The smaller size cliques were additional to Identification of cliques Cliques are entirely linked, conserved, and co expressed inside the networks.We formulated a graph based mostly this network of cliques. These CCPs could then be utilised to comprehend the commonality and uniqueness as gene signatures in CRC across populations. Background New proteomics methods enabled huge scale experi ments that check phosphorylation states of a lot of proteins underneath various physiological stimuli and. or pharmacological treatments. Every single measurement captures a static image of how the cellular signaling network responses for the binding of the ligand to its receptor, however the interconnections among a variety of ligand acti vated pathways are complex and dynamic. As a result, it truly is of biological value to infer which signaling path is at get the job done in response to a particular ligand and how path techniques cross talk to each other inside a cell variety distinct method, and sooner or later to develop computational mod els capable of predicting cellular responses below differ ent stimuli.
One particular on the most typical approaches to signaling net do the job modeling is always to signify the dynamic procedure being a set of ordinary differential equations making use of mass action kinetics, by which the concentration of species with time may be analyzed.In addition, selleck chemical when spatial facts is significant for such modeling, a process of partial differential equations is considered for being more precise device to model biochemical processes in each room and time dimension.ODEs or PDEs mathema tically signify signal transduction by introducing lots of parameters while in the model, which becomes impractical for particularly significant networks due to the expanding difficulty in parameter estimation.To strategy substantial scale sig naling network modeling, quite a few information driven methods have emerged and utilized to simulate signal transduction.
constraint primarily based network analysis that allows reconstruc tion of substantial programs of biochemical reactions you can look here in analyzing genome scale metabolic networks.network compo nent analysis which incorporates prior information of network topology to infer signaling pathways.partial least squares regression analysis to investi gate challenging signaling networks by identifying optimum principle part based mostly dimensions from a proposed romantic relationship.Many of the approaches described over are deterministic versions, which are not aimed at accommodating the noise inherent in biologically information.In contrast, Bayesian network evaluation is surely an alterna tive probabilistic graphical strategy to model signaling pathways.Bayesian network, which might explicitly manage the uncertainty of unobserved events.supplies a compact graphical representation from the joint probability distributions over all random variables, and has become utilised for reconstruction of signaling networks.

Briefly, tumor cells were harvested and resuspended in binding buffer.a hundred ul of the cell suspension was stained with Annexin V FITC. Right after 10 minutes, 0. 4 ml of binding buf fer and ten ul of 7 AAD were extra, as well as the cells had been ana lyzed promptly by movement cytometry. Early apoptotic cells have been defined as Annexin V 7 AAD.late apoptotic cells as Annexin V 7 AAD.dead cells as Saracatinib SRC inhibitor Annexin V. seven AAD and viable cells as Annexin V. seven AAD.Immunohistochemistry Organs and tissues extracted from mice have been fixed in 4% formalin, dehydrated via grades of ethanol in the Microm STP 420D sample processor.embedded in paraffin in EC 350 1 paraffin station and sliced in four um sections. For human CD20 inmmunohisto chemistry, the antigen retrieval was performed with citrate pH six in microwave. Slides have been stained with anti human CD20 antibody by using the VENTANA BenchMark ULTRA Procedure.
Statistical evaluation Statistical significance among untreated and handled samples was assessed with Students t test for unpaired data.Differences were viewed as substantial if P values have been 0. 05. Survival data had been analyzed from the Kaplan Meier technique plus the Tarone Ware test was made use of to check for significance amongst all of the groups. Effects Anti CCR7 mAb blocks migration of mantle cell lymphoma cells in response selleck to CCL19 and CCL21 in vitro and mediates CDC and ADCC CCR7 is expressed in different main lymphoprolifera tive issues as we and some others have previously demon strated.Within this research, we have confirmed and extended these success inside a more substantial cohort of patients. Our final results show that tumor cells from CLL and MCL individuals consistently express CCR7 around the cell surface and at large density, in comparison with other lymphoproliferative syn dromes.
