Biomolecular study and conjugation of two para-aminobenzoic acid derivatives with serum proteins: drug binding efficacy and protein structural analysis.

Two aminobenzoic acid derivatives DAB-0 and DAB-1 showed distinct biological properties on murine bladder cancer (BCa) cell line MB49-I. In contrast to DAB-1, DAB-0 does not possess any anti-inflammatory activity and is less toxic. Furthermore, DAB-0 does not interfere with INFγ-induced STAT1 activation and TNFα-induced IκB phosphorylation, while DAB-1 does. In order to rationalize these results, the binding efficacy of DAB-0 and DAB-1 with serum proteins such a human serum albumin (HSA), bovine serum albumin (BSA) and beta-lactoglobulin (ß-LG) was investigated in aqueous solution at physiological pH. Multiple spectroscopic methods and thermodynamic analysis were used to determine the binding efficacy of DAB-0 and DAB-1 with serum proteins. Drug-protein conjugation was observed via through ionic contacts. DAB-1 forms stronger adducts than DAB-0, while ß-LG shows more affinity with the order of stability ß-LG > BSA > HSA. The stronger complexation of DAB-1 with serum proteins might account for its biological potential and transport in the blood. The binding efficacy ranged from 40 to 60%. Major alterations of protein secondary structures were detected upon drug complexation. Serum proteins are capable of delivering DAB-1 in vitro.Communicated by Ramaswamy H. Sarma.

Binding efficacy of aminobenzoic acid derivatives with DNA duplex: drug binding sites and DNA structure and dynamics.

Communicated by Ramaswamy H. Sarma.

Optimal germination condition impacts on the antioxidant activity and phenolic acids profile in pigmented desi chickpea (Cicer arietinum L.) seeds.

Legume sprouts are considered natural, healthy products that provide a source of bioactive compounds to fight against chronic diseases. This study aims to identify the optimal germination temperature (GT) and germination time (Gt) to maximize total phenolic and flavonoid contents (TPC, FC), and antioxidant activity (AoxA) of desi chickpea. Response surface methodology was used as an optimization tool. An experimental design with two factors (GT and Gt) and five levels was used (13 treatments). The sprouts from each treatment were lyophilized, tempered, and milled to obtain germinated chickpea flours (GCF). To predict the phytochemicals composition and AoxA in GCF, regression models were developed. Maximum TPC, FC, and AoxA were attained during germination 33.7 °C for 171 h. Optimized germinated chickpea flour produced applying the optimal germination conditions resulted in an increase of protein and total dietary fibre content, TPC, FC, phenolic acids profile, and AoxA. Germination at optimal conditions also increased the level of coumaric, ferulic, synapic, ellagic, and syringic acids. This study demonstrated that germination carried out under optimal conditions enhanced the nutraceutical value of desi chickpea seeds.

Chemical composition and physicochemical properties of Phaeodactylum tricornutum microalgal residual biomass.

The production of photosynthetic biofuels using microalgae is a promising strategy to combat the use of non-renewable energy sources. The microalgae residual biomass is a waste by-product of biofuel production; however, it could prove to have utility in the development of sustainable nutraceuticals and functional foods. In this study, a comprehensive characterisation of the under-utilised Phaeodactylum tricornutum microalgae residual biomass is presented. Proximal composition, antioxidant capacity (using three different antioxidant assays; oxygen radical absorbance capacity; radical cation activity, ABTS; and radical scavenging activity, DPPH), and total phenolic content of free and bound polyphenols were determined. Additionally, the physicochemical properties of water activity, pH, water absorption index, water solubility index, and dispersibility were evaluated. Results revealed that P. tricornutum microalgae residual biomass exhibits a relatively high protein and carbohydrate content, with values of 36.67% and 46.78%, respectively; and most carbohydrates were found as total dietary fibre (45.57%), of which insoluble dietary fibre was the most predominant (43.54%). Antioxidant capacity values for total phytochemicals of 106.22, 67.93, 9.54 µM TE g-1 dw were determined by oxygen radical absorbance capacity, ABTS, and DPPH assays, respectively. Total phenolic content was found to be 2.90 mg GAE g-1 dw. Interestingly, antioxidant capacity and total phenolic content were higher in bound than in free phytochemical extracts. The physicochemical analysis showed P. tricornutum microalgae residual biomass to have suitable properties for the generation of a beverage with Aw, pH, water absorption index, water solubility index, and dispersibility values of 0.45, 7.12, 3.40 g gel g-1 dw, 2.5 g solids 100 g-1 dw, and 90%, respectively. Hence, P. tricornutum microalgae residual biomass could be considered a potential source of bioactive compounds suitable for the production of functional food exhibiting antioxidant capacity and high dietary fibre content.

