Effects of feeding a quebracho-chestnut tannin extract on lactating cow performance and nitrogen utilization efficiency.

The effects of feeding a quebracho-chestnut tannin extract mixture on performance and nitrogen (N) utilization were assessed with 36 multiparous lactating Holstein cows (mean ± standard deviation; 706 ± 59 kg of body weight; 126 ± 20 d in milk) randomly assigned to 3 dietary treatments in a randomized complete block design. Following a 2-wk covariate adjustment period, cows were fed their assigned treatment diets for 13 wk. Rice hulls were removed from a total mixed ration with a 54:46 forage:concentrate ratio (% of dry matter; DM), and a tannin extract mixture from quebracho and chestnut trees (2:1 ratio) was included at 0, 0.45, and 1.80% of dietary DM. There was no interaction between dietary treatments and experimental week for the reported measurements except milk lactose percentage. Overall, treatments did not affect milk yield (48.6 ± 7.8 kg/d), fat- and protein-corrected milk (46.1 ± 7.6 kg/d), milk fat content (3.88 ± 0.65%) and yield (1.85 ± 0.38 kg/d), and true protein yield (1.45 ± 0.21 kg/d). However, incremental levels of tannin extracts in the diet produced a linear increase in DM intake (29.2 to 30.9 kg/d) and a linear decrease in kilograms of milk per kilogram of DM intake (1.67 to 1.57 kg/kg) and MUN (12.2 to 10.8 mg/dL). Furthermore, there was a quadratic effect of tannin extracts on milk true protein content (2.96, 3.13, and 3.00% for 0, 0.45, and 1.80% tannin extract, respectively) and a tendency for linear and quadratic response for body weight gain (0.31, 0.16, and 0.44 kg/d for 0, 0.45, and 1.80% tannin, respectively). Intake of N increased linearly (782, 795, and 820 g/d) and N utilization efficiency (milk N/intake N) decreased linearly (0.300, 0.301, and 0.275 for 0, 0.45, and 1.80% tannin, respectively). Relative to the 0% diet, 1.80% tannin extract reduced estimated urinary N excretion by 11%. In this study, adding 0.45% tannin extract to the diet reduced feed efficiency but had a positive effect on milk protein content. Feeding a tannin extract mixture from quebracho and chestnut may reduce environmental labile urinary N excretion without affecting milk yield but at the expense of a lower feed utilization efficiency.

Short communication: Identifying challenges and opportunities for improved nutrient management through the USDA's Dairy Agroecosystem Working Group.

Nutrient management on US dairy farms must balance an array of priorities, some of which conflict. To illustrate nutrient management challenges and opportunities across the US dairy industry, the USDA Agricultural Research Service Dairy Agroecosystems Working Group (DAWG) modeled 8 confinement and 2 grazing operations in the 7 largest US dairy-producing states using the Integrated Farm System Model (IFSM). Opportunities existed across all of the dairies studied to increase on-farm feed production and lower purchased feed bills, most notably on large dairies (>1,000 cows) with the highest herd densities. Purchased feed accounted for 18 to 44% of large dairies' total operating costs compared with 7 to 14% on small dairies (<300 milk cows) due to lower stocking rates. For dairies with larger land bases, in addition to a reduction in environmental impact, financial incentives exist to promote prudent nutrient management practices by substituting manure nutrients or legume nutrients for purchased fertilizers. Environmental priorities varied regionally and were principally tied to facility management for dry-lot dairies of the semi-arid western United States (ammonia-N emissions), to manure handling and application for humid midwestern and eastern US dairies (nitrate-N leaching and P runoff), and pasture management for dairies with significant grazing components (nitrous oxide emissions). Many of the nutrient management challenges identified by DAWG are beyond slight modifications in management and require coordinated solutions to ensure an environmentally and economically sustainable US dairy industry.

Invited review: Enteric methane in dairy cattle production: quantifying the opportunities and impact of reducing emissions.

