Computed Tomography-Based Angiographic Evaluation of Graft Patency Rate After Coronary Artery Bypass Graft Surgery in Bangladesh.

Background This study aims to assess the graft patency rate following coronary artery bypass graft (CABG) surgery using noninvasive CT angiography.  Materials and methods A total of 68 patients were retrospectively evaluated with CT angiography (group I: 34 patients with coronary endarterectomy (CE) and group II: 34 patients without CE). CE was performed in multi-segmental diffuse coronary artery disease (CAD) or when calcified or extremely thick plaques made anastomosis troublesome. A team of two experts, an interventional radiologist and a cardiac surgeon, did the evaluations of graft patency rate. Results A total of 205 bypass grafts were evaluated in 68 post-CABG status patients (110 grafts in group I and 95 grafts in group II; moreover, 82 were arterial and 123 were venous grafts). Post CABG, CT angiography demonstrated a graft patency rate of about 90% in both study groups at five years follow up, which was statistically insignificant (P > 0.05) in terms of graft patency rate. Following CE, five-year angina-free survival rates were 89% and 91% in groups I and II, respectively. Conclusion CABG surgery with endarterectomy is reliable and effective. It achieves the desired surgical myocardial revascularization in patients with diffuse calcified CAD having no alternative options for adequate myocardial revascularization.

High-throughput FTIR-based bioprocess analysis of recombinant cyprosin production.

To increase the knowledge of the recombinant cyprosin production process in Saccharomyces cerevisiae cultures, it is relevant to implement efficient bioprocess monitoring techniques. The present work focuses on the implementation of a mid-infrared (MIR) spectroscopy-based tool for monitoring the recombinant culture in a rapid, economic, and high-throughput (using a microplate system) mode. Multivariate data analysis on the MIR spectra of culture samples was conducted. Principal component analysis (PCA) enabled capturing the general metabolic status of the yeast cells, as replicated samples appear grouped together in the score plot and groups of culture samples according to the main growth phase can be clearly distinguished. The PCA-loading vectors also revealed spectral regions, and the corresponding chemical functional groups and biomolecules that mostly contributed for the cell biomolecular fingerprint associated with the culture growth phase. These data were corroborated by the analysis of the samples' second derivative spectra. Partial least square (PLS) regression models built based on the MIR spectra showed high predictive ability for estimating the bioprocess critical variables: biomass (R 2 = 0.99, RMSEP 2.8%); cyprosin activity (R 2 = 0.98, RMSEP 3.9%); glucose (R 2 = 0.93, RMSECV 7.2%); galactose (R 2 = 0.97, RMSEP 4.6%); ethanol (R 2 = 0.97, RMSEP 5.3%); and acetate (R 2 = 0.95, RMSEP 7.0%). In conclusion, high-throughput MIR spectroscopy and multivariate data analysis were effective in identifying the main growth phases and specific cyprosin production phases along the yeast culture as well as in quantifying the critical variables of the process. This knowledge will promote future process optimization and control the recombinant cyprosin bioprocess according to Quality by Design framework.

Metabolic profiling of recombinant Escherichia coli cultivations based on high-throughput FT-MIR spectroscopic analysis.

Escherichia coli is one of the most used host microorganism for the production of recombinant products, such as heterologous proteins and plasmids. However, genetic, physiological and environmental factors influence the plasmid replication and cloned gene expression in a highly complex way. To control and optimize the recombinant expression system performance, it is very important to understand this complexity. Therefore, the development of rapid, highly sensitive and economic analytical methodologies, which enable the simultaneous characterization of the heterologous product synthesis and physiologic cell behavior under a variety of culture conditions, is highly desirable. For that, the metabolic profile of recombinant E. coli cultures producing the pVAX-lacZ plasmid model was analyzed by rapid, economic and high-throughput Fourier Transform Mid-Infrared (FT-MIR) spectroscopy. The main goal of the present work is to show as the simultaneous multivariate data analysis by principal component analysis (PCA) and direct spectral analysis could represent a very interesting tool to monitor E. coli culture processes and acquire relevant information according to current quality regulatory guidelines. While PCA allowed capturing the energetic metabolic state of the cell, e.g. by identifying different C-sources consumption phases, direct FT-MIR spectral analysis allowed obtaining valuable biochemical and metabolic information along the cell culture, e.g. lipids, RNA, protein synthesis and turnover metabolism. The information achieved by spectral multivariate data and direct spectral analyses complement each other and may contribute to understand the complex interrelationships between the recombinant cell metabolism and the bioprocess environment towards more economic and robust processes design according to Quality by Design framework. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:285-298, 2017.