Transmission efficiency of tomato apex necrosis virus by Bemisia tabaci (Hemiptera: Aleyrodidae) biotype B in tomato.

Tomato apex necrosis virus (ToANV) is a new virus that causes important damage in tomato crops from the Culiacan Valley, Sinaloa, Mexico. To understand the relationship between ToANV and its vector Bermisia tabaci (Hemiptera: Aleyrodidae) (Gennadius) biotype B, laboratory and greenhouse trials were completed to: 1) determine the acquisition and inoculation access periods of ToANV by B. tabaci from tomato to tomato, 2) understand the transmission efficiency at different B. tabaci population densities, 3) estimate the time from inoculation of the virus at different B. tabaci densities to manifestation of symptoms in the plants, and 4) determine the retention time of the virus by the insect vector. The presence of the virus in plants was determined by reverse transcription-polymerase chain reaction amplification ofa 795-bp fragment (GenBank JN704068), which is phylogenetically related to ToANV (GenBank EF063242). The results showed that B. tabaci is an effective vector for ToANV with relatively long acquisition (12 h) and inoculation (9 h) access periods; a single adult is capable of transmitting and retaining the virus for up to 7d, suggesting a persistent mode of transmission. These results will help in the development of management strategies for controlling the vector and the disease.

A possible mechanism of metabolic regulation in Gibberella fujikuroi using a mixed carbon source of glucose and corn oil inferred from analysis of the kinetics data obtained in a stirrer tank bioreactor.

A nonstructured model was used to study the dynamics of gibberellic acid production in a stirred tank bioreactor. Experimental data were obtained from submerged batch cultures of Gibberella fujikuroi (CDBB H-984) grown in varying ratios of glucose-corn oil as the carbon source. The nitrogen depletion effect was included in mathematical model by considering the specific kinetic constants as a linear function of the normalized nitrogen consumption rate. The kinetics of biomass growth and consumption of phosphate and nitrogen were based on the logistic model. The traditional first-order kinetic model was used to describe the specific consumption of glucose and corn oil. The nitrogen effect was solely included in the phosphate and corn oil consumption and biomass growth. The model fit was satisfactory, revealing the dependence of the kinetics with respect to the nitrogen assimilation rate. Through simulations, it was possible to make diagrams of specific growth rate and specific rate of substrate consumptions, which was a powerful tool for understanding the metabolic interactions that occurred during the various stages of fermentation process. This kinetic analysis provided the proposal of a possible mechanism of regulation on growth, substrate consumptions, and production of gibberellic acid (GA3 ) in G. fujikuroi.

Physicochemical, nutritional and antioxidant properties of tempeh flour from common bean ( Phaseolus vulgaris L.).

The effects of solid state fermentation (SSF) on physicochemical, nutritional and antioxidant properties of common bean flour were studied. SSF increased protein content (21.7%) and decreased lipids (-38.4%), carbohydrates (-3.5%) and phytic acid (-58.3%). Fermented (tempeh) flour showed higher dispersability, lower water solubility index and pH than unfermented flour. Fermentation also increased an average of 0.21 g/100 g protein, six of the essential amino acids (EAAs), including total sulfur (Met + Cys), the limiting EAAs in unfermented flour (score = 0.91); Lys and Trp decreased 0.21 and 0.09 g/100 g protein, respectively. SSF improved the in vitro protein digestibility and the calculated protein efficiency ratio. Tempeh flour had 2.2-fold more phenolics than the bean flour and exhibited antiradical activity (43%) and antioxidant activity (38%) correlated with total phenolics content. Common bean tempeh flour may be considered for the fortification of widely consumed legume-based food products and also for the prevention of pathologies associated with oxidative stress.

Nutritional properties of quality protein maize and chickpea extruded based weaning food.

