Use of coumarins as complementary medicine with an integrative approach against cervical cancer: background and mechanisms of action.

Cervical cancer is characterized by the cellular transformation caused by Human Papillomavirus (HPV), favoring cell proliferation, migration, invasion, and metastasis. Cervical cancer is conventionally treated with radiation therapy, and chemotherapy focused on the destruction of tumor cells. However, chemoresistance and low selectivity between tumor and non-tumor cells have been reported, causing side effects in patients. Metabolites of natural origin have shown selectivity against tumor cells, suggesting their use for reducing the side effects caused by drugs used in conventional therapy. Among these compounds, several natural coumarins stand out, such as auraptene, scopoletin, osthole, and praeruptorin, of which antiproliferative, anti-migratory, and anti-invasive activity have been reported. Auraptene, scopoletin, osthole, and praeruptorin show a cytotoxic or antiproliferative effect on cervical tumor cells, arresting the cell cycle by inducing the overexpression of negative regulators of the cell cycle, or inducing cell death by increasing the expression of pro-apoptotic proteins and decreasing that of anti-apoptotic proteins. On the other hand, auraptene, scopoletin, and praeruptorin inhibit the capacity for migration, invasion, and metastasis of cervical tumor cells, mainly by inhibiting the expression and activity of matrix metalloproteinase-2 and -9. The PI3K/Akt signal pathway appears to be central to the anti-tumor activity of the coumarins analyzed in this review. In addition, auraptene, osthole, and praeruptorin are useful in sensitizing tumor cells to radiotherapy or chemotherapeutic molecules, such as FOLFOX, cisplatin, or DOX. Coumarins offer an excellent possibility for developing new drugs as complementary medicine with an integrative approach against cervical cancer.

Possible role of available phosphorus in potentiating the use of low-protein diets for broiler chicken production.

A total of 945 male Ross 308 broiler chicks were used in a growth study to explore the interaction between dietary crude protein concentration and available phosphorus. Nine experimental treatments were constructed factorially by offering low, medium, or standard protein concentrations without or with low, standard, or high available phosphorus. Diets were based on corn, wheat, and soybean meal and all nutrients other than protein/amino acids and available phosphorus were maintained at or above breeder guidelines. Additional synthetic amino acids were used in the diets with low protein concentration in attempt to maintain digestible amino acid supply. Diets were offered to 7 replicate pens of 15 chicks per pen from day 8 to 35. Growth performance was measured during the grower (day 8-24) and finisher (day 25-35) periods. On day 35 carcass composition was determined, blood was drawn for various biochemical measurements and the tibia was excised for mechanical and compositional analyses. Birds that received the low-protein diet had lower terminal body weight and higher feed conversion ratio compared with those that received diets with adequate crude protein content. However, addition of available phosphorus to the low-protein diet resulted in significant reductions in weight-corrected feed conversion that were not evident in the diet with adequate protein content. Bone architecture was only moderately influenced by dietary treatment but birds that ingested the diets containing low and medium protein concentrations had relatively heavier abdominal fat pad weight. Blood biochemistry, especially ammonia, uric acid, and phosphorus, was influenced by both dietary protein and available phosphorus and trends suggested that both axes are involved in protein accretion and catabolism. It can be concluded that performance losses associated with feeding low protein diets to broiler chickens may be partially restored by additional available phosphorus. The implications for use of exogenous enzymes such as protease and phytase and protein nutrition per se warrants further examination.

Supplementation of broiler diets with high levels of microbial protease and phytase enables partial replacement of commercial soybean meal with raw, full-fat soybean.