Monitoring the ex-vivo expansion of human mesenchymal stem/stromal cells in xeno-free microcarrier-based reactor systems by MIR spectroscopy.

Human mesenchymal stem/stromal cells (MSCs) have received considerable attention in the field of cell-based therapies due to their high differentiation potential and ability to modulate immune responses. However, since these cells can only be isolated in very low quantities, successful realization of these therapies requires MSCs ex-vivo expansion to achieve relevant cell doses. The metabolic activity is one of the parameters often monitored during MSCs cultivation by using expensive multi-analytical methods, some of them time-consuming. The present work evaluates the use of mid-infrared (MIR) spectroscopy, through rapid and economic high-throughput analyses associated to multivariate data analysis, to monitor three different MSCs cultivation runs conducted in spinner flasks, under xeno-free culture conditions, which differ in the type of microcarriers used and the culture feeding strategy applied. After evaluating diverse spectral preprocessing techniques, the optimized partial least square (PLS) regression models based on the MIR spectra to estimate the glucose, lactate and ammonia concentrations yielded high coefficients of determination (R(2) ≥ 0.98, ≥0.98, and ≥0.94, respectively) and low prediction errors (RMSECV ≤ 4.7%, ≤4.4% and ≤5.7%, respectively). Besides PLS models valid for specific expansion protocols, a robust model simultaneously valid for the three processes was also built for predicting glucose, lactate and ammonia, yielding a R(2) of 0.95, 0.97 and 0.86, and a RMSECV of 0.33, 0.57, and 0.09 mM, respectively. Therefore, MIR spectroscopy combined with multivariate data analysis represents a promising tool for both optimization and control of MSCs expansion processes. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:447-455, 2016.

A comprehensive high-throughput FTIR spectroscopy-based method for evaluating the transfection event: estimating the transfection efficiency and extracting associated metabolic responses.

Reporter genes are routinely used in every laboratory for molecular and cellular biology for studying heterologous gene expression and general cellular biological mechanisms, such as transfection processes. Although well characterized and broadly implemented, reporter genes present serious limitations, either by involving time-consuming procedures or by presenting possible side effects on the expression of the heterologous gene or even in the general cellular metabolism. Fourier transform mid-infrared (FT-MIR) spectroscopy was evaluated to simultaneously analyze in a rapid (minutes) and high-throughput mode (using 96-wells microplates), the transfection efficiency, and the effect of the transfection process on the host cell biochemical composition and metabolism. Semi-adherent HEK and adherent AGS cell lines, transfected with the plasmid pVAX-GFP using Lipofectamine, were used as model systems. Good partial least squares (PLS) models were built to estimate the transfection efficiency, either considering each cell line independently (R (2) ≥ 0.92; RMSECV ≤ 2 %) or simultaneously considering both cell lines (R (2) = 0.90; RMSECV = 2 %). Additionally, the effect of the transfection process on the HEK cell biochemical and metabolic features could be evaluated directly from the FT-IR spectra. Due to the high sensitivity of the technique, it was also possible to discriminate the effect of the transfection process from the transfection reagent on KEK cells, e.g., by the analysis of spectral biomarkers and biochemical and metabolic features. The present results are far beyond what any reporter gene assay or other specific probe can offer for these purposes.

In situ near-infrared (NIR) versus high-throughput mid-infrared (MIR) spectroscopy to monitor biopharmaceutical production.

