilized in the pathway analysis.or, the primary instance with the protein. gene was used in the pathway analysis while in the absence of an expression worth for example mapping pro teomic information and nitroprotein data. Every subdataset con tained ID, notes, molecules, description, spot, sort of biofunction, and medicines.along with the fold adjust.The name of every molecule seems from the pathway network nodes.
The Network eligible IDs proceeded into the pathway network analysis by comparing the network eligible mol ecules together with the Ingenuity Pathways Evaluation Understanding hop over to this site Base.IPAKB is a curated database that includes a lot of scientific findings which have been extracted from a huge selection of thousands of journal posts, textbooks, together with other information sources, and many canonical pathways which are constructed from people scientific findings.The significance from the association among the dataset along with the canonical path way was measured by comparing the quantity of use spe cific proteins of interest that participate in a given pathway towards the complete amount of occurrences of those genes in all pathway annotations which might be stored from the IPAKB. A Fishers actual test was used to calculate the p value to determine the probability the association amongst the genes inside the dataset as well as canonical path way is explained only by chance. The level of statistical significance was set to p 0. 05. Just about every Pathway analysis produced the leading networks, biofunctions.

In vivo, we observed quite a few hypotheses associated with Amiodarones recommended mechanisms of action by cellular Ca and potassium modulation.and reported negative effects such as binding to thyroid antagon ism and hypothyroidism.None in the mechanism related hypotheses had been found in vitro. In addition, all key causal reasoning supported biological networks have been substantially different. Irritation is one of the important signaling networks predicted, albeit with opposite directionality staying predicted decreased in vivo and pre dicted improved in vitro. Advised downstream results varied substantially likewise, decreased cell cycle in vivo ver sus apoptosis in vitro and a greater tissue remodeling. struc tural signal principally driven by decreased TGFB in vitro. In the hypothesis level incredibly few similarities had been located in between in vivo cardiac tissue and in vitro principal rat cardiomyoctes, e. g.
Hypoxia and SRF hypotheses. Contrary to Amiodarone, Dexamethasone exhibits higher degree of in vivo to in vitro translatability at the two the course of action and personal hypothesis ranges. Figure three demonstrates the causal reasoning inferred molecular response to Dexamethasone in rat cardiac tissue and Pri mary rat cardiomyocytes.Causal reasoning generated a variety of individual hypotheses reflective of dexamethasone selleckchem action such as Dexamethasone.NR3C1 and glucocorticoid.Recognized dexamethasone ef fect can also be reflected by supported biological processes this kind of since the anti inflammatory sub network both in vivo and in vitro. Dexamethasone is also very translatable to H9C2 cells too using a causal network that is very related to that of primary rat cardiomyocytes.In vivo to in vitro translatability in the main biological processes The leading ranking causal networks from every single in vivo or in vitro experiment have been summarized with the biological system level in Figure four.
A network was established for being leading ranking if it had been supported by a cluster of at the least three hypotheses and considered one of which ranks during the major 25 hy Delanzomib potheses as previously described.For every com pound at the least 1 process was translatable to no less than considered one of the two cell lines applied. Total, H9C2 cells exhibited larger quantity of biological networks, probably a reflection of greater sensitivity as in contrast to each primary rat cardiomyocytes and in vivo cardiac tissue. H9C2 cells also demonstrated a trend of common cell anxiety. cytotoxicity responses that don’t always trans late to in vivo occasions, this kind of as endoplasmic reticulum strain and oxidative tension. On the other hand, for each compound there was not less than one biological process that translated properly from in vivo to H9C2 cells. Some of the biological processes which can be supported to translate equally nicely in H9C2s and RCMs are decreased cell cycle signaling, in creased tissue remodeling and elevated DNA harm and fix.