A 3 × 3 + 1 factorial, involving three levels of protease (0, 15,000 or 30,000 PROT/kg) and three levels of phytase (1,000, 2,000 or 3,000 FYT/kg), was used to evaluate the effect of replacing commercial soybean meal (SBM) with raw, full-fat soybean (RFSB) at 75 g/kg of diet for broilers. A control diet was used for comparison. Each treatment was replicated six times, with nine birds per replicate. The concentration of trypsin inhibitors (TIs) in the test diets was approximately 10,193.4 TIU/kg. Regardless of enzyme supplementation, feed intake (FI) and body weight gain (BWG) of birds in the control group were superior to those on the test diets. Birds that received the protease-free test diets had reduced FI and BWG, but when supplemented with protease, were similar to the control diet in BWG, FI (except 0-35 days) and feed conversion ratio (FCR). When the test diet was supplemented with elevated levels (extradose) of protease and phytase, the BWG was improved during 0-10 days (p = .05) and 0-24 days (p < .01). Regardless of protease supplementation, the weight of thighs was lower for birds fed the test diets. Birds that received the control diet had smaller weight of pancreas. Increasing the level of phytase supplementation reduced (p < .05) the weight of the pancreas. The apparent ileal digestibility (AID) of CP and AA was higher in birds on the control diets, but this was also improved in test diets by protease supplementation. The activities of trypsin (7%), general proteolytic (11%) and lipase (12%) were slightly increased because of protease supplementation. Mucosal depth and apparent villus surface areas were increased by about 2.9% and 20%, respectively, due to supplementation of elevated level of phytase. It can be concluded that RFSB could partially replace SBM in broiler diets, provided the diets are supplemented with elevated levels of protease and phytase.

Physiological and health-related response of broiler chickens fed diets containing raw, full-fat soya bean meal supplemented with microbial protease.

A 2 × 3 factorial study (protease: 0 or 1,5000 PROT/kg and raw full-fat soya bean meal [RSBM] replacing the commercial SBM at 0, 45 and 75 g/kg of diet) was conducted to examine the performance of broilers. Phytase (2000 FYT/kg) was uniformly added to each diet, each also replicated six times, with eight birds per replicate. Birds were raised in climate-controlled rooms using sawdust as the bedding material and offered starter, grower and finisher diets. Feed intake (FI) and body weight gain (BWG) were reduced (p < .05) due to increasing levels of RSBM, but feed conversion ratio (FCR; 0-35 days) was unaffected. Over the first 24 days, neither RSBM nor protease supplementation affected (p > .05) mortality, footpad dermatitis or intestinal lesions in birds. At day 24, the weight, length, width and strength of tibia bone were reduced in chickens that received an elevated level of RSBM (75 g/kg of diet), but this was not significant at day 35. At day 24 (p < .05) and 35 (p < .01), Ca concentration in the litter was reduced when the RSBM level was increased in the diet, but P content was not affected. On days 24 (p < .05) and 35 (p < .01), the N content in litter was also increased with increase in dietary RSBM. Protease supplementation increased (p < .05) the uric acid concentration in the litter (at day 35), but the reverse was the case for ammonia concentration. Overall, the results of this study indicate that there are no major health-related risks, associated with the replacement of commercial SBM with RSBM (≤25%) in broiler diets.

Pasture, multi-enzymes, benzoic acid and essential oils positively influence performance, intestinal organ weight and egg quality in free-range laying hens.

1. The objective of this study was to investigate the effect of range type, multi-enzyme applications, and a combination of benzoic acid (BA) and essential oils (EO) on the productive performance, organ weight and egg quality of free-range laying hens. 2. Three hundred laying hens were evaluated for the short-term (6 weeks) and long-term (12 weeks) effects of range type (G = no pasture, P = pasture) and feed additives (T1 = control; T2 = betaglucanase/pectinase/protease; T3 = BA/EO). Body weight, feed intake (FI), feed conversion ratio (FCR), egg production (EP), digestive organ weight, and egg quality (EQ) were evaluated. Data were analysed using SPSS 2.2 in a 2×2×3 factorial arrangement. 3. Hens that ranged on pasture were significantly heavier (2043 g vs. 1996 g; p < 0.001), laid heavier eggs (61.9 g vs. 60.3 g; p < 0.001) and produced darker yolk colour (4.3 vs. 7.0; p < 0.001) compared to hens ranged on gravel. Hens fed T2 were significantly heavier (2050 g) compared to hens fed T1 (2005 g) or T3 (2008 g). Organ weights (gizzard, liver and pancreas) were significantly heavier in hens ranged on pasture (16.8 g/kg BW, 22.3 g/kg BW and 1.89 g/kg BW, respectively) compared to hens ranged on gravel (14.2 g/kg BW, 21.7 g/kg BW and 1.83 g/kg BW, respectively). Over time, body weight (1970-2070 g; p < 0.001) and egg weight (59.5-62.8 g; p < 0.001) increased, FI (123-120 g; p = 0.024) was reduced and FCR (2.36-2.10; p = 0.002) improved 4. In conclusion, hens housed on pasture and fed multi-enzyme supplemented diets had significantly heavier body weight and produced heavier eggs with darker yolk colour. Pasture intake and enzyme supplementation increased digestive organ weight significantly.