The development of biopharmaceutical manufacturing processes presents critical constraints, with the major constraint being that living cells synthesize these molecules, presenting inherent behavior variability due to their high sensitivity to small fluctuations in the cultivation environment. To speed up the development process and to control this critical manufacturing step, it is relevant to develop high-throughput and in situ monitoring techniques, respectively. Here, high-throughput mid-infrared (MIR) spectral analysis of dehydrated cell pellets and in situ near-infrared (NIR) spectral analysis of the whole culture broth were compared to monitor plasmid production in recombinant Escherichia coli cultures. Good partial least squares (PLS) regression models were built, either based on MIR or NIR spectral data, yielding high coefficients of determination (R(2)) and low predictive errors (root mean square error, or RMSE) to estimate host cell growth, plasmid production, carbon source consumption (glucose and glycerol), and by-product acetate production and consumption. The predictive errors for biomass, plasmid, glucose, glycerol, and acetate based on MIR data were 0.7 g/L, 9 mg/L, 0.3 g/L, 0.4 g/L, and 0.4 g/L, respectively, whereas for NIR data the predictive errors obtained were 0.4 g/L, 8 mg/L, 0.3 g/L, 0.2 g/L, and 0.4 g/L, respectively. The models obtained are robust as they are valid for cultivations conducted with different media compositions and with different cultivation strategies (batch and fed-batch). Besides being conducted in situ with a sterilized fiber optic probe, NIR spectroscopy allows building PLS models for estimating plasmid, glucose, and acetate that are as accurate as those obtained from the high-throughput MIR setup, and better models for estimating biomass and glycerol, yielding a decrease in 57 and 50% of the RMSE, respectively, compared to the MIR setup. However, MIR spectroscopy could be a valid alternative in the case of optimization protocols, due to possible space constraints or high costs associated with the use of multi-fiber optic probes for multi-bioreactors. In this case, MIR could be conducted in a high-throughput manner, analyzing hundreds of culture samples in a rapid and automatic mode.

In situ near infrared spectroscopy monitoring of cyprosin production by recombinant Saccharomyces cerevisiae strains.

Near infrared (NIR) spectroscopy was used to in situ monitoring the cultivation of two recombinant Saccharomyces cerevisiae strains producing heterologous cyprosin B. NIR spectroscopy is a fast and non-destructive technique, that by being based on overtones and combinations of molecular vibrations requires chemometrics tools, such as partial least squares (PLS) regression models, to extract quantitative information concerning the variables of interest from the spectral data. In the present work, good PLS calibration models based on specific regions of the NIR spectral data were built for estimating the critical variables of the cyprosin production process: biomass concentration, cyprosin activity, cyprosin specific activity, the carbon sources glucose and galactose concentration and the by-products acetic acid and ethanol concentration. The PLS models developed are valid for both recombinant S. cerevisiae strains, presenting distinct cyprosin production capacities, and therefore can be used, not only for the real-time control of both processes, but also in optimization protocols. The PLS model for biomass yielded a R(2)=0.98 and a RMSEP=0.46 g dcw l(-1), representing an error of 4% for a calibration range between 0.44 and 13.75 g dcw l(-1). A R(2)=0.94 and a RMSEP=167 Um l(-1) were obtained for the cyprosin activity, corresponding to an error of 6.7% of the experimental data range (0-2509 Um l(-1)), whereas a R(2)=0.93 and RMSEP=672 U mg(-1) were obtained for the cyprosin specific activity, corresponding to an error of 7% of the experimental data range (0-11,690 Um g(-1)). For the carbon sources glucose and galactose, a R(2)=0.96 and a RMSECV of 1.26 and 0.55 g l(-1), respectively, were obtained, showing high predictive capabilities within the range of 0-20 g l(-1). For the metabolites resulting from the cell growth, the PLS model for acetate was characterized by a R(2)=0.92 and a RMSEP=0.06 g l (-1), which corresponds to a 6.1% error within the range of 0.41-1.23 g l(-1); for the ethanol, a high accuracy PLS model with a R(2)=0.97 and a RMSEP=1.08 g l(-1) was obtained, representing an error of 9% within the range of 0.18-21.76 g l(-1). The present study shows that it is possible the in situ monitoring and prediction of the critical variables of the recombinant cyprosin B production process by NIR spectroscopy, which can be applied in process control in real-time and in optimization protocols. From the above, NIR spectroscopy appears as a valuable analytical tool for online monitoring of cultivation processes, in a fast, accurate and reproducible operation mode.

Cyclooxygenase/lipoxygenase shunting lowers the anti-cancer effect of cyclooxygenase-2 inhibition in colorectal cancer cells.