haracteristic down regulation of E cadherin is regarded as the key step to EMT. HCCs with EMT options regularly exhibit additional venous invasion, metastases, and also a poorer prognosis than those without having EMT traits.Whether or not inadequate RFA directly induces the EMT of residual HCC cells and even more promotes the metastasis stays unclear. Inside the existing study, we investigated the morpho logical changes, cell growth, migration and invasion of HCC cell lines after inadequate RFA in vitro. Moreover, we analyzed the adjustments of epithelial and mesenchymal markers, and Akt and ERK1. two signaling pathways associated with the course of action in HCC cells after insufficient RFA. We also performed in vivo experiments to research the development and metastasis of HCC cells after insufficient RFA within a BALB. c nu. nu mice model. Approaches Cell culture Established human HCC cell lines, SMMC7721 and Huh7 were in the American Form Culture Assortment.
All cells have been maintained in higher glucose Dulbeccos modified Eagle medium supplement with 10% fetal bovine serum.100 U. ml penicillin and 100 ug. ml streptomycin in a humidi fied atmosphere of 5% CO2 at 37 C. selleck chemical Raf Inhibitors Chemicals and antibodies LY294002 and PD98059 were purchased from Beyotime.Antibodies with specificity to the phos phorylated forms of Akt and ERK1. 2 had been purchased from Cell signaling.Antibodies recognizing E cadherin, N cadherin, vimentin, snail and SMA were bought from Abcam.Antibodies recognizing B actin, MMP two and MMP 9 antibodies were obtained from Santa Cruz.Heat remedy Inadequate RFA was simulated in vitro as described be fore.Briefly, SMMC7721 or Huh7 cells had been seeded into the six nicely plates.After 24 h, the plates had been sealed and submerged within a water bath set to 47 C for 5 min.
Thereafter, cells were allowed to recover, and when the surviving populations reached 80% conflu ence, cells had been propagated to the six effectively plates and exposed to over heat therapy for ten min. Then the procedure was repeated and cells had been sequentially exposed to above heat treatment for 15 min, selleck 20 min and 25 min. Cells survived in the therapy have been designated as SMMC7721 H and Huh7 H respectively. The morpho logical traits of HCC cells have been observed by microscopy.Proliferation assay Cell proliferation was analyzed applying the three 2, 5 diphenyltetrazolium bromide assay. Briefly, HCC cells had been cultured in 96 properly plates at a concentration of three 103 cells. very well, and incu bated for 24 h, 48 h, or 72 h. MTT remedy was added to every single very well at a last concentration of 0. five mg. ml and incubated for 4 h. At the end of incubation, formazan crystals resulting from MTT reduction had been dissolved by addition of 150 ul dimethyl sulfoxide per effectively.

L 8 gene expression levels are acknowledged to become strikingly variable, ranging from not detectable amounts to over 100 fold activation based about the inducing stimuli.The core IL 8 promoter is made up of a NF kB component likewise as activating protein one and CAAT. enhancer binding protein binding websites.As opposed to the NF kB binding web sites, the AP 1 and C. EBP internet sites are not essential for IL 8 induction but are demanded for maximum gene expression in some cell types.We hypothesized that the robust expression of IL eight gene following sulindac sulfide therapy may very well be due to activation of both NF kB and AP one transcription aspects. AP 1 is often a transcription aspect composed of homo or heterodimers in the Jun, Fos and ATF household members.We studied the nuclear and cytoplasmic protein ranges from the NF kB subunit p65 along with the AP 1 members c Fos and c Jun in control and sulindac sulfide handled cells.
Treatment with sulindac sulfide kinase inhibitor erismodegib enhanced the quantity of nuclear p65, connected which has a reduce in cyto plasmic p65 amounts, indicative of p65 nuclear translocation following the drug therapy.The amounts of total c Jun and c Fos had been up regulated in each the cytoplasmic and nuclear fraction on sulindac sulfide therapy and there was a marked maximize in phosphorylated c Jun and JunD indicative of activation on the JNK pathway.In consensus together with the western blot success, the mRNA expres sion amounts of c JUN and c FOS were markedly upregulated inside the presence of 50 and 120 uM sulindac sulfide and c FOS ranges also enhanced following treatment method with twenty uM sulindac sulfide for 4 hours.Upcoming we assessed irrespective of whether each transcription components are required for the strong up regulation of IL eight gene expression employing an IL eight promoter construct cloned within a luciferase reporter vector, with or with no mutated NF kB and AP 1 binding web sites.
