Understanding the regulation of B-catenin expression and activity in colorectal cancer carcinogenesis: beyond destruction complex

Aberrant Wnt/β-catenin signaling is central to colorectal cancer carcinogenesis. The well-known potential of targeting the canonical Wnt signaling pathway for the treatment of CRC is largely attributed to the ability of this pathway to regulate various cellular processes such as cell proliferation, metastasis, drug resistance, immune response, apoptosis, and cellular metabolism. However, with the current approach of targeting this pathway, none of the Wnt-targeted agents have been successfully implicated in clinical practice. Instead of using classical approaches to target this pathway, there is a growing need to find new and modified approaches to achieve the same. For this, a better understanding of the regulation of β-catenin, a major effector of the canonical Wnt pathway is a must. The present review addresses the importance of understanding the regulation of β-catenin beyond the destruction complex. Few recently discovered β-catenin regulators such as ZNF281, TTPAL, AGR2, ARHGAP25, TREM2, and TIPE1 showed significant potential in regulating the development of CRC through modulation of the Wnt/β-catenin signaling pathway in both in vitro and in vivo studies. Although the expression and activity of β-catenin is influenced by many protein regulators, the abovementioned proteins not only influence its expression and activation but are also directly involved in the development of CRC and various other solid tumors. Therefore, we hypothesise that focusing the current research on finding the detailed mechanism of action of these regulators may assist in providing with a better treatment approach or improve the current therapeutic regimens (AU)

Understanding the regulation of ß-catenin expression and activity in colorectal cancer carcinogenesis: beyond destruction complex.

Aberrant Wnt/ß-catenin signaling is central to colorectal cancer carcinogenesis. The well-known potential of targeting the canonical Wnt signaling pathway for the treatment of CRC is largely attributed to the ability of this pathway to regulate various cellular processes such as cell proliferation, metastasis, drug resistance, immune response, apoptosis, and cellular metabolism. However, with the current approach of targeting this pathway, none of the Wnt-targeted agents have been successfully implicated in clinical practice. Instead of using classical approaches to target this pathway, there is a growing need to find new and modified approaches to achieve the same. For this, a better understanding of the regulation of ß-catenin, a major effector of the canonical Wnt pathway is a must. The present review addresses the importance of understanding the regulation of ß-catenin beyond the destruction complex. Few recently discovered ß-catenin regulators such as ZNF281, TTPAL, AGR2, ARHGAP25, TREM2, and TIPE1 showed significant potential in regulating the development of CRC through modulation of the Wnt/ß-catenin signaling pathway in both in vitro and in vivo studies. Although the expression and activity of ß-catenin is influenced by many protein regulators, the abovementioned proteins not only influence its expression and activation but are also directly involved in the development of CRC and various other solid tumors. Therefore, we hypothesise that focusing the current research on finding the detailed mechanism of action of these regulators may assist in providing with a better treatment approach or improve the current therapeutic regimens.

Study of twenty preparations of human albumin solution which failed in quality control testing due to elevated sodium content, a poor internal quality control at manufacturing unit.

Current study is conducted in our laboratory due to failure in quality control testing of twenty batches of Human Albumin solution in which sodium content is higher than the prescribed limit. These batches are received in short duration from indigenous manufacturer and is the first incident of failure of Human albumin preparation in sodium content of manufacturer. On request of manufacturer, study is conducted to rule out the cause. Repeat testing of each out of specification batch is conducted and a trend analysis is drawn between our findings and manufacturer's results, also study of trend analysis of manufacturer for the last one year. Trend analysis data indicated towards poor consistency of batches with major shift at various time intervals in sodium content of human albumin preparation. Further analysis rule out that non-traceable quality of standard used in the internal quality control testing by manufacturer is the root cause of the problem.

Ras signaling pathway in the chemopreventive action of different ratios of fish oil and corn oil in experimentally induced colon carcinogenesis.