BACKGROUND: Arachidonic acid metabolite, generated by cyclooxygenase (COX), is implicated in the colorectal cancer (CRC) pathogenesis. Inhibiting COX may therefore have anti-carcinogenic effects. Results from use of non-steroidal anti-inflammatory drugs inhibiting only COX have been conflicting. It has been postulated that this might result from the shunting of arachidonic acid metabolism to the 5-lipoxygenase (5-LOX) pathway. Cancer cell viability is promoted by 5-LOX through several mechanisms that are similar to those of cyclooxygenase-2 (COX-2). Expression of 5-LOX is upregulated in colorectal adenoma and cancer. The aim of this study was to investigate the shunting of arachidonic acid metabolism to the 5-LOX pathway by cyclooxygenase inhibition and to determine if this process antagonizes the anti-cancer effect in colorectal cancer cells. METHODS: Three colorectal cancer cell lines (HCA7, HT-29 & LoVo) expressing 5-LOX and different levels of COX-2 expression were used. The effects of aspirin (a non-selective COX inhibitor) and rofecoxib (COX-2 selective) on prostaglandin E2 (PGE2) and leukotriene B4 (LTB4) secretion were quantified by ELISA. Proliferation and viability were studied by quantifying double-stranded DNA (dsDNA) content and metabolic activity. Apoptosis was determined by annexin V and propidium iodide staining using confocal microscopy, and caspase-3/7 activity by fluorescent substrate assay. RESULTS: COX inhibitors suppressed PGE2 production but enhanced LTB4 secretion in COX-2 expressing cell lines (P <0.001). The level of COX-2 expression in colorectal cancer cells did not significantly influence the anti-proliferative and pro-apoptotic effects of COX inhibitors due to the shunting mechanism. CONCLUSIONS: This study provides evidence of shunting between COX and 5-LOX pathways in the presence of unilateral inhibition, and may explain the conflicting anti-carcinogenic effects reported with use of COX inhibitors.

A novel method for the extraction and culture of progenitor stem cells from human peripheral blood for use in regenerative medicine.

Human peripheral blood (HPB) contains both circulating endothelial cells (CECs) and endothelial progenitor stem cells (EPCs), which may be suitable for use in regenerative medicine. There has been considerable interest in using these cells, but there is no "gold standard" technique for isolating these cells. The aim of this study was to characterize and compare a number of different extraction and culture techniques to develop a system to isolate and culture cells. EPC and CEC were isolated from HPB using either Histopaque-1077 or Lymphoprep. The two isolation methods were compared for the number of cells isolated, cell metabolism, and RNA expression. Both isolations produced viable cells and were comparable. The tissue culture method employed does have a significant effect on the cell population with regard to medium choice, fetal bovine serum concentration, and surface modification of the culture surface. In conclusion, it can be seen that although this study and previous work can suggest a basis for culture, further work to develop an optimized and agreed "gold standard" culture regime for EPC from HPB is required to maximize the potential of this source of cells for regenerative medicine and to translate its clinical use in the future.

The long-term stability in gene expression of human endothelial cells permits the production of large numbers of cells suitable for use in regenerative medicine.

Tissue engineering has been conducted in the study of cardiovascular grafts for many years. Many obstacles have been overcome in this rapidly changing field, but one difficulty has remained until now: the large number of endothelial cells (ECs) needed for seeding the inner layer of bypass graft. Recent advances in endothelial progenitor cell (EPC) isolation and culture techniques have increased the interest in genetic studies. Despite these advances in EPC studies, the "gold standard" for the seeding of tissue engineering constructs or hybrid grafts remains mature human umbilical vein endothelial cells (HUVECs). This study investigates the ability of HUVECs to be expanded in culture to provide sufficient cells for graft seeding. The levels of gene expression of key genes are then examined to ensure that these cells retain the EC phenotype. This study demonstrates that HUVECs may be cultured for up to 12 passages without alteration in phenotype. Subsequent passage numbers are sufficiently similar to those preceding them to allow cells of different passages to be mixed without gene expression anomalies.

Inhibition of the p38 MAPK pathway sensitises human colon cancer cells to 5-fluorouracil treatment.