Performance, nutrient utilization, and energy partitioning in broiler chickens offered high canola meal diets supplemented with multicomponent carbohydrase and mono-component protease.

Two broiler chicken experiments were conducted to investigate the effects of canola meal (CM) replacing soybean meal (SBM) in diets supplemented with carbohydrase and protease on performance and partitioning of energy. First, a 2 × 2 × 2 factorial arrangement of treatments was employed to evaluate: protein meals (CM vs. SBM), carbohydrase (none or 300 mg/kg), protease (none or 200 mg/kg), and their interactions. Each treatment was fed to 6 replicated pens of 16 male broilers (Ross 308) from d 10 to 35. In the second experiment, 32 broiler chicks were used in a 2 × 2 factorial arrangement to investigate CM and carbohydrase effects on energy partitioning. Birds were transferred into 16 closed-circuit calorimeter chambers (4 chambers/diet; 2 birds/chamber) to measure heat production (HP), metabolizable and net energy (NE) by gaseous exchange, and total excreta collection from d 25 to 28. There were no 3-way interactions among experimental factors for any of the performance parameters measured. Birds given CM diets consumed less feed, had lower BW, and exhibited higher FCR compared to the control birds (P < 0.01). Both enzymes, alone or in combination, improved final BW and FCR (P < 0.05). There was an interaction between carbohydrase and protease for FCR over the grower period (P < 0.01), in which the combination of the enzymes resulted in further improvement of FCR. Energy, DM, and crude protein digestibility values were higher in control birds (P < 0.05). There was an interaction of protein meal and carbohydrase for HP, respiratory quotient (P < 0.05), and NE:ME ratio of the diets (P = 0.06). Inclusion of CM without carbohydrase increased HP and decreased NE and NE:ME ratio of the diets (P < 0.05). Carbohydrase decreased HP and increased retained energy (P = 0.06) and NE and NE:ME ratio (P < 0.05). In conclusion, high CM in the diet negatively affects growth performance through reduction in feed consumption, nutrient digestibility, and NE of the diet, which could partly be restored by enzyme supplementation.

Central administration of aminoprocalcitonin inhibits food intake and stimulates the hypothalamic-pituitary-adrenal axis in rats via the corticotrophin-releasing factor system.

Aminoprocalcitonin (N-PCT), a neuroendocrine peptide derived from procalcitonin, reduces food intake and body weight when administered centrally in rats. We have recently shown that N-PCT is expressed in brain areas known to be involved in energy homeostasis, including the paraventricular nucleus (PVN) of the hypothalamus, which contains a prominent population of corticotrophin-releasing factor (CRF)-synthesising neurones. CRF plays a pivotal role in the regulation of the hypothalamic-pituitary adrenal (HPA) axis and food intake. However, little is known about functional interactions of N-PCT and CRF. In the present study, we found endogenous N-PCT protein in the rat PVN. We also showed N-PCT immunoreactivity in PVN co-localised with NeuN, a neuronal marker, or glial fibrillary acidic protein, an astrocyte marker. Double staining immunohistochemistry revealed that N-PCT co-localised with CRF in parvocellular neurones of the PVN. Intracerebroventricular N-PCT administration increased CRF mRNA and content in the hypothalamus, suggesting that N-PCT stimulates the HPA axis and suppresses food intake and body weight via CRF-dependent pathways. In keeping with this, i.c.v. co-injection of D-Phe-CRF(12-41), a CRF receptor antagonist, significantly attenuated N-PCT-induced reduction in food intake and body weight in a dose-dependent manner. Furthermore, i.c.v. administration of N-PCT increased plasma adrenocorticotrophic hormone and corticosterone concentrations and induced the expression of Fos protein, a marker of neuronal activity, in parvocellular CRF neurones. These data collectively support the hypothesis that N-PCT inhibits food intake and body weight and stimulates the HPA axis via CRF-mediated pathways.

Effects of heat treatment on the nutritional value of raw soybean selected for low trypsin inhibitor activity.