Many opportunities exist to reduce enteric methane (CH4) and other greenhouse gas (GHG) emissions per unit of product from ruminant livestock. Research over the past century in genetics, animal health, microbiology, nutrition, and physiology has led to improvements in dairy production where intensively managed farms have GHG emissions as low as 1 kg of CO2 equivalents (CO2e)/kg of energy-corrected milk (ECM), compared with >7 kg of CO2 e/kg of ECM in extensive systems. The objectives of this review are to evaluate options that have been demonstrated to mitigate enteric CH4 emissions per unit of ECM (CH4/ECM) from dairy cattle on a quantitative basis and in a sustained manner and to integrate approaches in genetics, feeding and nutrition, physiology, and health to emphasize why herd productivity, not individual animal productivity, is important to environmental sustainability. A nutrition model based on carbohydrate digestion was used to evaluate the effect of feeding and nutrition strategies on CH4/ECM, and a meta-analysis was conducted to quantify the effects of lipid supplementation on CH4/ECM. A second model combining herd structure dynamics and production level was used to estimate the effect of genetic and management strategies that increase milk yield and reduce culling on CH4/ECM. Some of these approaches discussed require further research, but many could be implemented now. Past efforts in CH4 mitigation have largely focused on identifying and evaluating CH4 mitigation approaches based on nutrition, feeding, and modifications of rumen function. Nutrition and feeding approaches may be able to reduce CH4/ECM by 2.5 to 15%, whereas rumen modifiers have had very little success in terms of sustained CH4 reductions without compromising milk production. More significant reductions of 15 to 30% CH4/ECM can be achieved by combinations of genetic and management approaches, including improvements in heat abatement, disease and fertility management, performance-enhancing technologies, and facility design to increase feed efficiency and life-time productivity of individual animals and herds. Many of the approaches discussed are only partially additive, and all approaches to reducing enteric CH4 emissions should consider the economic impacts on farm profitability and the relationships between enteric CH4 and other GHG.

Anaphylaxis: mechanisms and management.

Anaphylaxis is a serious allergic reaction that can be rapid in onset and occasionally fatal. This is a review of the literature on the epidemiology of anaphylaxis, diagnostic criteria, mechanisms, and management. This current review of the literature is intended for clinicians, trainees, and researchers as a comprehensive review of anaphylaxis with a focus on incorporating the latest advances in the field.

Estimating phosphorus loss in runoff from manure and fertilizer for a phosphorus loss quantification tool.

Nonpoint-source pollution of fresh waters by P is a concern because it contributes to accelerated eutrophication. Given the state of the science concerning agricultural P transport, a simple tool to quantify annual, field-scale P loss is a realistic goal. We developed new methods to predict annual dissolved P loss in runoff from surface-applied manures and fertilizers and validated the methods with data from 21 published field studies. We incorporated these manure and fertilizer P runoff loss methods into an annual, field-scale P loss quantification tool that estimates dissolved and particulate P loss in runoff from soil, manure, fertilizer, and eroded sediment. We validated the P loss tool using independent data from 28 studies that monitored P loss in runoff from a variety of agricultural land uses for at least 1 yr. Results demonstrated (i) that our new methods to estimate P loss from surface manure and fertilizer are an improvement over methods used in existing Indexes, and (ii) that it was possible to reliably quantify annual dissolved, sediment, and total P loss in runoff using relatively simple methods and readily available inputs. Thus, a P loss quantification tool that does not require greater degrees of complexity or input data than existing P Indexes could accurately predict P loss across a variety of management and fertilization practices, soil types, climates, and geographic locations. However, estimates of runoff and erosion are still needed that are accurate to a level appropriate for the intended use of the quantification tool.

Modeling phosphorus losses from soils amended with cattle manures and chemical fertilizers.