Other lymphomas such as follicular lymphomas or lymphoplasmacytic lymphomas also include tumor populations expressing CCR7 but mixed with CCR7 adverse tumor cells.For that reason, CCR7 may represent a whole new therapeutic target for the treatment method of particular hematological cancers, in particular MCL and CLL. To assess the potential of anti CCR7 antibodies to inhibit in vitro migration of MCL cells in direction of the ligands of CCR7, the homeostatic chemokines CCL19 and CCL21, we made use of the CCR7 positive Granta 519 MCL cell line.As shown in Figure 1B, we demon strated in transwell chamber assays that anti CCR7 mAb was extremely productive in preventing Granta 519 cells chemotactic response to CCL19 or CCL21. An other anti CCR7 mAb was also in a position to partially block Granta 519 migration, albeit significantly less successfully than clone 150503.The ability of your anti human CCR7 antibodies to in duce CDC or ADCC in MCL cells was also evaluated while in the MCL cell line Granta 519. To execute the CDC ex periments, the cells have been preincubated with two unique clones with the anti CCR7 antibody or using the correspond ing isotype manage for 1 hour and incubated with rabbit complement during an additional hour.

Depletion of cyclin D1 and p21 prevents mammary tumor development and area invasion Overexpression of p21 and cyclin D1 is correlated with poor prognosis and aggressiveness in breast cancer. To deal with the importance of p21 and cyclin D1 on breast cancer advancement in vivo, we injected either SCP2 con trol or double p21 and cyclin D1 knockdown cells to the mammary body fat pads of female Balb. c nude mice to watch main tumor growth and community invasiveness. Silencing p21 and cyclin D1 expression applying siRNAs sig nificantly lowered the price of primary tumor formation and tumor size.As depletion of p21 alone did not impact tumor formation in the Xenograft transplan tation in vivo model.it can be possible the observed phenotype on tumor formation in the double knockdown is mediated by cyclin D1. This is certainly in agreement with previous studies showing that depletion of cyclin D1 pre vented tumor development in oncogenic HER2 overex pressing transgenic mice.
Importantly, 3 out of 6 mice inside the manage group had tumors ulcerating through the overlaying skin, although all of the mice in the double knockdown group had intact skin. Breast tumor with ulcerated skin continues to be clinically classified as locally superior breast cancer. All tumors order RO4929097 were taken with the overlaying skin and surrounding tissues and subjected to hematoxylin and eosin staining. As proven in Figure 5B, the deep tumor margins from the manage group have been significantly less distinct, invading nearby structures, together with skeletal muscles as well as mammary extra fat pad, and showed frequent lymphovascular invasion. Nevertheless, the tumor margins within the knockdown group were effectively encapsulated using a non invasive nature. Also, we performed immuno histochemistry on principal mammary tumor derived from animals injected with parental and p21.
cyclin D1 depleted SCP2 cells. We assessed the expression with the TGFb regulated gene PTGS2, which we now have previously proven to be involved in mediating the TGFb result on cell migration and invasion.As proven in Figure 5C, applying tumors from four different mice in every single group, we uncovered expression of PTGS2 to become obviously higher in paren tal tumors when compared to p21. MG-132 Proteasome inhibitor cyclin D1 depleted tumors, even more confirming that the p21. cyclin D1 depleted tumors displayed much less invasive characteristics. To investigate the part of p21 and cyclin D1 about the improvement of bone osteolytic lesions, parental and dou ble knockdown SCP2 cells were injected intramuscularly into the left tibia of two groups of nude mice. As proven in Figure 5D, following X ray examination of the bones, both group of mice produced secondary tumors that triggered extreme osteolytic bone lesions, suggesting that p21.

In addition a phosphatase was absolutely suppressed after HIV infection.The downregulation of PPAC is deemed to be vital due to the fact its absence is important for maintain ing phosphorylation of a variety of tyrosine kinases and acti vation of endothelial cell development in vivo.The biological significance of all 31 proteins recognized on this examine was computed in relation to protein interaction networks concerned in angiogenesis.This, we believe, will be the to start with phase toward establishing a better insight to the molecular mechanisms by which pathogenic viruses such as HIV may possibly initiate and. or promote angio genesis while in the contaminated host. Stepwise Analyses of Necessary Biological Processes in Angiogenesis Angiogenesis is known as a multifactorial biological course of action involv ing numerous ways as well as endothelial cell activation, degradation of basement membrane, cell proliferation, invasion, morphogenesis, sprouting, migration and stabi lization of microvessel formation.