Colorectal cancer is the third most common cause of cancer-related deaths in the Western world. 5-Fluorouracil (5-FU) based chemotherapeutic regimes have been the mainstay of systemic treatment for disseminated colorectal cancer for many years. However, it only produces a 25% response rate due to the drug-resistance. The mitogen-activated protein kinase (MAPK) pathway is involved in the anti-apoptotic process; its activation provides cancer cells with a survival advantage to escape the apoptotic challenge. This study assessed whether the p38 MAPK pathway is involved in 5-FU resistance in colorectal cancer cells. 5-FU only or 5-FU combined with a p38 MAPK pathway inhibitor (SB203580) was used to treat 5-FU-resistant colorectal cancer cells. The effect of the treatment on cell viability, death and caspase activities was assessed. Western blotting was used to investigate the responses of apoptosis-related proteins following the treatment. Results showed that p38 MAPK inhibitor significantly increased colorectal cancer cell sensitivity to 5-FU. SB203580 in combination with 5-FU significantly reduced cell viability (P<0.01), and increased cell death and cellular caspase activity (P<0.01). Western blotting data revealed that SB203580 sensitises cancer cells to 5-FU due to an increase in Bax expression. These findings suggest that p38 MAPK is involved in cancer cell survival, and that the inhibition of p38 MAPK can enhance 5-FU to kill colorectal cancer cells.

Haemodynamic regulation of gene expression in vascular tissue engineering.

Synthetic grafts, namely expanded polytetrafluoroethlene (ePTFE) and poly(ethylene terephthalate) (Dacron), used for cardiovascular bypass surgery are thrombogenic. Lining the inner lumen ("seeding") of synthetic grafts with endothelial cells (ECs) increases patency rates similar to those of autologous grafts (e.g. saphenous vein). The major drawback with seeding grafts is the retention of cells present on the graft after implantation in vivo, where large portions of cell wash off. Preconditioning the seeded EC monolayer with shear stress has been shown to promote the reorganisation of the EC cytoskeleton and production of extracellular matrix, resulting in higher EC retention after exposure to blood flow. Vascular ECs have a number of essential and complex roles. ECs synthesise and secrete vasoconstrictors, vasodilators, growth factors, fibrinolytic factors, cytokines, adhesion molecules, matrix proteins and mitogens that modulate many physiological processes such as wound healing, hemostasis, vascular remodelling, inflammatory and immune responses. Vascular cells in vivo are exposed to hemodynamic forces created by the pulsatile flow of blood through the vessel. Due to their unique anatomical position, ECs are constantly exposed to shear stress forces and allow the vessel wall to adapt to changes by modulating EC structure and function. This review describes the mainly in vitro and in vivo studies used to define the molecular role hemodynamics have in vascular disease and its usage in developing tissue engineered vascular bypass grafts.

Doxycycline in mitochondrial mediated pathway of apoptosis: a systematic review.

Tetracyclines have been long known for their antimicrobial role. They are one of the most widely used antibiotics in clinical practice since last 5 decades. Recently their role as matrix metalloproteinase inhibitor and in apoptosis has widely attracted attention in biological field. Of them, doxycycline is one with long duration of actions and has recently been shown to have various anti-cancer properties, especially cytotoxic and anti-proliferative activities. Here, we systematically reviewed the role of doxycycline in the mitochondrial mediated apoptosis in various tissues. MEDLINE and EMBASE databases were searched using a formal search strategy with definite inclusion and exclusion criteria. Data extraction was performed for each included study using a custom designed data extraction form. A total of 81 references were identified through MEDLINE and 5 were identified through EMBASE. 74 references from MEDLINE and all 5 in EMBASE were excluded through reading titles, abstracts and full text. In total, 7 studies fulfilled inclusion criteria. Following systematic review of these studies, we concluded that doxycycline induces apoptosis through mitochondrial mediated pathway in different tissue cells however it may be cell specific. The caspase independent apoptosis as one of the mechanisms of actions of doxycycline needs further studies for better understanding.

Lowering the apoptotic threshold in colorectal cancer cells by targeting mitochondria.

BACKGROUND: Colorectal cancer is the third most-common cancer and the second most-common cause of cancer related death in UK. Although chemotherapy plays significant role in the treatment of colorectal cancer, morbidity and mortality due to drug resistance and cancer metastasis are yet to be eliminated. Recently, doxycycline has been reported to have cytotoxic and anti-proliferating properties in various cancer cells. In this study, whether doxycycline was apoptosis threshold lowering agent in colorectal cancer cells by targeting mitochondria was answered. RESULTS: This study showed dose-dependent cytotoxic effects of cisplatin, oxaliplatin and doxycycline in HT29 colorectal cancer cells. Doxycycline showed inhibition of cytochrome-c-oxidase activity in these cells over a time-period. The pre-treatment of doxycycline reported statistically significant increased cytotoxicity of cisplatin and oxaliplatin compared to cisplatin and oxaliplatin alone. The caspase studies revealed significantly less expression and activity of caspase 3 in HT29 cells pre-treated with doxycycline compared to the cells treated with cisplatin and oxaliplatin alone. CONCLUSIONS: It was concluded that doxycycline lowered the apoptotic threshold in HT 29 cells by targeting mitochondria. This also raised possible caspase-independent mechanisms of apoptosis in HT29 cells when pre-treated with doxycycline however this needs further research work.