Dietary factors play a significant role in colon cancer. The essential polyunsaturated fatty acids (PUFAs), n-3 PUFAs, and n-6 PUFAs exert inverse effect on cancer. This study was designed to understand the mechanism of chemopreventive action of different ratios of fish oil (FO) and corn oil (CO) in colon carcinoma. Wistar rats were divided into 3 groups: Group 1 received purified diet whereas Groups 2 and 3 received modified diet with FO:CO (1:1) and FO:CO (2.5:1), respectively. The groups were further subdivided into controls receiving ethylenediamine-tetra acetic-acid and treated groups received dimethylhydrazine-dihydrochloride (DMH)/wk for 4 wk. Animals sacrificed 48 h after last injection constituted initiation phase and that sacrificed after 16 wk constituted post-initiation phase. Differential effect of different ratios of FO and CO was analyzed in isolated colonocytes. In both phases, DMH treatment showed an increase in pan Ras, Raf, MEK1/2, extracellular signal regulated kinase (Erk)1/2, and c-fos levels. Akt levels were increased in post-initiation phase only. Treatment with FO + CO (1:1) + DMH decreased pan Ras, MEK1/2 and Erk1/2 levels in post-initiation phase whereas Raf and c-fos were decreased in both phases. Treatment with FO + CO (2.5:1) + DMH decreased Ras, Raf, MEK1/2, Erk1/2, and c-fos levels in both phases. Akt was decreased in post-initiation phase only. The chemo-preventive action of FO and CO may be mediated by time- and dose-dependent effect.

Bystander effects of ionizing radiation can be modulated by signaling amines.

Actual risk and risk management of exposure to ionizing radiation are among the most controversial areas in environmental health protection. Recent developments in radiobiology especially characterization of bystander effects have called into question established dogmas and are thought to cast doubt on the scientific basis of the risk assessment framework, leading to uncertainty for regulators and concern among affected populations. In this paper we test the hypothesis that small signaling molecules widely used throughout the animal kingdom for signaling stress or environmental change, such as 5-Hydroxytryptamine (5-HT, serotonin), l-DOPA, glycine or nicotine are involved in bystander signaling processes following ionizing radiation exposure. We report data which suggest that nano to micromolar concentrations of these agents can modulate bystander-induced cell death. Depletion of 5-HT present in tissue culture medium, occurred following irradiation of cells. This suggested that 5-HT might be bound by membrane receptors after irradiation. Expression of 5-HT type 3 receptors which are Ca(2+) ion channels was confirmed in the cells using immunocytochemistry and receptor expression could be increased using radiation or 5-HT exposure. Zofran and Kitryl, inhibitors of 5-HT type 3 receptors, and reserpine a generic serotonin antagonist block the bystander effect induced by radiation or by serotonin. The results may be important for the mechanistic understanding of how low doses of radiation interact with cells to produce biological effects.

Communication of radiation-induced stress or bystander signals between fish in vivo.

We report data in this paper suggesting that fish irradiated to 0.5 Gy total body dose can release factors into the water that signal other unexposed fish and cause induction of bystander effects expressed as increased cell death in a reporter system. Radiation-induced bystander effects, resulting in the appearance of radiation damage or induction of typical radiation responses in unirradiated cells and tissues are now an established consequence of exposure to low doses of ionizing radiation, however little work has been done in vivo or in species other than humans or mice. In these experiments rainbow trout were irradiated and then paired with unirradiated fish for two hours. Additionally, unirradiated fish were placed in water which had previously been used to hold irradiated fish for 2 h. Sham-irradiated fish and absolute control fish were also examined all using blind protocols. Following a two h incubation period, at these various exposure regimes, the fish were killed by a blow to the head and dissected. Five organs were removed from each fish and tissue explants were cultured using an established technique. After 2 days, the culture medium was harvested and used in a reporter assay to determine whether a bystander effect had been induced. The explants were cultured on in Clonetics growth medium for a further 14 days then fixed for assay of radiation response proteins. The responses varied according to the cell type in the original explants, with the gill and fin showing the most pronounced response. The results suggest that communication signals leading to a typical radiation response can be passed between fish and seem to involve secretion of a chemical messenger into the water.

Aerobic bacterial isolates from burn wound infections and their antibiograms--a five-year study.

A retrospective study of major aerobic bacterial isolates from pus/wound swabs taken from patients admitted to the burn unit at Govt. Medical College Hospital, Chandigarh, India, over a period of 5 years (June 1997-May 2002) was undertaken. The study was carried out to determine the bacterial profile and antimicrobial susceptibility of the isolates and to describe the change in trends over the study period. The pus/wound swabs yielded very high culture positivity (96%) for 665 total isolates. Pseudomonas aeruginosa was found to be most common isolate (59%) followed by Staphylococcus aureus (17.9%), Acinetobacter spp. (7.2%), Klebsiella spp. (3.9%), Enterobacter spp. (3.9%), Proteus spp. (3.3%) and others (4.8%). Although P. aeruginosa continued to remain the predominant isolate over the five years, a constant and significant increase in the incidence of Acinetobacter spp. was found. Amikacin was found to be the most effective drug against gram negative bacteria, however, resistance to it was significantly increased over 5 years. For S. aureus and P. aeruginosa netilmicin and piperacillin were found to be the most effective drugs. Most of the isolates showed high level resistance to antimicrobial agents.