While applied manure/fertilizer is an important source of P loss in surface runoff, few models simulate the direct transfer of phosphorus (P) from soil-surface-applied manure/fertilizer to surface runoff. The SurPhos model was tested with 2008-2010 growing season daily surface runoff data from clay loam experimental plots subject to different manure/fertilizer applications. Model performance was evaluated on the basis of the coefficient of determination (R2), Nash-Sutcliffe efficiency (NSE), percent bias (PBIAS), and the ratio of the root mean square error to the standard deviation of observed values (RSR). The model offered an acceptable performance in simulating soil labile P dynamics (R2 = 0.75, NSE = 0.55, PBIAS = 10.43%, and RSR = 0.67) and dissolved reactive P (DRP) loss in surface runoff (R2 ≥ 0.74 and NSE ≥ 0.69) for both solid and liquid cattle manure, as well as inorganic fertilizer. Simulated direct P loss in surface runoff from solid and liquid cattle manure accounted for 39% and 40% of total growing season DRP losses in surface runoff. To compensate for the unavailability of daily surface runoff observations under snow melt condition, the whole four years' (2008-2011) daily surface runoff predicted by EPIC (Environmental Policy Integrated Climate) was used as SurPhos input. The accuracy of simulated DRP loss in surface runoff under the different manure/fertilizer treatments was acceptable (R2 ≥ 0.55 and NSE ≥ 0.50). For the solid cattle manure treatment, of all annual DRP losses, 19% were derived directly from the manure. Beyond offering a reliable prediction of manure/fertilizer P loss in surface runoff, SurPhos quantified different sources of DRP loss and dynamic labile P in soil, allowing a better critical assessment of different P management measures' effectiveness in mitigating DRP losses.

A model for phosphorus transformation and runoff loss for surface-applied manures.

Agricultural P transport in runoff is an environmental concern. An important source of P runoff is surface-applied, unincorporated manures, but computer models used to assess P transport do not adequately simulate P release and transport from surface manures. We developed a model to address this limitation. The model operates on a daily basis and simulates manure application to the soil surface, letting 60% of manure P infiltrate into soil if manure slurry with less than 15% solids is applied. The model divides manure P into four pools, water-extractable inorganic and organic P, and stable inorganic and organic P. The model simulates manure dry matter decomposition, and manure stable P transformation to water-extractable P. Manure dry matter and P are assimilated into soil to simulate bioturbation. Water-extractable P is leached from manure when it rains, and a portion of leached P can be transferred to surface runoff. Eighty percent of manure P leached into soil by rain remains in the top 2 cm, while 20% leaches deeper. This 2-cm soil layer contributes P to runoff via desorption. We used data from field studies in Texas, Pennsylvania, Georgia, and Arkansas to build and validate the model. Validation results show the model accurately predicted cumulative P loads in runoff, reflecting successful simulation of the dynamics of manure dry matter, manure and soil P pools, and storm-event runoff P concentrations. Predicted runoff P concentrations were significantly related to (r2=0.57) but slightly less than measured concentrations. Our model thus represents an important modification for field or watershed scale models that assess P loss from manured soils.

Effect of methodology in estimating and interpreting water-extractable phosphorus in animal manures.

Manure water-extractable phosphorus (WEP) data are used in indices and models to assess P transport in runoff. Methods to measure WEP vary widely, often without understanding the effect on how much P is extracted. We conducted water extractions on five dairy, swine, and poultry manures to assess single and sequential extractions, drying manures, solution to solid (cm3 g(-1)) extraction ratios, and P determination method. We found little difference in WEP of single or sequential extractions. Increasing extraction ratio from 10:1 to 250:1 resulted in more WEP recovered, but in a diminishing fashion so that ratios of 200:1 and 250:1 were not significantly different. Patterns of increased WEP with extraction ratio varied with manure type, presence of bedding material, and drying treatment. Fresh and air-dried manures had similar patterns, but differed substantially from oven-dried (90 degrees C) manures. The differential effect of oven-drying on WEP was greatest for dairy and poultry manure, and less for swine manure. We analyzed water extracts colorimetrically before and after digestion, to examine the potential effect of P determination by inductively coupled plasma (ICP) spectroscopy. Digested extracts always contained more P. For manures with bedding, drying decreased the difference in P measured before and after digestion. The opposite was true for manures without bedding. Results highlight the influence of methodology on manure WEP measurement and caution needed when comparing data across studies using different WEP methods. Overall, our results point to a need for a standard manure water extraction method.

Group II phospholipases A2 are indirectly cytolytic in the presence of exogenous phospholipid.