Malnutrition is one of the major causes of morbidity and mortality among young children in most of the developing countries. To minimize the adversities of malnutrition, low-cost infant supplementary foods have been developed and are being supplied to the needy through state-sponsored nutrition intervention programmers. The present study had two objectives: to determine the best combination of nixtamalized extruded quality protein maize (NEMF) and extruded chickpea (ECF) flours for producing a weaning food, and to evaluate the nutritional properties of the optimized NEMF/ECF mixture and the weaning food. The NEMF and ECF were produced applying combinations of extrusion temperature/screw speed of 79.4 degrees C/73.5 rpm, and 150.5 degrees C/190.5 rpm, respectively. Response surface methodology was applied to determine the optimum combination NEMF/ECF; the experimental design generated 11 assays. Mixtures from each assay were evaluated for true protein (TP) and available lysine (AL). Each one of 11 mixtures were used for preparing 11 weaning foods which were sensory evaluated for acceptability (A). The best combination of NEMF/ECF for producing a weaning food was NEMF = 21.2%/ ECF = 78.8 %. This mixture had a global desirability (D) of 0.93; it contained 20.07% proteins (DM), 5.70% lipids (DM), and 71.14% carbohydrates (DM); its essential amino acids (EAA) profile satisfactorily covered the EAA requirements for children 2-5 years old, except for Trp. The weaning food prepared with the optimized mixture had high protein quality and digestibility and could be used to support the growth of infants.

Infant food from quality protein maize and chickpea: optimization for preparing and nutritional properties.

The present study had two objectives: to determine the best combination of nixtamalized maize flour (NMF) from quality protein maize and extruded chickpea flour (ECF) for producing an infant food, and to evaluate the nutritional properties of the optimized NMF/ECF mixture and the infant food. Response surface methodology (RSM) was applied to determine the best combination of NMF/ECF; the experimental design (Lattice simplex) generated 11 assays. Mixtures from each assay were evaluated for true protein and available lysine. Each one of 11 mixtures was used for preparing 11 infant foods that were sensory evaluated for acceptability. A common optimum value for the three response variables was obtained utilizing the desirability method. The best combination of NMF/ECF for producing an infant food was NMF = 26.7%/ECF = 73.3%; this optimized mixture had a global desirability of 0.87; it contained 19.72% dry matter (DM) proteins, 6.10% (DM) lipids, 71.45% (DM) carbohydrates, and 2.83% (DM) minerals; its essential amino acids profile covered the amino acids requirements for children 10-12 years old. The infant food prepared from optimized mixture had an in vitro protein digestibility of 87.9%, and a calculated protein efficiency ratio of 1.86. Infant food could be used to support the growth of infants in developing countries.

Localization of the chaperone proteins GRP78 and HSP60 on the luminal surface of bovine oviduct epithelial cells and their association with spermatozoa.

Upon their transit through the female genital tract, bovine spermatozoa bind to oviduct epithelial cells, where they are maintained alive for long periods of time until fertilization. Although carbohydrate components of the oviduct epithelial cell membrane are involved in these sperm/oviduct interactions, no protein candidate has been identified to play this role. To identify the oviduct factors involved in their survival, sperm cells were preincubated for 30 min with apical membranes isolated from oviduct epithelial cells, washed extensively, and further incubated for up to 12 h in the absence of apical membranes. During this incubation, sperm viability, motility, and acrosomal integrity were improved compared with cells preincubated in the absence of apical membranes. This suggests that, during the 30-min preincubation with apical membrane extracts, either an oviductal factor triggered intracellular events resulting in positive effects on spermatozoa or that such a factor strongly attached to sperm cells to promote a positive action. Similarly, spermatozoa were incubated with apical membranes isolated from oviduct epithelial cells labeled with [35S]-methionine and, upon extensive washes, proteins were separated by two-dimensional (2-D) gel electrophoresis to identify the factors suspected to have beneficial effects on spermatozoa. The six major proteins, according to their signal intensity on the autoradiographic film, were extracted from a 2-D gel of oviduct epithelial cell proteins run in parallel and processed for N-terminal sequencing of the first 15 amino acids. Of these, one was identical to heat shock protein 60 (HSP60) and one to the glucose-regulated protein 78 (GRP78). Their identities and association with spermatozoa were confirmed using an antibody directed against these proteins. This paper reports the localization of both GRP78 and HSP60 on the luminal/apical surface of oviduct epithelial cells, their binding to spermatozoa, and the presence of endogenous HSP60 in the sperm midpiece.