1. Two broiler experimentss and a layer experiments were conducted on Kunitz trypsin inhibitor (Kti) soybeans (SB) of low trypsin inhibitor (TI) activity to determine their nutritive value when included as mash in least-cost poultry diets. 2. Experiment 1 compared chick performance on the Kti or raw SB using a commercial full-fat SB meal (FFSBM) and a solvent extracted SB meal (SBM) as controls during a 20 d experimental period. Broiler experiment 2 compared Kti and raw SB, non-steamed, or steam-pelleted with and without DL-methionine supplementation added to every treatment containing 170 g SB/kg. For each broiler experiment the levels of each SB were 70, 120 and 170 g/kg with the control birds fed only 170 g SB/kg. 3. The layer experiment, compared steam-pelleted Kti and raw SB against a non-steamed Kti and raw SB each fed at two levels (70 and 110 g/kg) x 30 replicates from 29 weeks of age for 19 weeks in a completely randomised design. Production parameters were measured when diets were formulated to contain minimum required specifications and calculated apparent metabolisable energy (AME). At the completion of each trial, 2 broiler birds from each cage and 5 layer birds per treatment were killed, weighed, and their liver and pancreas weighed. 4. Both broiler experiments indicated that production parameters on the Kti SB treatments were significantly lower (P<0.05) than on the two commercial control SB treatments. However, the Kti treatments were superior to the raw SB treatments. 5. Pancreas weight increased with increasing inclusion of both raw and Kti SB, suggesting that a TI was causing the depression in performance. The AME of the Kti SB was similar to that of commercial FFSB meal. After steam conditioning, the raw SB meal AME value of 9.5 MJ/kg dry matter (DM) was improved to 14.1 MJ/kg DM by reduced TI activity, but this AME improvement with TI activity reduction, plus the supplementation with DL-methionine on birds fed the raw SB had no effect (P>0.05) on any parameter evaluated in experiment 2. 6. The layer experiment showed that hens on the Kti SB treatments had significantly greater live weight gain (LWG), egg weight and daily egg mass than birds given raw SB. A reduced food intake (FI) was observed in the Kti treatments but egg mass was generally similar to that on the FFSB control diet, indicating that Kti SB supported excellent egg production at an inclusion of 110 g/kg. The depressed performance observed for broiler chicks suggest that younger birds are more susceptible to the effects of SB TI.

Optimum inclusion of field peas, faba beans, chick peas and sweet lupins in poultry diets. I. Chemical composition and layer experiments.

1. Experiments were undertaken to determine the chemical composition and apparent metabolisable energy (AME) of field peas, faba beans, sweet lupins and chick peas and the production of hens when each was included in nutritionally similar diets at 250 g/kg in 2 experiments. 2. Amino acid composition, crude protein and AME agreed well with previously published measurements. Detailed analysis of the non-starch polysaccharides (NSPs) showed that sweet lupins were much higher than the other grain legumes in the soluble NSPs and that the NSPs were particularly high in arabinose. The condensed tannin content was highest in field peas followed by faba beans. 3. In the 1st layer experiment over 40 weeks, hen-day egg production was lowest on the faba bean-based diet and egg weight and egg mass were also lowest. Relative viscosity of digesta in the small intestine of hens fed on the sweet lupin-based diet was highest, followed by that of hens fed on field peas. Enlargement of the pancreas was observed in hens consuming chick peas. 4. In experiment 2, in which only sweet lupins and faba beans were used, steam or cold pelleting showed few effects, nor did dehulling of faba beans but egg weight was lower when diets were steam pelleted. Daily food intake was 5.7 g/bird lower on the steam than cold pelleted diets and food conversion ratio tended to be improved (P=0.082). 5. It was concluded that field peas could support good production at 250 g/kg of layer diet. Although chick peas and sweet lupins supported good performance, there was concern about the increased weight of the pancreas and high gut viscosity respectively. Faba beans showed similar hen-d egg production in the 2nd experiment to that of sweet lupins but egg weight tended to be about 0.8 g lower than when on the sweet lupin-based diets.

Optimum inclusion of field peas, faba beans, chick peas and sweet lupins in poultry diets. II. Broiler experiments.