Treatment with TNF was utilised being a optimistic manage and resulted within a robust enhance in luciferase action, which was somewhat down regulated in cells transfected using the AP 1 mutant binding sites but fully abolished in cells transfected with mutated NF kB binding web sites.In contrast, in sulindac KRN-633 sulfide taken care of cells each the mutated AP 1 and NF kB binding web-sites strongly diminished the up regulation in luciferase action.These benefits indicate that the powerful up regulation of IL eight gene expression induced by sulindac sulfide treatment is dependent on both the NF kB and AP one transcription elements, whereas TNF up regulates IL eight primarily by way of NF kB. Sulindac sulfide modulates TNF induced IkB phosphorylation and degradation Considering the fact that sulindac sulfide induced a lower in IkB protein ranges in basal ailments, and induced up regulation of NF kB target genes, we following tested the result of sulindac sulfide on IkB in conditions where the canonical NF kB pathway is activated through stimulation from the cyto kine TNF.

utations on the Smad4 gene in other tumor kinds are less regular, which has a 16% charge in biliary tract cancers, 13% in colorectal carcinomas, 12% in breast cancers, bladder cancers and ovarian cancers, 7% in lung cancers, 6% in hepatocellular carcinoma and 4% in cervical cancers.Studies in cervical tissue have shown similar levels of Smad4 mRNA expression in non malignant and premalignant tissue. Nevertheless, Smad4 expression is decreased or misplaced in 90% of cervical squamous cell carcinomas.An immunohistological review in cervical squamous cell motor vehicle cinomas uncovered a significant correlation of weak cyto plasmic Smad4 staining with the two the presence of positive lymph nodes and recurrent disease.Fur thermore, absence of nuclear Smad4 protein expression strongly correlated with tumors size and infiltration depth.
Both weak cytoplasmic Smad4 and the ab sence of nuclear Smad4 staining had been associated with bad survival in cervical cancer individuals.The kinase inhibitor Vorinostat HPV16 good SiHa human cervical carcinoma cell line is refractory to growth inhibition by TGF B, which can be explained, a minimum of in part, by decreased expression of Smad4 in these cells.Transfection of SiHa by using a Smad4 expression construct recovered the growth in hibitory effects of TGF B in these cells.Our scientific studies of Smad4 protein expression in our in vitro model of HPV16 mediated transformation indicate that immortal ization by HPV16 DNA triggers an early raise in Smad4 protein in HKc. HPV16, as compared to standard HKc, and this maximize is maintained at later phases of in vitro progression. The functional significance of this getting stays to become determined.
Smad7 works as a feedback loop attenuating TGF B signaling. Each deletions and amplifications with the Smad7 gene happen to be reported in colorectal tumors.None theless, amplifications are additional prevalent than deletions, as in contrast to your same genetic alterations of Smad2 and Smad4, which suggests that retention as well as amplification of Smad7 could be the picked selleckchem MEK Inhibitor occasion during colorectal tumorigenesis.These findings are in agreement with frequent Smad7 overexpression located in endometrial and thyroid follicular carcinomas.Scientific studies have also discovered upregulation of Smad7 mRNA in pancreatic cancer as in contrast to normal tissue.On top of that, transfection of Smad7 to the TGF B sensi tive pancreatic cell line COLO 357 rendered them re fractory towards the antiproliferative impact in the cytokine, and significantly enhanced soft agar colony formation.In one more research, primary mouse keratinocytes were transduced together with the Smad7 gene resulting in enhanced keratinocyte proliferation, blocked normal differentiation, and induced keratin 8, a marker of malignant conversion, but did not lead to tumor formation.W