Nixtamalized flour from quality protein maize (Zea mays L). optimization of alkaline processing.

Quality of maize proteins is poor, they are deficient in the essential amino acids lysine and tryptophan. Recently, in Mexico were successfully developed nutritionally improved 26 new hybrids and cultivars called quality protein maize (QPM) which contain greater amounts of lysine and tryptophan. Alkaline cooking of maize with lime (nixtamalization) is the first step for producing several maize products (masa, tortillas, flours, snacks). Processors adjust nixtamalization variables based on experience. The objective of this work was to determine the best combination of nixtamalization process variables for producing nixtamalized maize flour (NMF) from QPM V-537 variety. Nixtamalization conditions were selected from factorial combinations of process variables: nixtamalization time (NT, 20-85 min), lime concentration (LC, 3.3-6.7 g Ca(OH)2/l, in distilled water), and steep time (ST, 8-16 hours). Nixtamalization temperature and ratio of grain to cooking medium were 85 degrees C and 1:3 (w/v), respectively. At the end of each cooking treatment the steeping started for the required time. Steeping was finished by draining the cooking liquor (nejayote). Nixtamal (alkaline-cooked maize kernels) was washed with running tap water. Wet nixtamal was dried (24 hours, 55 degrees C) and milled to pass through 80-US mesh screen to obtain NMF. Response surface methodology (RSM) was applied as optimization technique, over four response variables: In vitro protein digestibility (PD), total color difference (deltaE), water absorption index (WAI), and pH. Predictive models for response variables were developed as a function of process variables. Conventional graphical method was applied to obtain maximum PD, WAI and minimum deltaE, pH. Contour plots of each of the response variables were utilized applying superposition surface methodology, to obtain three contour plots for observation and selection of best combination of NT (31 min), LC (5.4 g Ca(OH)2/l), and ST (8.1 hours) for producing optimized NMF from QPM.

Addition of specific metabolites to bovine epididymal cell culture medium enhances survival and motility of cryopreserved sperm.

We have developed a cell culture system of bovine epididymal epithelium in which cryopreserved bovine sperm motility was efficiently maintained for many hours. The culture conditions to maintain viable epididymal cells are quite different from conditions normally used to incubate sperm cells. Thus, we have modified a previously described principal cell medium (PCM; Moore et al, 1992) using HEPES as a buffer and supplemented media with myo-inositol, pyruvate, lactate, glycerol, and carnitine to mimic epididymal intraluminal conditions. In the first experiments the effects of PCM and our epididymal cell medium (ECM) on sperm motility were compared in the absence of cells and evaluated by microscopic analysis under a phase contrast microscope or using the Hamilton Thorn Image Analyzer System. Our results showed that motility of cauda epididymal sperm was significantly higher in ECM than in PCM during a 48-hour incubation period when both media were supplemented with 10% fetal bovine serum (FBS). We then replaced FBS with bovine serum albumin (BSA) or no proteins at all to verify if ECM was able to enhance sperm survival. To test this aspect we used frozen-thawed sperm, which survived up to 48 hours when sperm cells were coincubated with epididymal cell monolayers. Hence, PCM, ECM, and different media containing each metabolite of ECM were supplemented with 0.5% BSA to assess motility of thawed sperm after an incubation period of 6 hours. A positive effect on sperm motility was observed in all fresh and unconditioned media containing 1 mM pyruvate. Motion parameters were more efficiently maintained in all conditioned media than in unconditioned media. Our results showed, however, that pyruvate was almost completely oxidized or consumed by epididymal cells during preincubation of culture media. We conclude that motility of frozen-thawed bovine spermatozoa can be improved using a culture medium or a medium conditioned by epididymal cell cultures without carnitine but containing mainly pyruvate, inositol, glycerol, and lactate.

Optimization of the solid state fermentation process to obtain tempeh from hardened chickpeas (Cicer arietinum L.).