Each step entails a series of really complicated but well orchestrated protein protein interactions along various signaling pathways. To understand the biological significance of every protein, we’ve got divided all proteins into 10 well acknowledged biologi cal occasions in the course of neovascularization or angiogenesis.and mentioned putative functions of each professional tein in that group. Considering the fact that u0126 Uo126 most proteins are multifunc tional, some overlap from the protein functions was inevitable. Phase one Activation of T Cells. Transcriptional and Translational Reprogramming The moment the HIV envelope glycoproteins bind to your T cell receptor and co receptors.the cell surface proteins are clustered. This generates a cascade of signals from the plasma membrane on the cytoplasm and nucleus. Because the new proteins are expressed, the HIV contaminated cells are activated and therefore are driven towards apoptotic pathways.
However, most activated cells also make numerous cytokines, enzymes together with other signal transduction molecules that invoke innate cellular immunity and might be crucial for that survival with the infected cells. These proteins retain cellular integrity through var ious phases of HIV replication and cell development. Numerous professional teins that happen to be upregulated, downregulated or induced de novo publish HIV infection can also be necessary selleck chemical to compen sate for your reduction or disruption of essential physiological functions performed by the T lymphocytes before HIV infection. Between a diverse relatives of multifunctional signaling professional teins induced de novo in HIV contaminated cells, the protein tyrosine kinases, the serine. threonine kinases and many regulatory enzymes seem to play major roles in T cell activation and global reprogramming in the transcrip tional and translational actions that bring about novel inter action pathways.

In addition a phosphatase was entirely suppressed soon after HIV infection.The downregulation of PPAC is regarded as to be important simply because its absence is crucial for maintain ing phosphorylation of several tyrosine kinases and acti vation of endothelial cell growth in vivo.The biological significance of all 31 proteins recognized on this research was computed in relation to protein interaction networks involved in angiogenesis.This, we think, may be the initial stage towards creating a greater insight into the molecular mechanisms by which pathogenic viruses such as HIV may well initiate and. or encourage angio genesis in the contaminated host. Stepwise Analyses of Very important Biological Processes in Angiogenesis Angiogenesis can be a multifactorial biological process involv ing several techniques which includes endothelial cell activation, degradation of basement membrane, cell proliferation, invasion, morphogenesis, sprouting, migration and stabi lization of microvessel formation.
Every step includes a series of really complicated but properly orchestrated protein protein interactions along many signaling pathways. To comprehend the biological significance of every protein, we have divided all proteins into 10 effectively acknowledged biologi cal occasions during neovascularization or angiogenesis.and mentioned putative functions of each professional tein in that category. Due to the fact recommended reading most proteins are multifunc tional, some overlap from the protein functions was inevitable. Step 1 Activation of T Cells. Transcriptional and Translational Reprogramming As soon as the HIV envelope glycoproteins bind on the T cell receptor and co receptors.the cell surface proteins are clustered. This generates a cascade of signals from your plasma membrane on the cytoplasm and nucleus. Because the new proteins are expressed, the HIV contaminated cells are activated and are driven toward apoptotic pathways.
However, most activated cells also create a number of cytokines, enzymes along with other signal transduction molecules that invoke innate cellular immunity and might be essential to the survival with the infected cells. These proteins retain cellular integrity for the duration of var ious phases of HIV replication and cell growth. Lots of professional teins which might be upregulated, downregulated or induced de novo submit HIV infection can also be needed selleck to compen sate for that loss or disruption of necessary physiological functions carried out from the T lymphocytes prior to HIV infection. Amid a diverse family members of multifunctional signaling professional teins induced de novo in HIV infected cells, the protein tyrosine kinases, the serine. threonine kinases and lots of regulatory enzymes appear to play leading roles in T cell activation and worldwide reprogramming with the transcrip tional and translational routines that cause novel inter action pathways.

Tortuous capillaries are noticeable with the surface of LP 1D1b tumors when LP 1K tumors, characterized by lack of size progress, were not perfused. Hence, cyclin D1b promotes neoangiogenesis and consequently, tumor growth in vivo. To confirm the involvement of neoangiogenesis in tum origenesis of LP 1D1b cells in xenografts, we injected either as soon as VEGF siRNA on the vicinity of your injection web page or biweekly, chemical FGFR or VEGFR inhibitors, SSR and SAR respectively. As shown Figure 6c, as expected, scrambled siRNA had no results on tumor evolution. Administration of VEGF siRNA markedly diminished the volume of LP 1D1b derived tumors for a 15 day time period. Following 15 days, no far more effects of VEGF siRNA were observed very likely resulting from siRNA degradation along with the tumor grew having a rate similar to the one of control. This is often in agreement together with the reported stability of siRNA during the delivery gel.