1. Three experiments were undertaken to determine the optimum inclusion rates of field peas, faba beans, chick peas and sweet lupins in broiler starter and finisher diets in amounts up to 360 g/kg. 2. In experiment A chickens in cages grown to 21 d on diets with field peas and faba beans gave better growth rate and feed efficiency than those with sweet lupins and chick peas. Growth rate and Food conversion ratio (FCR) improved with increasing amounts of faba beans in the diet while for chick peas growth rate and FCR declined. Digesta viscosity and excreta stickiness scores were much higher on diets with sweet lupins. Steam pelleting improved growth rate and FCR on all diets. 3. In experiment B birds were in cages and grown from 21 to 42 d. There were no differences between grain legumes (when combined for all inclusions) for growth rate, food intake or FCR. Viscosity was again much higher on the sweet lupin-based diets while the pancreas was significantly enlarged on the diets with chick peas, as observed previously in chickens grown to 21 d. Steam pelleting of diets gave a consistent and positive response for weight gain and FCR. 4. Experiment C was carried out in pens each holding 60 birds under semi-commercial conditions and grown to 42 d on starter and finisher diets with the same grain legumes as used previously but each at 2 rates of inclusion similar to those in commercial practice. Field peas at 200 to 300 g/kg and chick peas at 150 to 220 g/kg gave inferior growth to faba beans (150 to 180 g/kg) and sweet lupins (120 g/kg). 5. The results of these experiments allowed tentative recommendations to be made to industry for inclusion rates of these cultivars of the 4 grain legumes. These were: field peas 300 g/kg; faba beans 200 g/kg, chick peas 100 g/kg and sweet lupins <100 g/kg. Wet droppings and high gut viscosity were serious problems with sweet lupins although these were not so obvious in experiment C.

Digestion of 14C-labelled condensed tannins from Desmodium intortum in sheep and goats.

An experiment was conducted to investigate the metabolism of condensed tannin (CT) in sheep and goats offered a mixture of Digitaria decumbens (700 g/kg) and Desmodium intortum (300 g/kg) hay. Radioactive 14CO2 was used to label CT in young growing desmodium plants, [14C]CT was extracted, purified and infused intraruminally, and the metabolism of [14C]CT was followed in the rumen and lower digestive tract of both species. Digestion of DM, organic matter (OM), cell-wall constituents (CWC), N and the efficiency of rumen microbial synthesis were determined using a continuous intraruminal infusion of 51Cr EDTA, YbCl3 and Na235SO4. The measurements taken for sheep and goats respectively were: intake, 21 and 30 g/kg0.9 per d; digestibilities (g/g) of DM, 0.566 and 0.505; OM 0.578 and 0.508; neutral-detergent fibre, 0.584 and 0.532; and acid-detergent fibre, 0.535 and 0.435. None of these measurements was significantly different (P > 0.05) between animal species. There was an apparent net gain in lignin across the rumen and whole intestinal tract for both animal species (19 and 29% for sheep and goats respectively). There were no significant differences between sheep and goats (P > 0.05) detected for any measurements of N excretion and utilization. The overall efficiency of N digestion and utilization was also similar between species. The routes of CT metabolism were compared for both colorimetric estimates (butanol-HCl) of dietary CT (DCT) and the specific radioactivity of [14C]CT in digesta (abomasum) and excreta (urine and faeces) of both sheep and goats. [14C]CT showed total losses of 57 and 56% in sheep and goats respectively whilst losses of DCT of 71 and 70% were detected with butanol-HCl in sheep and goats respectively. The apparent losses of DCT across the rumen of sheep and goats were 12 and 9% whilst higher losses (49 and 42% for sheep and goats respectively) were observed for [14C]CT. Losses of DCT in the lower intestinal tract accounted for 69 and 71% of the total CT leaving the abomasum. By comparison, only 40 and 35% of [14C]CT was lost during intestinal passage in sheep and goats respectively. It was concluded that the infused free [14C]CT interacted with DCT and entered the protein and fibre-bound DCT pools. The loss of DCT during passage through the intestines was considered to be a consequence of either absorption of free CT or the degradation products of CT. It was assumed that free CT arose in the lower gastrointestinal tract from protein-CT and fibre-CT dissociation to be digested and/or absorbed. The higher recoveries of [14C]CT in faeces (32 and 35%) compared with DCT (27 and 26%) for sheep and goats respectively) were associated with the excretion of [14C] degradation products or conjugates which were not reactive to butanol-HCl. It was concluded that both methods (butanol-HCl and labelling CT with 14C) detected a substantial disappearance of CT (free, protein, and fibre-bound) during metabolism in the gastrointestinal tract in sheep and goats.

The effects of condensed tannins from Desmodium intortum and Calliandra calothyrsus on protein and carbohydrate digestion in sheep and goats.