Solid state fermentation (SSF) represents a technological alternative for a great variety of cereals and legumes, or combination of them, to improve their nutritional quality and to obtain edible products with palatable sensorial characteristics. The objective of this work was to find the best conditions of fermentation temperature and time to obtain tempeh from hardened chickpeas (Cicer arietinum L.) applying SSF. Response surface methodology (RSM) was applied over three response variables (phytic acid, in vitro protein digestibility and available lysine) to find best conditions of fermentation to carry out the process. A central composite experimental design with two factors [X1 = temperature (31-36 degrees C) and X2 = time (48-72 h)] in five levels (2 factorials, 2 axial, I central) was used. Spores from Rhizopus stolonifer were suspended in distilled water (1 x 10(6) spores/mL) and used as starter. According to regression models, minimum and maximum levels of the response variables were 1.24-2.66 mg phytic acid/g of sample DM, 77.6-83.5% in vitro protein digestibility and 2.18-4.63 g available lysine/16 g N. The superposition of contour plots of each one of the response variables allowed researchers to find, graphically, the best conditions for the SSF process: 35.8 degrees C for 42.7 h.

Hard-to-cook phenomenon in chickpeas (Cicer arietinum L): effect of accelerated storage on quality.

Storage, at high temperature (> or = 25 degrees C) and high relative humidity (> or = 65%), causes development of hard to cook (HTC) phenomenon in grain legumes. The objective of this work was to study the effect of storage simulating tropical conditions on chickpeas quality. The hardening of the Surutato 77, Mocorito 88, and Blanco Sinaloa 92 chickpea varieties was produced using adverse storage (32 +/- 1 degrees C, RH = 75%, 160 days) conditions. For all samples, the Hunter 'L' values decreased and deltaE values increased during storage, meaning a loss of color lightness and development of darkening. Accelerated storage caused a significant decrease in the water absorption capacities and cooking times of whole seeds, cotyledons and seed coats of all samples, being more pronounced in The Blanco Sinaloa 92 variety. Furthermore, storage produced significant decreases in the seed coat tannin content of the three materials; this parameter increased significantly in the cotyledon. In all samples, the levels of phytic acid decreased significantly with the seed hardness. Hardening of chickpea grains caused a decrease in the in vitro protein digestibilities of all varieties. These results suggest that both the cotyledon and seed coat play a significant role in the process of chickpea hardening. Blanco Sinaloa 92 and Mocorito 88 might be classified as varieties with high and low proneness, respectively, to the development of the HTC condition.

Chemotherapy cytotoxicity of human MCF-7 and MDA-MB 231 breast cancer cells is altered by osteoblast-derived growth factors.

One-third of women with breast cancer will develop bone metastases and eventually die from disease progression at these sites. Therefore, we analyzed the ability of human MG-63 osteoblast-like cells (MG-63 cells), MG-63 conditioned media (MG-63 CM), insulin-like growth factor I (IGF-I), and transforming growth factor beta 1 (TGF-beta1) to alter the effects of adriamycin on cell cycle and apoptosis of estrogen receptor negative (ER-) MDA-MB-231 and positive (ER+) MCF-7 breast cancer cells, using cell count, trypan blue exclusion, flow cytometry, detection of DNA fragmentation by simple agarose gel, and the terminal deoxynucleotidyl transferase (TdT)-mediated nick end-labeling method for apoptosis (TUNEL assay). Adriamycin arrested MCF-7 and MDA-MB-231 cells at G2/M phase in the cell cycle and inhibited cell growth. In addition, adriamycin arrested the MCF-7 cells at G1/G0 phase and induced apoptosis of MDA-MB-231 cells. Exogenous IGF-I partially neutralized the adriamycin cytotoxicity/cytostasis of cancer cells. MG-63 CM and TGF-beta1 partially neutralized the adriamycin cytotoxicity of MDA-MB-231 cells but enhanced adriamycin blockade of MCF-7 cells at G1/G0 phase. MG-63 osteoblast-like cells inhibited growth of MCF-7 cells while promoting growth and rescued MDA-MB-231 cells from adriamycin apoptosis in a collagen co-culture system. These data suggest that osteoblast-derived growth factors can alter the chemotherapy response of breast cancer cells. Conceivably, host tissue (bone)-tumor cell interactions can modify the clinical response to chemotherapy in patients with advanced breast cancer.

Response of PC-3 prostate cancer cells to combination therapy using irradiation with glucocorticoids or doxorubicin.