Importantly, SSR and SAR inhibitors entirely abol ished the growth of tumors indicating a position of FGFR and VEGFR in the tumor evolution. The capacity of VEGF siRNA as well as TK inhibitors to inhibit tumor development strongly supports microarray and CAM data and the con clusion that cyclin D1b favors tumorigenesis by way of activation of a neoangiogenic procedure. Discussion Cyclin D1 is overexpressed in a broad array of solid malignancies, selleck expressed in lymphoid tumors such as MM and MCL and not within their regular counterparts. Nevertheless, in vivo scientific studies failed to reveal a powerful oncogenic poten tial from the traditional cyclin D1, referred to cyclin D1a.By contrast, the cyclin D1 isoform b as well as mutant cyclin D1 T286A are capable to transform cells in vitro and to induce tumors in vivo.These two varieties of cyclin D1 share a strict nuclear localization sug gesting that nuclear functions of cyclin D1 are vital and.
or ample for tumor formation. Mutations in the CCND1 gene disrupting the phosphorylation at Thr286 and thereby leading to nuclear accumulation of cyclin D1 have been described in endometrial and esophageal chk inhibitor automobile cinomas additional reinforcing this notion.On the other hand, the molecular mechanisms of cyclin D1b driven tumori genesis aren’t entirely elucidated. In cultured cells, cyclin D1b will not be capable to activate its catalytic spouse CDK4 and in flip, won’t regulate positively the cell cycle.retains a strong transcriptional co repressor activ ity, displays lowered binding to p27Kip1 and isn’t going to con trol cell migration.Right here we show that, during the context of MM cells, cyclin D1b confers a complete malignant pheno style and enables cells engraftment in immune compro mised mice. The genome broad examination of LP 1D1b cells extends our knowing on the biological properties of cyclin D1b. Furthermore, we’ve got identified genes regulated by cyclin K, a viral oncogenic homolog of cyclin D1a and confirm the basic differences between the two cyclin D1 isoforms.

n MCF7 cells, HRG B1 induced nuclear colocalization of phospho Smad2 and Snail, and pretreat ment with LY294002 and SB203580 suppressed the nu clear translocation induced by HRG B1.The imply percentages of fluorescence of phospho Smad2 and Snail may also be shown in Figure six. HRG B1 induces EMT by means of phospho Smad2 mediated Snail through the PI3k. Akt signaling pathway As talked about earlier, HRG B1 enhanced the expres sions of vimentin and fibronectin all through EMT in SK BR three and MCF7 cells. As shown in Figure 7a, b, the HRG B1 induced expressions of vimentin and fibronectin have been inhibited by the indicated inhibi tors. Taken together, HRG B1 induced EMT via phospho Smad2 mediated expression of Snail by way of the PI3k. Akt signaling pathway in both breast cancer cell lines.
Knockdown of Smad2 expression suppresses HRG B1 induced expressions of Snail and fibronectin SK BR 3 and MCF7 cells have been transfected with control and Smad2 siRNAs. As shown in Figure 8a, b, the HRG B1 elevated expressions of Snail and fibronectin in con trol siRNA transfected selleck cells compared with un taken care of management cells had been downregulated in Smad2 siRNA transfected cells.Taken to gether, Smad2 activation plays roles inside the expression of Snail and induction of EMT by HRG B1 in SK BR 3 and MCF7 cells. HRG B1 and ErbB3 induces cancer cell migration and invasion through Smad2 activation We carried out in vitro wound healing assays. Pretreat ment with LY294002 and PD169316 or SB203580 inhibited the cell migration of SK BR three and MCF7 cells inside the presence of HRG B1.
