Upregulation of CD271 transcriptome in breast cancer promotes cell survival via NFκB pathway

Molecular Biology Reports - Biological treatment of many cancers currently targets membrane bound receptors located on a cell surface. We are in a great to need identify novel membrane proteins...     

Novel Changes in NF-κB Activity during Progression and Regression Phases of Hyperplasia: ROLE OF MEK,ERK, AND p38*

Utilizing the Citrobacter rodentium-induced transmissible murine colonic hyperplasia (TMCH) model, we measured hyperplasia and NF-κB activation during progression (days 6 and 12 post-infection) and regression (days 20–34 post-infection) phases of TMCH. NF-κB activity increased at progression in conjunction with bacterial attachment and translocation to the colonic crypts and decreased 40% by day 20. NF-κB activity at days 27 and 34, however, remained 2–3-fold higher than uninfected control. Expression of the downstream target gene CXCL-1/KC in the crypts correlated with NF-κB activation kinetics. Phosphorylation of cellular IκBα kinase (IKK)α/β (Ser^176/180) was elevated during progression and regression of TMCH. Phosphorylation (Ser^32/36) and degradation of IκBα, however, contributed to NF-κB activation only from days 6 to 20 but not at later time points. Phosphorylation of MEK1/2 (Ser^217/221), ERK1/2 (Thr²⁰²/Tyr²⁰⁴), and p38 (Thr¹⁸⁰/Tyr¹⁸²) paralleled IKKα/β kinetics at days 6 and 12 without declining with regressing hyperplasia. siRNAs to MEK, ERK, and p38 significantly blocked NF-κB activity in vitro, whereas MEK1/2-inhibitor (PD98059) also blocked increases in MEK1/2, ERK1/2, and IKKα/β thereby inhibiting NF-κB activity in vivo. Cellular and nuclear levels of Ser⁵³⁶-phosphorylated (p65⁵³⁶) and Lys³¹⁰-acetylated p65 subunit accompanied functional NF-κB activation during TMCH. RSK-1 phosphorylation at Thr³⁵⁹/Ser³⁶³ in cellular/nuclear extracts and co-immunoprecipitation with cellular p65-NF-κB overlapped with p65⁵³⁶ kinetics. Dietary pectin (6%) blocked NF-κB activity by blocking increases in p65 abundance and nuclear translocation thereby down-regulating CXCL-1/KC expression in the crypts. Thus, NF-κB activation persisted despite the lack of bacterial attachment to colonic mucosa beyond peak hyperplasia. The MEK/ERK/p38 pathway therefore seems to modulate sustained activation of NF-κB in colonic crypts in response to C. rodentium infection.     

Human kallikrein 10, a predictive marker for breast cancer

Our laboratory is involved in identifying genes that can be used as early diagnostic or prognostic markers in breast cancer. We previously identified a gene (NES1) that is expressed in normal but not in transformed mammary epithelial cells (MECs). NES1 is located on chromosome 19q13.4 within the kallikrein locus and thus was designated as human kallikrein 10 (hK10), although we have been unable to detect any protease activity. Importantly, hK10 expression is decreased in a majority of breast cancer cell lines. Transfection of hK10 into hK10-negative breast cancer cells reduces the tumorigenicity. Using methylation-specific PCR and subsequent sequencing, we demonstrate a strong correlation between hypermethylation of hK10 and loss of mRNA expression. Further analysis showed that essentially 100% of normal breast specimens had hK10 expression, whereas 46% of ductal carcinoma in situ (DCIS) and the majority of infiltrating ductal carcinoma (IDC) samples lacked the hK10 mRNA. Importantly, hK10-negative DCIS diagnosed at the time of biopsy were subsequently diagnosed as IDC at the time of definitive surgery. It has been shown that hK10 protein expression is regulated by steroids. In addition to breast cancers, hK10 is downregulated in cervical cancer, prostate cancer and acute lymphocytic leukemia, whereas it is upregulated in ovarian cancers. These results point to the paradoxical role of hK10 in human cancers and underscore the importance of further studies of this kallikrein.     

Protective role of curcumin in ameliorating AFB1-induced apoptosis via mitochondrial pathway in liver cells

It is well documented that liver is the primary target organ of aflatoxin B₁ (AFB₁) and curcumin proved to be effective against AFB₁-induced liver injury. In the present study, we investigated the preventive effects of curcumin against AFB₁-induced apoptosis through the molecular regulation of p53, caspase-3, Bax, caspase-9, Bcl-2 and cytochrome-C associated with mitochondrial pathway. Liver antioxidant levels were measured. The hallmarks of apoptosis were analysed by methyl green-pyronin-Y staining, transmission electron microscopy, RT-PCR and western blot. Results revealed that dietary curcumin ameliorated AFB₁-induced oxidative stress in a dose-dependent manner. Methyl green-pyronin-Y staining and transmission electron microscopy showed that AFB₁ induced apoptosis and caused abnormal changes in liver cells morphology such as condensation of chromatin material, reduces cell volume and damaged mitochondria. Moreover, mRNA and protein expression results manifested that apoptosis associated genes showed up-regulation in AFB₁ fed group. However, the supplementation of dietary curcumin (dose-dependently) alleviated the increased expression of the apoptosis associated genes at mRNA and protein level, and restored the hepatocytes normal morphology. The study provides an insight and a better understanding of the preventive mechanism of curcumin against AFB₁-induced apoptosis in hepatocytes and provide scientific basis for the therapeutic uses of curcumin.