A factorial experiment was conducted to study the effects of condensed tannins (CT) from the tropical legumes Desmodium intortum and Calliandra calothyrsus on the digestion and utilization of protein and carbohydrate in sheep and goats. CT-free Centrosema pubescens was also fed for comparison with the CT legumes, and each legume was included (300 g/kg DM) in a basal diet of pangola grass (Digitaria decumbens). Pangola grass alone was used as a control diet. There were no significant (P > 0.05) differences between sheep and goats for the efficiency of digestion of N (0.574, SE 0.013), organic matter (OM; 0.519, SE 0.010), neutral-detergent fibre (NDF; 0.524, SE 0.011) and acid-detergent fibre (ADF; 0.407, SE 0.016). Diets containing desmodium and calliandra were digested less well in the rumen (64 and 62% of total OM digested) when compared with the pangola and centrosema diets (74 and 73% of total OM digested in rumen). There was an apparent net gain of 30% in ADF across the digestive tract of sheep and goats given calliandra, and this gain was ascribed to the formation of 'artifact' fibre as a result of fibre-tannin interaction. Overall, inclusion of legume at 300 g/kg in the diet significantly increased (P < 0.05) the concentration of acetic acid and decreased butyric acid concentration in the rumen fluid of sheep and goats. Significantly higher proportions of dietary N apparently reached the abomasum of animals given the diets containing desmodium (50%) and calliandra (56%) when compared with animals given the centrosema and pangola diets (35%). Sheep and goats given the CT diets also had higher excretions of faecal N. This increment of faecal N (14%) did not affect post-rumen N digestion (P > 0.05) since animals given CT diets absorbed more N (19%) per kg total OM digested than those given the control diets. It was concluded that whilst the low levels of CT provided in desmodium (1.0%) and calliandra (2.3%) diets protected dietary protein from degradation in the rumen, there were no overall beneficial or detrimental effects of CT in these diets for sheep or goats. A method was developed to categorize CT into fractions representative of their forms (free, protein-bound, and fibre-bound) during the digestion process. A quantitative model of CT metabolism during passage through the digestive tract was developed from the measured exchanges of CT between free, protein-bound and fibre-bound pools in the rumen and lower digestive tract. CT interchange mainly occurred in the reticulo-rumen of both animal species. Desmodium and calliandra free CT showed net losses of 68 and 78% in the rumen respectively and 57 and 68% of the fibre-bound CT was lost in the same site for sheep and goats respectively. However, protein-bound CT increased across the rumen by 73 and 56% for both animal species. Post-rumen losses of the total CT abomasal flow were 86 and 83% (free CT) for sheep and goats respectively, 70 and 66% (protein-bound CT), whilst 28% loss of fibre-bound CT occurred in sheep and goats respectively.

Phosphorus-containing peptides as mixed inhibitors of endopeptidase 3.4.24.15 and 3.4.24.16: effect on neurotensin degradation in vitro and in vivo.

1. We have examined several phosphorus-containing peptides as potential mixed inhibitors of two neurotensin-degrading zinc metallopeptidases, endopeptidase 3.4.24.15 and endopeptidase 3.4.24.16. 2. Among a series of 13 phosphonamide peptides, N-(2-(2-naphtyl)ethylphosphonyl-glycyl-prolyl-norleucine (phosphodiepryl 08) was found to inhibit potently the hydrolysis of neurotensin by purified endopeptidase 3.4.24.15 and 3.4.24.16 with an identical Ki value of 0.4 nM. 3. Phosphodiepryl 08 displayed a strong selectivity towards the two peptidases since it failed to inhibit several other zinc-containing peptidases such as endopeptidase 3.4.24.11, angiotensin-converting enzyme, aminopeptidase M, leucine aminopeptidase and carboxypeptidases A and B. 4. The protective effect of phosphodiepryl 08 on neurotensin degradation was examined in vitro and in vivo in central and peripheral bioassays. 5. Phosphodiepryl 08 virtually abolished neurotensin degradation by 4-day-old plated pure cultured neurones from mouse embryos and greatly potentiated neurotensin-induced antinociception in the mouse hot plate test. 6. In the periphery, phosphodiepryl 08 inhibited neurotensin degradation by membranes prepared from isolated longitudinal smooth muscle of guinea-pig ileum and greatly potentiated the neurotensin-induced contraction of the same longitudinal smooth muscle preparation. 7. Our study indicates that phosphodiepryl 08 behaves as a potent and selective mixed inhibitor of endopeptidase 3.4.24.15 and 3.4.24.16 and can be used as a powerful agent to prevent neurotensin degradation, in vitro and in vivo, in central and peripheral assays.