Systemic inflammatory response syndromes including septic shock and salicylate poisoning are associated with high circulating levels of secretory phospholipase A2 (sPLA2). In septic shock, sPLA2 has been implicated in the pathogenesis of multisystem organ failure, presumably by a direct cytotoxic effect on cells. The cytotoxicity of recombinant human sPLA2 and a venom PLA2 were examined on human erythrocytes, erythroleukemia cells and U937 cells. Neither the human nor venom PLA2's were directly injurious to target cells. However, in the presence of liposomal phospholipids, both PLA2's induced irreversible cell injury. Whereas venom PLA2 was cytolytic in the presence of either phosphatidylcholine or phosphatidylethanolamine (PE), rh-sPLA2 caused cell death only in the presence of PE. These data show that normal unperturbed cells are resistant to injury from PLA2, and that additional cofactors such as PE are required to induce cell injury.

Regulation of the cellular expression of secretory and cytosolic phospholipases A2, and cyclooxygenase-2 by peptide growth factors.

Secretory group II (sPLA2) and cytosolic (cPLA2) phospholipases A2 and cyclooxygenase-2 (Cox-2) play a pivotal role in release of proinflammatory eicosanoids. Excessive activity of sPLA2 per se can also propagate inflammation. Endogenous control of the above enzymes has not been completely elucidated. We investigated the combined impact of promoting cytokines and inhibitory peptide growth factors on the expression of mRNA of the above enzymes, on protein content and extracellular release of sPLA2 and on PGE2 production in osteoblasts (FRCO). The synthesis and release of sPLA2 were enhanced by about 20-fold by 0.5 ng/ml IL-1beta or by 50 ng/ml of TNFalpha. Coaddition of both cytokines resulted in synergistic 150-fold increase in the release of sPLA2 implying the existence of two paths of induction. IL-1beta and TNFalpha markedly enhanced the transcription of sPLA2 mRNA. Kinetic study showed that IL-1/TNF initiated sPLA2 release after 12 h, reaching maximum at 48 h. IL-1alpha was a weak stimulator of sPLA2 release, whereas IL-6, IL-8, IGF, IFN-gamma, growth hormone, insulin and GM-CSF were not stimulatory. Peptide growth hormones TGFbeta, PDGF-BB, EGF and bFGF markedly inhibited the extracellular release of sPLA2. TGFbeta and PDGF-BB significantly reduced the level of sPLA2 mRNA, thus acting upon transcription whereas EGF and bFGF were not inhibitory, acting rather upon the translational or posttranslational steps. IL-1/TNF and growth factors had no significant effect on cPLA2 mRNA expression. Cox-2 mRNA expression was markedly enhanced by IL-1/TNF and suppressed by all growth factors tested. Cytokines enhanced the extracellular release of PGE2 and further enhancement was induced by growth factors with the exception of TGFbeta. Cycloheximide abolished completely the release of sPLA2 and markedly reduced the release of PGE2 from cytokine-stimulated FRCO, regardless of whether growth factors were present or not. NS-398, a specific inhibitor of Cox-2 abolished almost completely the release of PGE2 from cytokine-stimulated cells, regardless of the presence of growth factors. Thus, different signalling mechanisms are involved in the impact of growth factors on mRNA expression of sPLA2, cPLA2 and Cox-2. The differences between the impact on FRCO sPLA2 and that reported in other cells, imply that endogenous control of arachidonic acid cascade is cell-specific.

Predicting dissolved phosphorus in runoff from manured field plots.

Dissolved inorganic P transport in runoff from agricultural soils is an environmental concern. Models are used to predict P transport but rarely simulate P in runoff from surface-applied manures. Using field-plot data, we tested a previously proposed model to predict manure P in runoff. We updated the model to include more data relating water to manure ratio to manure P released during water extractions. We verified that this update can predict P release from manure to rain using published data. We tested the updated model using field-plot and soil-box data from three manure runoff studies. The model accurately predicted runoff P for boxes, but underpredicted runoff P for plots. Underpredictions were caused by runoff to rain ratios used to distribute P into runoff or infiltration. We developed P distribution fractions from manure water extraction data to replace runoff to rain ratios. Calculating P distribution fractions requires knowing rainfall rate and times that runoff begins and rain stops. Using P distribution fractions gave accurate predictions of runoff P for soil boxes and field plots. We observed relationships between measured runoff to rain ratios and both P distribution fractions and a degree of error in original predictions, calculated as (measured runoff P/predicted runoff P). Using independent field-plot data, we verified that original underpredictions of manure runoff P can be improved by calculating P distribution fractions from measured runoff to rain ratios or adjusting runoff to rain ratios based on their degree of error. Future work should test the model at field or watershed scales and at longer time scales.