Pretreatment with insulin-like growth factor I protects skeletal muscle cells against oxidative damage via PI3K/Akt and ERK1/2 MAPK pathways.

Oxidative stress has an important role in the pathogenesis of many muscle diseases. The major contributors to oxidative stress in muscle tissue are reactive oxygen species such as oxygen ions, free radicals, and peroxides. Insulin-like growth factor I (IGF-I) has been shown to increase muscle mass and promote muscle cell proliferation, differentiation, and survival. We, therefore, hypothesized that IGF-I might also be cytoprotective for muscle cells during oxidative stress. Exogenous hydrogen peroxide (H(2)O(2)) was used to induce oxidative stress/damage in two types of skeletal muscle cells. Apoptotic pathways were assessed after the oxidative damage and the effects of IGF-I on oxidative stress in muscle cells were examined. Different IGF-I sub-pathways were analyzed with measurement of the expression of pro-and anti-apoptotic proteins. It was found that H(2)O(2) diminishes muscle cell viability and induces a caspase-independent apoptotic cell death. Pretreatment with IGF-I protects muscle cells from H(2)O(2)-induced cell death and enhances muscle cells survival. This effect appears to result from the promotion of the anti-apoptotic protein, Bcl2. Further investigation shows that protection is via an IGF-I sub-pathway: PI3K/Akt and ERK1/2 MAPK pathways. Protecting muscle cells from oxidative damage presents a potential application in the treatment of the muscle wasting, which appears in many muscle pathologies including Duchenne muscle dystrophy and sarcopenia.

In vitro small intestinal epithelial cell growth on a nanocomposite polycaprolactone scaffold.

Tissue engineering of the small intestine remains experimental despite worldwide attempts to develop a functional substitute for short bowel syndrome. Most published studies have reported predominant use of PLLA (poly-L-lactide acid)/PGA (polyglycolic acid) copolymer as the scaffold material, and studies have been limited by in vivo experiments. This lack of progress has inspired a fresh perspective and provoked further investigation and development in this field of tissue engineering. In the present paper, we exploit a relatively new nanocomposite of POSS (polyhedral oligomeric silsesquioxane) and PCL [poly(caprolactone-urea)urethane] as a material to develop porous scaffolds using a solvent casting/particulate leaching technique to fabricate porous scaffolds in different pore sizes and porosities. Scaffolds were characterized for pore morphology and porosity using scanning electron microscopy and micro-computed tomography. Rat intestinal epithelial cells were then seeded on to the polymer scaffolds for an in vitro study of cell compatibility and proliferation, which was assessed by Alamar Blue and lactate dehydrogenase assays performed for 21 days post-seeding. The results obtained demonstrate that POSS-PCL nanocomposite was produced as a macroporous scaffold with porosity over the range of 40-80% and pore size over the range of 150-250 microm. This scaffold was shown to support epithelial cell proliferation and growth. In conclusion, as a further step in investigating small intestinal tissue engineering, the nanocomposite employed in this study may prove to be a useful alternative to poly(lactic-co-glycolic acid) in the future.

IGF-I activates caspases 3/7, 8 and 9 but does not induce cell death in colorectal cancer cells.

BACKGROUND: Colorectal cancer is the third most common cancer in the western world. Chemotherapy is often ineffective to treat the advanced colorectal cancers due to the chemo-resistance. A major contributor to chemo-resistance is tumour-derived inhibition or avoidance of apoptosis. Insulin-like growth factor I (IGF-I) has been known to play a prominent role in colorectal cancer development and progression. The role of IGF-I in cancer cell apoptosis is not completely understood. METHODS: Using three colorectal cancer cell lines and one muscle cell line, associations between IGF-I and activities of caspase 3/7, 8 and 9 have been examined; the role of insulin-like growth factor I receptor (IGF-IR) in the caspase activation has been investigated. RESULTS: The results show that exogenous IGF-I significantly increases activity of caspases 3/7, 8 and 9 in all cell lines used; blocking IGF-I receptor reduce IGF-I-induced caspase activation. Further studies demonstrate that IGF-I induced caspase activation does not result in cell death. This is the first report to show that while IGF-I activates caspases 3/7, 8 and 9 it does not cause colorectal cancer cell death. CONCLUSION: The study suggests that caspase activation is not synonymous with apoptosis and that activation of caspases may not necessarily induce cell death.