Modulation of acid secretion in common bile duct ligation-related gastropathy in Wistar rats.

BACKGROUND: Portal hypertensive gastropathy is associated with fundic gland atrophy, resulting in a decrease in chief and parietal cells, and diminished acid secretion. METHODS: Acid secretion by isolated parietal cells was measured (acridine orange retention), along with the levels of various second messengers (intracellular Ca(2+), cyclic adenosine monophosphate and protein kinase C) in the common bile duct, ligated portal hypertensive rats and compared with sham-operated controls. RESULTS: There was a significant decrease in the response of isolated parietal cells to the secretagogues histamine and carbachol. This resulted in the blunted acid secretion in the common bile duct ligated group. In addition, all the second messengers studied were significantly decreased as compared with the sham-operated controls. CONCLUSION: These results suggest that the blunted acid secretory response in the portal hypertensive rat is caused by an alteration in the intracellular signal transduction mechanism.

Activity of brush border membrane enzymes in proximal jejunum of portal hypertensive rats.

PURPOSE: Malabsorption accompanies portal hypertension, especially when associated with chronic liver disease. The development of portal hypertension is accompanied by significant alterations in the splanchnic microcirculation. In this study, the effect of extrahepatic and intrahepatic portal hypertension on brush border membrane enzymes was estimated. METHODS: Portal hypertension was induced in rats by partial portal vein ligation (PPVL) (n = 6) and common bile duct ligation (CBDL) (n = 6), and the activity of sucrase, lactase, alkaline phosphatase, and leucine aminopeptidase (LAP) in the intestinal homogenate was measured. RESULTS: Intrasplenic pulp pressure (ISPP) (in cm of saline) was found to be elevated in PPVL (21.3 +/- 1.47) and CBDL animals (21.5 +/- 1.79) as compared with findings in their respective sham-operated controls (12.74 +/- 0.86, 11.83 +/- 1.04). Only sucrase and LAP activity was significantly elevated (P < 0.05) in the PPVL group. No changes were observed in the CBDL group. CONCLUSION: Only sucrase and LAP activities were increased in PPVL rats.

Simultaneous determination of gallium(III) and indium(III) by derivative spectrophotometry.

A simple, selective and sensitive derivative spectrophotometric method is proposed for the simultaneous determination of gallium(III) and indium(III) in mixtures using 1-(2-pyridylazo)-2-naphthol in cationic micellar medium, without any prior separation. Beer's law is obeyed between 2.80x10(-1)-3.63 and 4.60x10(-1)-9.20 mug ml(-1) concentration of Ga(III) and In(III) at 550 and 542 nm, the isodifferential points of indium and gallium complexes in the first-order derivative mode, respectively. The proposed method is successfully applied for the determination of gallium and indium in standard reference materials and synthetic binary mixtures with a relative error of +/-2.07 and +/-2.55%, respectively.

Inhibition of Helicobacter pylori adherence by a peptide derived from neuraminyl lactose binding adhesin.

Helicobacterpylori, like many other gut colonizing bacteria, binds to sialic acid rich macromolecules present on the gastric epithelium. NLBH (neuraminyl lactose binding haemagglutinin) a 32 kDa adhesin located on the surface of H. pylori has been shown to have specific affinity towards NeuAcalpha2,3Galbeta1,4Gluc(3'SL). This sialic acid moiety is over-expressed in an atrophic stomach undergoing parietal cell depletion. Antibodies against a lysine rich peptide fragment of NLBH inhibit agglutination of human erythrocytes. This lysine rich sequence from NLBH was proposed to be the receptor-binding site. In order to elucidate the binding of NLBH to gastric epithelium, a peptide (D-P-K-R-T-I-Q-K-K-S) was synthesized. A series of experiments were performed involving adherence inhibition assays, 2D-NMR, molecular modelling and measurement of modulation in acid secretion. Results indicated that the peptide fragment could be involved in receptor recognition, which is important for the binding of H. pylori to gastric epithelium. The binding is possibly through hydrogen bonding. Two lysines and a threonine residue seem to be within the hydrogen bonding distance of NeuAcalpha2,3Galbeta1,4Gluc. Further, in vitro assays were performed to evaluate the role of the peptide on acid secretion by parietal cells isolated from human fundal biopsies. Interestingly, the peptide increases acid secretion only in H. pylori negative and in treated patients but not in H. pylori positive patients. This highlights the role of NLBH in acid secretion and could be of some consequence in the prognosis of the disease.