Relating soil phosphorus to dissolved phosphorus in runoff: a single extraction coefficient for water quality modeling.

Phosphorus transport from agricultural soils contributes to eutrophication of fresh waters. Computer modeling can help identify agricultural areas with high potential P transport. Most models use a constant extraction coefficient (i.e., the slope of the linear regression between filterable reactive phosphorus [FRP] in runoff and soil P) to predict dissolved P release from soil to runoff, yet it is unclear how variations in soil properties, management practices, or hydrology affect extraction coefficients. We investigated published data from 17 studies that determined extraction coefficients using Mehlich-3 or Bray-1 soil P (mg kg(-1)), water-extractable soil P (mg kg(-1)), or soil P sorption saturation (%) as determined by ammonium oxalate extraction. Studies represented 31 soils with a variety of management conditions. Extraction coefficients from Mehlich-3 or Bray-1 soil P were not significantly different for 26 of 31 soils, with values ranging from 1.2 to 3.0. Extraction coefficients from water-extractable soil P were not significantly different for 17 of 20 soils, with values ranging from 6.0 to 18.3. The relationship between soil P sorption saturation and runoff FRP (microg L(-1)) was the same for all 10 soils investigated, exhibiting a split-line relationship where runoff FRP rapidly increased at P sorption saturation values greater than 12.5%. Overall, a single extraction coefficient (2.0 for Mehlich-3 P data, 11.2 for water-extractable P data, and a split-line relationship for P sorption saturation data) could be used in water quality models to approximate dissolved P release from soil to runoff for the majority of soil, hydrologic, or management conditions. A test for soil P sorption saturation may provide the most universal approximation, but only for noncalcareous soils.

Interleukin-1 alpha stimulates the release of prostaglandin E2 and phospholipase A2 from fetal rat calvarial cells in vitro: relationship to bone nodule formation.

We have shown previously that interleukin-1 (IL-1) has biphasic effects on the formation of bone nodules in long-term cultures of fetal rat calvarial (RC) cells (Ellies and Aubin, Cytokine 2:430-437, 1990). To determine the role of arachidonic acid metabolism in this process, we have examined the release of prostaglandin E2 (PGE2) and phospholipase A2 (PLA2) from RC cells under conditions that allowed concomitant analysis of the formation of bone nodules. Recombinant human IL-1 alpha (rhIL-1 alpha) stimulated PGE2 and PLA2 release in a time- and dose-dependent manner. PGE2 release was highest in preconfluent cultures (days 1-6) and was stimulated up to 8.5-fold in response to 50 U/ml of rhIL-1 alpha. In contrast, extracellular PLA2 activity was maximal in postconfluent cultures, with 50 U/ml of rhIL-1 alpha causing a 20-fold increase by day 15. PLA2 release by RC cells was not significantly affected by PGE2, the glucocorticoid dexamethasone, or the cyclooxygenase inhibitor indomethacin. Indomethacin partially blocked the inhibition of bone nodule formation caused by rhIL-1 alpha, and exogenous PGE2 reversed this effect. Addition of group I PLA2 from Naja naja venom to RC cells had no effect on bone nodule development; however, group II PLA2 from Crotalus adamanteus venom inhibited the formation of bone nodules in a dose range similar to that induced by rhIL-1 alpha. These results indicate that PGE2 release does not have a direct temporal correlation with increases in PLA2 activity. In addition, the data show that only part of the inhibition of bone formation seen with rhIL-1 alpha is mediated by PGE2 and suggest that extracellular PLA2 also accounts for part of the inhibition.

Induction of circulating phospholipase A2 activity by intravenous infusion of endotoxin in patients with neoplasia.