OBJECTIVES: We evaluated the effects of irradiation, doxorubicin and dexamethasone on human PC-3 prostate cancer cells, investigating whether dexamethasone and doxorubicin can alter the irradiation cytotoxicity of PC-3 cells. METHODS: We used the human PC-3 prostate cancer cells, analyzing cell growth with trypan blue exclusion, indices of the cell cycle with flow cytometry and apoptosis with flow cytometry and analysis of DNA fragmentation on simple agarose gel. RESULTS: Doxorubicin (100 nM) arrested cell cycle at the G2/M phase, decreased cell growth and produced apoptosis of PC-3 cells in a time-dependent manner. Dexamethasone (100 nM) increased the distribution of PC-3 cells at G0/G1 phase in the cell cycle, exerting an inhibitory effect on the proliferation of PC-3 cells after 48 and 72 hr, but it did not produce apoptosis. Irradiation (4 Gy) initially arrested cells at the G2/M phase in the cell cycle (24 hr) which was gradually overcome and the PC-3 cells were shifted into G0/G1 phase or apoptosis after 48 and 72 hr. Irradiation decreased the PC-3 cell growth by 40-50% after 48 and 72 hr, respectively. Treatment with doxorubicin (100 nM) for 24, 48, and 72 hr after irradiation potentiated irradiation cytotoxicity of PC-3 cells. Dexamethasone treatment 24 hr before and 24, 48 and 72 hr after irradiation increased the number of surviving PC-3 cells and partially neutralized the irradiation effects on cell cycle. CONCLUSION: Doxorubicin potentiated while dexamethasone partially reversed the irradiation cytotoxicity of PC-3 cells. These data may be of clinical importance for the treatment of hormone refractory prostate cancer.

Osteoblast-derived survival factors protect PC-3 human prostate cancer cells from adriamycin apoptosis.

OBJECTIVES: Hormone-independent and cytotoxic drug-resistant tumor growth in osteoblastic metastases defines poor survival in patients with advanced prostate cancer. Therefore, we analyzed the ability of human osteoblast-like cells (MG-63 cells) and MG-63 conditioned media (MG-63 CM) to protect PC-3 human prostate cancer cells from adriamycin cytotoxicity in vitro. METHODS: Adriamycin cytotoxicity was assessed in MG-63 osteoblast-like and PC-3 prostate cancer monolayer and three-dimensional collagen coculture systems using the DNA content and trypan blue exclusion assays, analysis of indexes of cell cycle by flow cytometry, determination of DNA fragmentation on simple agarose gel and terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) assay, and immunocytochemistry. RESULTS: Adriamycin (100 nM) arrested both the PC-3 and MG-63 cells at the G2/M phase in the cell cycle but induced apoptosis only in PC-3 cells, as assessed by flow cytometry, trypan blue exclusion, and agarose gel. Optimal doses of MG-63 CM (50 microg/mL), insulin-like growth factor I (50 ng/mL), and transforming growth factor-beta-1 (25 ng/mL), as determined by DNA content assay, partially neutralized the adriamycin cytotoxicity of PC-3 cells detected by flow cytometry and trypan blue exclusion. In addition, MG-63 cells rescued PC-3 cells from adriamycin apoptosis in the three-dimensional type I collagen gel coculture system, as analyzed by TUNEL assay. CONCLUSIONS: These data suggest that osteoblast-like cells and osteoblast-derived growth factors can optimize survival of metastatic prostate cancer cells, thereby helping to develop cytotoxic drug-resistant growth in vitro.

Osteoblast-derived growth factors enhance adriamycin-cytostasis of MCF-7 human breast cancer cells.