Apoptosis and colorectal cancer: implications for therapy.

Colorectal cancer (CRC) is characterized by the partial suppression of apoptosis, which in turn gives tumours a selective advantage for survival and can cause current chemotherapy approaches to be ineffective. Recent progress in understanding the mechanisms of apoptosis in colorectal carcinogenesis has provided potential new targets for therapy. Here, we review recent studies of the regulation of apoptosis and its role in CRC initiation and progression, and we discuss the relationship between chemoresistance and the suppression of apoptosis. Recent progress in targeting apoptotic pathways and their regulators provide strategies for the exploration of novel therapies for CRC.

Does doxycycline work in synergy with cisplatin and oxaliplatin in colorectal cancer?

BACKGROUND: In recent years, apart from antibacterial properties, doxycycline is reported to have cytotoxic and anti-proliferative actions in various cancers including colorectal cancer. Colorectal cancer constitutes one of the most common cancers in the western population. Apart from surgery, chemotherapy plays crucial role in the treatment of colorectal cancer. Cisplatin and oxaliplatin are most commonly used platinum compounds for the cancer chemotherapy. This study has looked for any impact of doxycycline on the cytotoxic effects of platinum compounds in colorectal cancer including its mechanisms of actions. METHODS: HT 29 colorectal cancer cells were used for this study. These cells were treated with cisplatin and oxaliplatin with or without doxycycline treatment. The caspase 3 gene expression was quantitated by gel electrophoresis and qualitated by real time polymerase chain reactions. The caspase 3 activity was assessed in HT 29 cells with fluorescence kit. RESULTS: The results revealed increased caspase 3 gene expressions and activities in HT 29 cells treated with cisplatin, oxaliplatin and doxycycline; however the combination of doxycycline with cisplatin and oxaliplatin did not report increased caspase 3 gene expressions and activity compared to cisplatin and oxaliplatin alone. CONCLUSION: We concluded that doxycycline has role in apoptosis induction in the colorectal cancer. However, it did not show any synergy with platinum compounds in the colorectal cancer cells. This study also pointed towards possible caspase-independent actions of doxycycline with cisplatin and oxaliplatin. However, further work is required to underpin the mechanisms of actions of doxycycline.

Degradation studies on biodegradable nanocomposite based on polycaprolactone/polycarbonate (80:20%) polyhedral oligomeric silsesquioxane.

The development of biocompatible polymers has greatly advanced the field of tissue engineering. Some tissues can be propagated on a nondegradable scaffold. Tissue such as cartilage, however, is a complex tissue in which the chondrocytes require their own synthesized extracellular matrix (ECM) to function. Suitable scaffolds for tissue engineering cartilage should provide mechanical strength and degrade at a similar rate to that of cell growth and ECM production. We have developed a biodegradable nanocomposite based on polycaprolactone and polycarbonate polyurethane (PCU) with an incorporated polyhedral oligomeric silsesquioxane (POSS) (POSS modified Poly(caprolactone/carbonate) urethane/urea). Previous work on POSS incorporated into PCU (POSS-PCU) has been shown to possess good mechanical strength, elasticity and resistance to degradation. This series of experiments involved exposing this polymer to a selection of accelerated degradative solutions for up to 8 weeks. The samples were analyzed by infra-red spectroscopy, scanning electron microscopy, X-ray microanalysis, contact angle analysis, and stress-strain mechanical analysis. Degradation of hard and soft segments of the nanocomposite was evident by infra-red spectroscopy in all conditioned samples. POSS nanocage degradation was evident in some oxidative/peroxidative systems accompanied by gross changes in surface topography and significant changes in mechanical properties. The hydrophobic polymer became more hydrophilic in all conditions. This biodegradable nanocomposite demonstrated steady degradation with protection of mechanical properties when exposed to hydrolytic enzymes and plasma protein fractions and exhibited more dramatic degradation by oxidation.This pattern may be potentially employed in tissue engineering scaffolds where controlled degradation and retained structural stability of the scaffold is required.