The purpose of this study was to evaluate the impact of repeated intravenous infusions of endotoxin (EN) in patients with cancer on the systemic release of extracellular proinflammatory phospholipase A2 (PLA2) and its relationship to the release of tumor necrosis factor (TNF) and interleukin-6 (IL-6). Six patients received 15 infusion of EN isolated from Salmonella abortus equi at a dose of 4 ng/kg. Marked increase in the activity of circulating PLA2 was noted within 3 h after the first EN infusion and reached a maximal level of 20.4-fold greater than baseline 24 h after infusion. In five patients challenged with EN 2 weeks later, PLA2 reached peak levels 15.5-fold greater than baseline. In two patients who received three sequential daily infusions, the incremental increase in PLA2 activity after the second and third challenge reached maximum levels 6 h after EN infusion. PLA2 response followed those of TNF and IL-6 but was quantitatively different. Whereas maximal levels of TNF and IL-6 declined substantially after repeat EN challenges, no such decline occurred in PLA2 activity. Since, in the clinical setting of gram-negative sepsis, there is recurrent increase in circulating EN, our study approximates this clinical situation and shows that extracellular release of PLA2 follows temporally that of proximal cytokines such as TNF and IL-6. These cytokines may be related to PLA2 release and sustained high activity in the systemic circulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Inflammatory effect of intradermal administration of soluble phospholipase A2 in rabbits.

Extracellular phospholipase A2 (PLA2) has been found in association with inflamed sites in experimental animals and in humans. The tissue effects of soluble PLA2 have not been defined. We studied the development of inflammatory changes in rabbit skin subsequent to intradermal injection of active and inactivated venom and pancreatic PLA2, over a broad concentration range. PLA2, at concentrations encountered in human disease, caused acute inflammatory changes characterized grossly by erythema and induration, and histologically by inflammatory cell infiltration, vascular and tissue damage, and abscess formation. Extracellular PLA2 may be considered as one of the pathogenic factors in inflammatory reaction.

Extracellular phospholipase A2 secretion is a common effector pathway of interleukin-1 and tumour necrosis factor action.

Inflammatory processes are characterized by increased levels of extracellular phospholipase A2 (PLA2) and cytokines such as interleukin 1 (IL-1) and tumour necrosis factor (TNF). IL-1, TNF and PLA2 share a number of proinflammatory, arthritogenic effects. The sequential induction, first of the cytokines followed by PLA2, suggests that these cytokines may regulate synthesis and secretion of PLA2. To test this postulate, foetal rat calvarial bone-forming cells (FRCC) were treated with recombinant human IL-1 and TNF and extracellular PLA2 release was quantitated. Both IL-1 and TNF induced the de novo synthesis of PLA2 in a concentration-dependent manner. Continuous exposure of FRCC in primary culture to IL-1 (50 units/ml) over 15 days resulted in as much as 100-fold increase in PLA2 secretion. IL-1 (50 units/ml) added to post-confluent cultures for a 48-h pulse increased PLA2 activity 9.4-fold. The combination of IL-1 (50 units/ml) and TNF (500 units/ml) was synergistic with an observed increase in extracellular PLA2 secretion of 146-fold following a 48-h pulse. Interleukin-6, alone or in combination with IL-1 or TNF, did not further enhance PLA2 synthesis of secretion. Cytokine-induced synthesis of PLA2 was inhibited 80% by 10 microM cycloheximide but not by dexamethasone over the range of 10(-6) to 10(-8) M. FRCC-derived PLA2 was neutral-active with a pH optimum of 6-7.5 and was calcium-dependent with optimal activity in the presence of 2-7 mM calcium. It had absolute 2-acyl specificity using micellar phosphatidylcholine.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibition of extracellular release of proinflammatory secretory phospholipase A2 (sPLA2) by sulfasalazine: a novel mechanism of anti-inflammatory activity.