Bone only metastasis in patients with estrogen receptor (ER) positive breast cancer reported to have favorable response to chemotherapy, favorable prognosis, and an "indolent" course. Therefore, we assessed the ability of MG-63 osteoblast-like human osteosarcoma cells (MG-63 cells) and MG-63 conditioned media (CM) to influence adriamycin-cytotoxicity of ER-positive MCF-7 human breast cancer cells. Estradiol (E2; 100 nM) increased the distribution at S and G2/M phases in the cell cycle and stimulated the growth of MCF-7 cells. Adriamycin (100 nM) inhibited the growth and arrested the MCF-7 cells supplemented with or without 100 nM of estradiol [(-E2) and (+E2) MCF-7 cultures] at G2/M phase in the cell cycle. In addition, adriamycin (100 nM) increased the distribution at G1/G0 phase in the cell cycle of (+E2) MCF-7 cultures. Adriamycin (100 nM and 10 microM) did not induce apoptosis of MCF-7 cells as assessed by flow cytometry and analysis of DNA fragmentation on simple agarose gel. Exogenous insulin-like growth factor I (IGF I) stimulated while transforming growth factor beta 1 (TGF beta 1) and MG-63 CM inhibited the growth of MCF-7 cells. Furthermore, MG-63 CM and TGF beta 1 enhanced while exogenous IGF I reversed adriamycin (100 nM)-cytostasis of MCF-7 cells. These data suggested that osteoblastic CM contained growth factors, such as TGF beta 1 capable of enhancing adriamycin-cytostasis, in vitro. Conceivably, these osteoblast-derived "enhancers" of chemotherapy-cytostasis can explain the favorable prognosis and "indolent" course of ER-positive breast cancer patients with bone only metastasis.

Growth factors mediate glucocorticoid receptor function and dexamethasone-induced regression of osteoblastic lesions in hormone refractory prostate cancer.

We investigated the ability of important regulators of osteoblast function, such as insulin-like growth factor I (IGF-I), transforming growth factor beta 1 (TGF beta 1), and urokinase-type plasminogen activator (uPA) to act as mediators in cell-cell interactions between osteoblast-like cells and metastatic prostate cancer cells, in vitro. In addition, we assessed whether these growth substances can (a) mediate glucocorticoid receptor (GR) function and (b) be implicated in dexamethasone-induced regression of osteoblastic tumors. Exogenous IGF-I, rat/human uPA, and PA-III (rat)/PC-3 (human) prostate cancer cells conditioned media (CM) stimulated the proliferation of rat (UMR 106 cells) and human (MG-63 cells) osteosarcoma cells. This mitogenic activity was completely neutralized by anti-IGF-I specific antibody. In addition, dexamethasone decreased cell growth, up regulated TGF beta 1 mRNA, and down regulated uPA mRNA expression in prostate cancer cells. Furthermore, it inhibited cell growth by activating latent-TGF beta 1 in osteoblast-like cells. In addition, dexamethasone down regulated the expression of IGF-I mRNA in osteoblast-like cells. Therefore, it is conceivable that uPA, TGF beta 1 and IGF-I mediate at least in part cell-cell interactions and GR function in osteoblastic metastases. Conceivably, regression of the osteoblastic tumors produced by high-dose dexamethasone treatment in hormone-refractory prostate cancer patients is been mediated by differential regulation of growth factors, locally.

Glucocorticoid receptor function possibly modulates cell-cell interactions in osteoblastic metastases on rat skeleton.

We analysed the glucocorticoid receptor (GR) function and its ability to modulate cell-cell interactions between the PA-III rat prostate cancer and UMR 106 osteoblast-like rat osteosarcoma cells as an in vitro model for studying GR function in PA-III cell-induced tumor and blastic reaction in rat bone. Intact GR was detected by ligand binding assays, DNA band-shift, and GR trans-activation analysis of PA-III and UMR 106 cells transiently transfected with the mouse mammary tumor virus thymidine kinase-chloramphenicol acetyltransferase reporter gene. Dexamethasone and transforming growth factor beta 1 (TGFbeta1) inhibited the growth of PA-III and UMR 106 cells. Dexamethasone's inhibition of PA-III and UMR 106 cells was reversed by anti-TGFbeta1 antibody and exogenous insulin-like growth factor I (IGF-I). Exogenous IGF-I, urokinase-type plasminogen activator (uPA), UMR 106 conditioned media (CM) and PA-III CM stimulated the proliferation of PA-III and UMR 106 cells. The mitogenic activity exerted by uPA and PA-III CM in UMR 106 cells was completely neutralized by anti-IGF-I specific antibody. In addition, dexamethasone up-regulated TGFbeta1 mRNA and down-regulated uPA mRNA expression in PA-III cells without affecting TGFbeta1 and uPA mRNA expression in UMR 106 cells. These data suggested that TGFbeta1, uPA, and IGF-I mediate at least in part cell-cell interactions and GR function in PA-III prostate cancer and UMR 106 osteosarcoma cells.