     

Spectral lights trigger biomass accumulation and production of antioxidant secondary metabolites in adventitious root cultures of Stevia rebaudiana (Bert.)

Stevia rebaudiana (S. rebaudiana) is the most important therapeutic plant species and has been accepted as such worldwide. It has a tendency to accumulate steviol glycosides, which are 300 times sweeter than marketable sugar. Recently, diabetic patients commonly use this plant as a sugar substitute for sweet taste. In the present study, the effects of different spectral lights were investigated on biomass accumulation and production of secondary metabolites in adventitious root cultures of S. rebaudiana. For callus development, leaf explants were excised from seed-derived plantlets and inoculated on a Murashige and Skoog (MS) medium containing the combination of 2,4-dichlorophenoxy acetic acid (2, 4-D, 2.0 mg/l) and 6-benzyladenine (BA, 2.0 mg/l), while 0.5 mg/l naphthalene acetic acid (NAA) was used for adventitious root culture. Adventitious root cultures were exposed to different spectral lights (blue, green, violet, red and yellow) for a 30-day period. White light was used as control. The growth kinetics was studied for 30 days with 3-day intervals. In this study, the violet light showed the maximum accumulation of fresh biomass (2.495 g/flask) as compared to control (1.63 g/flask), while red light showed growth inhibition (1.025 g/flask) as compared to control. The blue light enhanced the highest accumulation of phenolic content (TPC; 6.56 mg GAE/g DW), total phenolic production (TPP; 101 mg/flask) as compared to control (5.44 mg GAE/g DW; 82.2 mg GAE/g DW), and exhibited a strong correlation with dry biomass. Blue light also improved the accumulation of total flavonoid content (TFC; 4.33 mg RE/g DW) and total flavonoid production (TFP; 65 mg/flask) as compared to control. The violet light showed the highest DPPH inhibition (79.72%), while the lowest antioxidant activity was observed for control roots (73.81%). Hence, we concluded that the application of spectral lights is an auspicious strategy for the enhancement of the required antioxidant secondary metabolites in adventitious root cultures of S. rebaudiana and of other medicinal plants.     

Human sex hormone-binding globulin as a potential target of alternate plasticizers: an <Emphasis Type="Italic">in silico</Emphasis> study

Background

Currently, alternate plasticizers are used to replace phthalate plasticizers in children’s toys, medical equipments and food packaging, due to the adverse effects of phthalate compounds on human health and laws prohibiting their use. Current information regarding the safety and potential adverse effects of alternate plasticizers is limited and recent studies have found alternate plasticizers to display similar characteristics to those observed in phthalate plasticizers. This study was undertaken to evaluate and predict the potential endocrine disrupting activity of the three most commonly used alternate plasticizers: di(2-ethylhexyl)terephthalate (DEHT), tris(2-ethylhexyl)trimellitate (TOTM), and diisononyl hexahydrophthalate (DINCH) against human sex hormone-binding globulin (SHBG) using in silico approaches.

Materials and methods

The crystal structure of human SHBG (Id: 1D2S) was retrieved from Protein Data Bank. PubChem database was searched for the structures of alternate plasticizers, DEHT, TOTM, and DINCH. Docking was performed using Glide (Schrodinger) Induced Fit Docking module.

Results

Induced Fit Docking of three alternate plasticizer compounds indicated that each of the three compounds fitted well into the steroid binding pocket of SHBG. Docking displays showed interactions of alternate plasticizers with 25–30 amino-acid residues of SHBG; 18–20 amino residues overlapped between the natural ligand, DHT, and the three compounds (commonality of 82–91 %). The hydrogen-bonding interaction of the amino-acid residue, Asn-82, of SHBG was also present in displays of DHT and all the three alternate phthalates. The binding affinity of all the three alternate phthalates was higher than DHT; maximum in DINCH followed by TOTM and DEHT.

Conclusion

Our results suggested that the three alternate plasticizers have potential to engage the important interacting residues of SHBG and thus interfere in its steroid homeostatic function.

     

Review: role of carbon sources for in vitro plant growth and development

In vitro plant cells, tissues and organ cultures are not fully autotrophic establishing a need for carbohydrates in culture media to maintain the osmotic potential, as well as to serve as energy and carbon sources for developmental processes including shoot proliferation, root induction as well as emission, embryogenesis and organogenesis, which are highly energy demanding developmental processes in plant biology. A variety of carbon sources (both reducing and non-reducing) are used in culture media depending upon genotypes and specific stages of growth. However, sucrose is most widely used as a major transport-sugar in the phloem sap of many plants. In micropropagation systems, morphogenetic potential of plant tissues can greatly be manipulated by varying type and concentration of carbon sources. The present article reviews the past and current findings on carbon sources and their sustainable utilization for in vitro plant tissue culture to achieve better growth rate and development.     

Acetylation regulates subcellular localization of eukaryotic translation initiation factor 5A (eIF5A)

Eukaryotic translation initiation factor 5A (eIF5A) is a protein subject to hypusination, which is essential for its function. eIF5A is also acetylated, but the role of that modification is unknown. Here, we report that acetylation regulates the subcellular localization of eIF5A. We identified PCAF as the major cellular acetyltransferase of eIF5A, and HDAC6 and SIRT2 as its major deacetylases. Inhibition of the deacetylases or impaired hypusination increased acetylation of eIF5A, leading to nuclear accumulation. As eIF5A is constitutively hypusinated under physiological conditions, we suggest that reversible acetylation plays a major role in controlling the subcellular localization of eIF5A.