In cell inva sion assay, knockdown of ErbB3 and Smad2 by siRNA inhibitor CP-690550 transfection inhibited the cell invasive ability of SK BR three and MCF7 cells underneath HRG B1 stimulation in matrigel coated chamber.Collectively, these information suggested that HRG B1 induced cancer cell migration and invasion by induction of EMT by means of PI3k. Akt phospho Smad2 Snail signaling pathway. Discussion Breast cancer is the most common malignancy between gals worldwide. Knowing the mechanisms of cancer invasion and metastasis is a important concern in cancer investigate. Nearly all research regarding EMT have centered on TGF B signaling in numerous kinds of sickness settings.Therefore far, the basal like sort and triple negative type of breast carcinomas are charac terized to demonstrate mesenchymal and stem cell capabilities and are identified for being correlated with resistance to therapy.It’s been suggested that not merely TGF B but also numerous kind of signaling molecules, such as development fac tors, cytokines, integrins, and Wnts, are inducers of EMT.H

ontrol pair. Every single single microarray analysis was derived from a single cell culture. The taken care of cells were in contrast on the management. Probesets exhibiting a signal log2 ratio 1. 0 as well as a transform p value 0. 004 or perhaps a signal log2 ratio one. 0 along with a transform p value 0. 996 had been identified by filtering employing the Affymetrix Information Mining Device 3. 0.Addition ally unsupervised clustering was carried out in between the 19 management tumor samples. Genes associated with apoptotic pathways were selected primarily based on Gene Ontology examination.Expression improvements were correlated to the grading as well as the histological sub entity with the tumors. Only tumor samples have been included within the ultimate analysis whose gross sections had been diagnosed as liposarcomas by an seasoned soft tissue pathologist.
According to your amount of differentially expressed genes just after incubation with doxorubicin, liposarcomas have been cat egorized into substantial.intermediate and lower responders.The outcomes have been uploaded selleck Telatinib to NCBI GEO, quantity GSE12972.Authentic time PCR for microarray information validation Microarray data validation was carried out for picked gene merchandise with relevant up regulation in in excess of ten out of the 19 liposarcoma probes or relevant down regulation in no less than ten probes.CD14 and TIA weren’t examined because no higher high-quality PCR assays have been purchasable. Effects Sixteen tumors have been located in the extremities, 3 at the trunk. The tumors diameters ranged from one cm to 38,5 cm. Twelve principal cultures have been harvested from primary tumors, six from area recurrences, and one from a metastatic tumor. The majority of tumors were large grade.
only 4 had been diagnosed minimal grade.Atypical lipoma or very differentiated liposarcoma was by far the most selleck chemicals URB597 common histological subentity followed by dedifferentiated and pleomorphic.myxoid. rundcell and myxoid liposarcoma. All low grade tumors had been diagnosed as atypical lipoma, whereas 3 atypical lipomas had been categorized as G2 tumors because of aggres sive development or localized locations of dedifferentiation. Another subentities were all categorized as higher grade tumors.A summary with the sufferers information is given in table 1. Hierarchical clustering of expression profiles from untreated samples did not clearly separate according to histological subtype or grading, but unveiled two important branches, which showed some enrichment for G3 and G2 tumors.In accordance for the number of differentially expressed genes n immediately after incubation with doxorubicin, the 19 liposarcomas were categorized into high.intermediate and reduced responders.All poorly dif ferentiated tumors were large responders.G2 tumors were predominantly intermediate responders.and, many of the G1 tumors had been reduced responders. A heatmap of all differentially expressed genes after 24 h of doxorubicin tratment exhibits heterogeneous response patterns.C

n distinct, miR 335 and miR 365 have been one of the most very upregulated person miRs identified in Rasless cells that had been also simultan eously detected as becoming downregulated in each BRAF and MEK1 rescued MEFs.Moreover, miR 27a plus the miR 30a, miR 30b and miR 30c had been also upregulated in Rasless cells but transcriptional reversal was only detectable for miR 27a and miR 30a in MEK1 rescued cells. In other cases, this kind of as the downregulated allow seven loved ones members or even the upregulated miR 10b, miR 129, miR 215, miR 487b and miR 883, no reversal of their transcriptional pattern was detected.Direct visual proof with the reversibility within the miRNA profile of Rasless cells is provided through the heatmap in Figure 5B, depicting a multiclass comparison resulting from hierarchical clustering of your microarray primarily based miRNA profiles of control, Rasless, and BRAF or MEK1 rescued MEF clones.