Micro-perfusion flow cell for imaging cultured cells.

We present a unique design for a flow cell with a small working volume that allows rapid displacement of media viewed under high power and short working distance objectives. The flow cell has a small internal depth (ca. 0.033 cm) and volume (ca. 0.05 mL) and is easy to handle. Made of Delrin, the flow cell is biologically inert. We have used the flow cell for fluorescence imaging of PC12 cells loaded with tetramethylrhodamine dextran (TMRD) and other dyes.

Non-extractive derivative spectrophotometric determination of cobalt in neutral micellar medium.

A sensitive derivative spectrophotometric method using 1-nitroso-2-naphthol has been developed for determination of trace amounts of cobalt in the presence of a neutral surfactant. Photometric parameters, viz., lambda(max), molar absorption coefficient and analytical sensitivity of the complex formed in micellar media are 420 nm, 3.18x10(4) l mol(-1) cm(-1) and 2.05 ng ml(-1), respectively. Beer's law holds from 0.20 to 3.0 mug ml(-1) of the analyte concentration. The method has a high sensitivity with a detection limit of 1.68 ng ml(-1). A selective determination of cobalt in presence of copper(II) or iron(III) using derivative spectral profiles and without any masking or pre-separation is also reported. Samples of drugs and standard alloys analysed by the proposed method yielded results comparable to those obtained using recommended procedures.

Morphological changes associated with long-term potentiation.

Long-term potentiation (LTP) is a long-lasting form of synaptic plasticity induced by brief repetitive afferent stimulation that is thought to be associated with learning and memory. It is most commonly studied in the hippocampus where it may last for several weeks, and involves the synthesis of new proteins that might play a structural role. In this review we summarize the evidence in favor of modifications of neuronal architecture during LTP. We focus our attention on changes occurring at the level of single synapses, including components of postsynaptic dendrites (dendritic spines, the postsynaptic density, and synaptic curvature), of presynaptic terminals, and the formation of new synapses. We conclude that although many morphological changes at various sites have been observed during LTP, there is no definitive proof in favor of structural changes associated with LTP. However, morphological modifications remain a valid candidate for mechanisms of learning and memory.

Characterization of lymphocytic subsets and cytokine production in gastric biopsy samples from Helicobacter pylori patients.

BACKGROUND: This study characterized the phenotypic subsets of isolated gastric lymphocytes and the cellular immune response in cultured gastric biopsy specimens. METHODS: Endoscopy specimens from 40 Helicobacter pylori-positive and 40 H. pylori-negative patients were studied. a) Isolated gastric lymphocytes were analysed for CD4+, CD8+ T-lymphocyte subsets, activated T cells, and natural killer cells on a fluorescence-activated cell sorter, using monoclonal antibodies. b) The supernatant of cultured gastric biopsy specimens were assayed for interleukin (IL)-2, IL-4, and IL-6 levels. RESULTS: In H. pylori-positive patients there was (a) a decrease in CD4+/CD8+ T cells, no change in activated T cells, and an increase in natural killer cells, and (b) no change in IL-2 levels and a significant increase in IL-4 and IL-6 levels. CONCLUSIONS: There is an increase in CD8+ lymphocytes and natural killer cells, and the observed increase in IL-4 and IL-6 might be important in H. pylori-associated gastritis.

Extrahepatic portal hypertensive gastropathy in Wistar rats: modulation of acid secretion in isolated parietal cells.

Stimulated acid secretion in portal hypertensive gastropathy is blunted and could be due to defective signal transduction in the parietal cell. Therefore, an attempt was made to study the levels of second messengers in parietal cells in experimental extrahepatic portal hypertensive gastropathy. Our aim was to measure acid secretion, intracellular free calcium, calcium transport, cyclic AMP, and ATP levels in the parietal cells isolated from the gastric mucosa of portal hypertensive rats. Acid secretion using acridine orange, intracellular free calcium using Fura-2/AM, calcium influx and efflux by 45CaCl2 and cyclic AMP by RIA kits were measured in unstimulated and histamine- and carbachol-stimulated isolated parietal cells in rats with partial portal vein ligation and sham operation. ATP was measured by HPLC. In portal hypertensive gastropathy, stimulated acid secretion was blunted, and there was a decrease in basal intracellular free calcium. Calcium influx and efflux were at a higher level, and there was a decrease in elevation of intracellular free calcium and cyclic AMP levels with secretagogues. There was also a decrease in ATP. In conclusion, there exists a low energy state in addition to multiple aberrations at the second messenger level in parietal cells in portal hypertensive gastropathy.