Sulfasalazine is widely used in rheumatoid arthritis and inflammatory bowel diseases. The mechanisms of its activity have not been elucidated. In leukocytes, sulfasalazine and its analogue, CL 42A, inhibited the formation of leukotrienes and possibly of the second messenger compounds at the level of phospholipase C. Partial inhibition of interleukin-lbeta (IL-1beta), IL-6 and tumor necrosis factor-alpha (TNF-alpha) was also found. Since the synthesis of eicosanoids is induced by phospholipase A2 and since secretory phospholipase A2 (sPLA2) is proinflammatory, we investigated the impact of sulfasalazine and related compounds on mRNA, protein synthesis, and release of sPLA2 from osteoblasts. Sulfasalazine and CL 42A markedly inhibited extracellular release of sPLA2. The impact of sulfasalazine was evident at 50 microM (P < 0.001) and maximal at 400 microM, and that of CL 42A at 10 microM (P < 0.001) and 200 microM, respectively. Split products of sulfasalazine, 5-aminosalicylic acid (400 microM) and sulfapyridine (400 microM), had no impact. The effect of sulfasalazine and CL 42A was evident regardless of whether the cells were stimulated with IL-1beta/TNF-alpha, lipopolysaccharide/forskolin, or dibutyryl-cAMP. Sulfasalazine and CL 42A did not alter the level of sPLA2 mRNA. Exposure of stimulated fetal rat calvaria osteoblasts (FRCO) to sulfasalazine did not show accumulation of the intracellular sPLA2 protein as tested by western blot; however, enzymatic activity of PLA2 in disrupted cells was definitely increased. Thus, the impact is on the post-transcriptional release of sPLA2 rather than on the synthesis. There was also an increase in the extracellular release of prostaglandin E2 from FRCO exposed to sulfasalazine or to CL 42A. In contrast, sulfasalazine had no effect on the extracellular release of gelatinase from the cells or on mRNA of cytosolic PLA2 or cyclooxygenase 2. We conclude that the anti-inflammatory activity of sulfasalazine may be related, in part, to the selective inhibition of the extracellular release of proinflammatory sPLA2.

Inhibition of enzymatic activity of phospholipases A2 by minocycline and doxycycline.

Extracellular phospholipases A2 play an important role in articular and extra-articular inflammatory processes. Secretory non-pancreatic phospholipase A2 (PLA2) has been implicated in the pathogenesis of articular inflammation in rheumatoid arthritis, whereas pancreatic PLA2 contributes to the tissue damage associated with acute pancreatitis. Since in experimental models lipophilic tetracyclines such as minocycline and doxycycline are antiinflammatory, we examined their effects on PLA2 activity using two assay systems in vitro. We found that minocycline and to a lesser degree doxycycline were markedly inhibitory to both pancreatic and non-pancreatic PLA2. Using [14C]oleic acid labeled Escherichia coli membrane phospholipids as substrate, the IC50 values for minocycline and doxycycline were 3.6 x 10(-5) M (18 micrograms/mL) and 0.98 x 10(-4) M (47 micrograms/mL), respectively. In a scooting mode assay using the synthetic phospholipid 1-palmitoyl-2-(10-pyrenedecanoyl)-3-L-phosphatidylmethanol as substrate, IC50 values for minocycline were 5 microM (2.47 micrograms/mL) for non-pancreatic PLA2 and 8 microM (3.95 micrograms/mL) for pancreatic PLA2. Addition of excess calcium up to 50 mM did not reverse the inhibitory activity of tetracyclines. We conclude that lipophilic tetracyclines inhibit PLA2, probably by interaction with the substrate, and may be a useful adjunct in the therapy of inflammatory conditions in which PLA2 is implicated pathogenetically.

Benefits of access to on-site acute and critical care for the residential section of a multi-level geriatric center.

Of 220 residents living in the residential unit of a multi-level geriatric center, 95 required a total of 131 transfers to other facilities over a one-year period. Fifty-three of these patients were transferred to the associated chronic-care hospital for short-term acute medical investigation or treatment. The average length of stay of 17 days was similar to that of patients of comparable age with comparable medical conditions admitted to an affiliated acute general hospital from the community. In addition to continuity of medical care and the social and psychologic advantages of remaining within the geriatric center, this transfer potential resulted in substantial financial savings. The cost of hospitalization in the general hospital would have been more than twice that of the geriatric unit.

Purification of a soluble phospholipase A2 from synovial fluid in rheumatoid arthritis.

A soluble phospholipase A2 (PLA2) was purified 4,500-fold from human rheumatoid synovial fluid. Preparative sodium dodecyl sulfate polyacrylamide gel electrophoresis yielded two bands of PLA2 activity of molecular weights 15,000 and 17,000 and pl 4.2-5.0. Purified PLA2 had absolute 2-acyl specificity, and hydrolyzed phosphatidylcholine with optimal activity at pH 7.5-8.0 and phosphatidylethanolamine with optimal activity at pH 7.0. Human synovial fluid PLA2 did not cross-react with anti-human pancreatic PLA2, as tested by radioimmunoassay.