Antinociceptive response induced by mixed inhibitors of enkephalin catabolism in peripheral inflammation.

RB101 (N-((R,S)-2-benzyl-3[(S)(2-amino-4-methylthio)butyl dithio]-1-ox-opropyl)-L-phenylalanine benzyl ester) is a recently developed full inhibitor of the enkephalin-catabolizing enzymes able to cross the blood-brain barrier, whereas RB38A ((R)-3-(N-hydroxycarboxamido-2-benzylpropanoyl)-L-phenylalanine) is as potent as RB101 but almost unable to enter the brain. In this study, we have investigated the effects of systemic administration of morphine, RB101 and RB38A on nociception induced by pressure on inflamed peripheral tissues. Antinociceptive test was performed between 4 and 5 days after injection into the rat left hindpaw of Freund's complete adjuvant to produce localized inflammation. Morphine (1, 2 and 4 mg/kg, i.v.) induced antinociception in inflamed paws at all the doses used, and only at the highest dose in non-inflamed paws. RB101 (10 and 20 mg/kg, i.v.) induced an antinociceptive response only in the inflamed paws. RB38A, also induced an antinociceptive effect in the inflamed paws, but only at the highest dose (20 mg/kg, i.v.). The responses induced by morphine and the inhibitors of enkephalin catabolism were antagonized by the systemic administration of naloxone (1 mg/kg) or methylnaloxonium (2 mg/kg) which acts essentially outside the brain. Central injection (i.c.v.) of methylnaloxonium (2 micrograms) blocked the effect of morphine only in non-inflamed paws, and slightly decreased the response induced by RB101 on inflamed paws. These results indicate that the endogenous opioid peptides, probably enkephalins, are important in the peripheral control of nociception from inflamed tissues.

Regulation of rat glomerular epithelial cell proteoglycans by high-glucose medium.

In diabetic nephropathy the heparan sulfate proteoglycan (HSPG) content of the glomerular basement membrane (GBM) is reduced but the cellular mechanisms involved have not been studied. Glomerular epithelial cells (GEC) are thought to be the source of HSPG present in the GBM. In this study we examined if proteoglycan metabolism of the rat GEC in culture is dysregulated in a metabolic environment simulating diabetes. Following incubation for 8 days with a serum-supplemented medium containing 30 mM glucose and no added insulin, a significant increase in the overall synthesis of 35SO4-labeled molecules by the GEC was seen compared to control monolayers incubated with medium containing 5 mM glucose and insulin. Ion exchange chromatography revealed that 30 mM glucose did not alter the anionic charge density of proteoglycans, but significantly increased the amount of 35S-labeled low-anionic macromolecules in the medium; mannitol induced similar changes. Sepharose CL-4B chromatography, glycosaminoglycan analysis and immunoprecipitation of control cell layer proteoglycans demonstrated the presence of HSPG of hydrodynamic size, Kav 0.4, resembling rat GBM HSPG in size and antigenic nature. Incubation of GEC with 30 mM glucose resulted in a significant reduction (58%) in this HSPG species, an effect not seen with equimolar mannitol. Additionally, 30 mM glucose induced a significant increment in synthesis of a small HS species (Kav 0.71 on Sepharose CL-4B column) present in the cell layer. Our findings suggest that both osmotic and nonosmotic mechanisms are operative in dysregulation of glycopeptide metabolism by high-glucose medium and that reduced synthesis by the GEC may contribute to decreased content of GBM HSPG in diabetic nephropathy.

[Cicatricial, antibacterial and antimycotic effects of tepescohuite in experimental animals]. / Efectos cicatrizante, antibacteriano y antimicótico del tepescohuite en animales de experimentación.

The cicatricial and antibacterial effects of the sterile powder of the barks of tepescohuite (Mimosa tenuiflora), 2% mupirocin ointment, and 0.9% saline were compared. The experiment was performed in rabbits with chemically induced burns clinically, histopathologically, bacteriologically, and mycologically controlled. No statistically significant difference was found among the three treatment modalities. Due to the potentially hepatotoxic effects and low therapeutic efficacy of tepescohuite it should not be used in human beings.