This dendrogram displays a clear discrimination among TSA hdac inhibitor Trichostatin A three key vertical branches corresponding to control, proliferating K Raslox MEFs, the BRAF or MEK1 rescued MEFs along with the non proliferating, Rasless cells.Remarkably, the profiles in the BRAF and MEK1 rescued MEFs showed an expression pattern that was antagonistic to that of Rasless cells, hence regaining a miRNA profile that approached that within the unique proliferating handle K Raslox MEFs.Mechanistic implications inferred from your reversible miRNA expression patterns of Rasless cells We attempted to recognize just about the most salient reversible miRNA alterations with regards towards the generation and.or servicing from the Rasless status by means of Venn diagrams identifying miRs from Table 1 that showed an precisely opposite pattern of differential expression involving Rasless cells and both the BRAF rescued and MEK1 rescued proliferating MEFs.
This method identi fied at the very least 34 distinct repressed miRs and six overex pressed miRs of Rasless cells fulfilling that issue.This particular group of 40 miRs probably represents the PCI-32765 price core of most func tionally relevant miRs with regards for the mechanisms involved within the generation and. or reversal of your Rasless phenotype. An total see within the most significant cellular func tional classes predicted for being impacted by this pool of Rasless miRNAs is represented in Figure 6C. Interest ingly, this evaluation acknowledged a set of standard GO func tional classes that are tremendously coincident with these previously recognized inside a similar examination within the pool of re versible mRNAs of Rasless cells.Amongst some others, these incorporated the following.Transport.Protein phosphorylation.Tiny GTPase mediated signalling and DNA. RNA processing.Concentrating on the identity of the personal miRs within this group, it was striking to observe a significant enrichment in miRs belonging to a brief list of precise miR clusters and households characterized by their shared skill to target quite a few certain cellular regulators participating in modulation of cell cycle progression.

two,5 diphenyltetrazolium bromide],phosphate buffered saline,dimethyl sulf oxide,F twelve K medium,NGF 7 S from murine submax illary gland, MEK inhibitor,and PI3K inhibitor had been obtained from Sigma Co. Fetal bovine serum and horse serum were purchased from PAA Laboratories. Cultivation ailment of mushrooms Pleurotus giganteus was maintained on po tato dextrose agar at 4 10 C and on a regular basis sub cultured. The substrate formulation for your cultivation of P. giganteus is equivalent to that for oyster mushroom cultivation, i. e. 89 94% rubber wood sawdust, five 10% rice bran and 1% calcium carbonate. Polypropylene bags are applied for substrate bagging as well as moisture content material within the substrate was stored at 60% 65%. The temperature for mycelia development, spawn run, and fruiting entire body formation is 26 32 C. Relative hu midity of 70% and 80 90% throughout mycelia development and fruiting. respectively, must be maintained.
Direct illu mination need to be averted because it has become reported to inhibit the fruiting notch inhibitor body formation. A twenty day cycle right after total colonization in the artificial log is needed for each harvest and about four harvests can be obtained from each bag of 900 g. Cell culture The PC12 cells from ATCC have been maintained in F 12 K medium sup plemented with two. 5% heat inactivated fetal bovine serum and 15% horse serum with final pH 6. 8 7. two. All incubations were carried out at 37 C within a humidified surroundings of 5% CO2 and 95% air. The cells were maintained while in the logarithmic phase of growth and were subcultured at two three day intervals. For storage, the cells had been frozen at 70 C liquid nitro gen in full medium supplemented pop over here with 5% di methyl sulfoxide as a cryoprotectant. Extraction of P. giganteus fruiting bodies The fresh fruiting bodies have been sliced, weighed and freeze dried for 1 two days.
The freeze dried fruiting bodies were then ground making use of a blender. The resulting dried powder was weighed and kept in 4 8 C. Aqueous extraction technique was in accordance to Eik et al. Briefly, the freeze dried powder was soaked in distilled water and was left overnight at space temperature and 200 rpm inside a shaker. The mix ture was double boiled in water bath for thirty min and fil tered soon after cooling. The resulting aqueous extract was freeze dried and kept at 40 C just before use. For ethanol extraction, the freeze dried powder was soaked in 95% ethanol at room temperature for 3 days and the procedure was repeated 3 instances. The ethanol solvent was evaporated implementing a rotary evaporator to give a brownish vis cous extract. Nutritional composition of freeze dried fruiting bodies of P. giganteus Fifty grams sample of P. giganteus fruiting bodies was sent to Consolidated Laboratory Sdn. Bhd. for nutri tional analysis. Cell viability and cytotoxicity assay Cell viability and proliferation was established by MTT assay.