SAP Levels Research Articles

P53 contributes to T cell homeostasis through the induction of pro-apoptotic SAP

Lack of functional SAP protein, due to gene deletion or mutation, is the cause of X-linked lymphoproliferative disease (XLP), characterized by functionally impaired T and NK cells and a high risk of lymphoma development. We have demonstrated earlier that SAP has a pro-apoptotic function in T and B cells. Deficiency of this function might contribute to the pathogenesis of XLP. We have also shown that SAP is a target of p53 in B cell lines. In the present study, we show that activated primary T cells express p53, which induces SAP expression. p53 is functional as a transcription factor in activated T cells and induces the expression of p21, PUMA and MDM2. PARP cleavage in the late phase of activation indicates that T cells expressing high levels of SAP undergo apoptosis. Modifying p53 levels using Nutlin-3, which specifically dissociates the MDM2-p53 interaction, was sufficient to upregulate SAP expression, indicating that SAP is a target of p53 in T cells. We also demonstrated p53’s role as a transcription factor for SAP in activated T cells by ChIP assays. Our result suggests that p53 contributes to T cell homeostasis through the induction of the pro-apoptotic SAP. A high level of SAP is necessary for the activation-induced cell death that is pivotal in termination of the T cell response.

Protease Activated Receptor 2 (PAR-2) Promotes Vascular Inflammation in a Mouse Model of Sickle Cell Disease

Abstract 375Sickle cell disease (SCD) is a hematologic disorder caused by a single nucleotide mutation of the beta-globin gene. It is associated with increased tissue factor (TF) expression, activation of coagulation and chronic vascular inflammation. Using two mouse models of SCD (BERK and Townes mice), we have recently demonstrated that inhibition of TF with a rat anti-mouse TF (1H1) antibody not only abolishes activation of coagulation (measured by plasma levels of thrombin anti-thrombin (TAT) complexes) but also reduces inflammation and endothelial cell (EC) injury, indicated by attenuation of plasma levels of IL-6 and sVCAM-1, respectively. Furthermore, we showed that EC-specific deletion of TF gene significantly reduced plasma levels of IL-6 but had no effect on activation of coagulation (TAT) or EC injury (sVCAM-1). These data suggest that EC-TF is primarily involved in signaling rather than activation of coagulation. Since TF:factor VIIa complex-dependent activation of protease activated receptor-2 (PAR-2) has been shown to promote inflammation, we have now investigated the role of PAR-2 expressed by non-hematopoietic cells in the pathology of SCD.PAR-2+/+ and PAR-2−/− mice were lethally irradiated and transplanted with bone marrow from BERK SS (sickle cell mice) or BERK AA (non-sickle control) mice(n=6–10). Four months after bone marrow transplantation, mice were sacrificed and the reconstitution of bone marrow was confirmed by electrophoretic analysis of the different forms of hemoglobin. PAR-2+/+ mice transplanted with bone marrow from BERK SS mice had reduced number of red blood cells and hematocrit compared to PAR-2+/+ mice transplanted with bone marrow from BERK AA mice. PAR-2 deficiency in all non-hematopoietic cells had no effect on these hematologic parameters. Furthermore, PAR-2+/+ mice transplanted with bone marrow from BERK SS mice had increased number of monocytes (3.1 fold, p<0.0001) and neutrophils (2.5 fold, p<0.05) in the blood. Interestingly, sickle cell mice lacking PAR-2 in non-hematopoietic cells had significantly reduced neutrophil counts compared to the sickle cell mice with normal levels of PAR-2 (1.9+/−0.2 vs. 5.4+/−1.4 X103/ul; p<0.05), whereas monocytes counts were not affected. Compared to non-sickle controls, sickle cell mice had increased plasma levels of TAT (1.9 fold, p<0.01), IL-6 (6.8 fold, P<0.0001), serum amyloid protein SAP (6.5 fold, p<0.01; mouse homolog of human C reactive protein) and sVCAM-1 (1.4 fold, p<0.01). Moreover, increased levels of myeloperoxidase (MPO) were observed in the livers of sickle cell mice (3 fold, p<0.0001). Importantly, sickle cell mice lacking PAR-2 expression in all non-hematopoietic cells demonstrated significant reduction of plasma levels of IL-6 (9.4+/−0.9 vs. 18.9+/−4.5 pg/ml; p<0.05) and SAP (60.5+/−12.9 vs. 182.8+/−62.5ug/ml; p<0.05) compared to sickle cell mice with normal levels of PAR2 expression. In addition, deletion of PAR-2 also significantly reduced MPO levels in the liver of sickle cell mice (53.7+/−3.5 vs. 117.4+/−16.9 ng/mg protein; p<0.0001). In contrast, PAR-2 deficiency in non-hematopoietic cells had no effect on activation of coagulation (TAT) or EC injury (sVCAM-1) in sickle mice.Our data demonstrate that vascular inflammation observed in a mouse model of sickle cell disease is mediated, in part, by PAR-2 expressed by non-hematopoietic cells. Activation of EC (sVCAM-1) was not affected by PAR-2 deficiency. Ongoing studies are investigating the possible contribution of the TF-thrombin-PAR1 pathway to the EC activation in SCD. Disclosures:No relevant conflicts of interest to declare.

Alkaptonuria is a novel human secondary amyloidogenic disease

Alkaptonuria (AKU) is an ultra-rare disease developed from the lack of homogentisic acid oxidase activity, causing homogentisic acid (HGA) accumulation that produces a HGA-melanin ochronotic pigment, of unknown composition. There is no therapy for AKU. Our aim was to verify if AKU implied a secondary amyloidosis. Congo Red, Thioflavin-T staining and TEM were performed to assess amyloid presence in AKU specimens (cartilage, synovia, periumbelical fat, salivary gland) and in HGA-treated human chondrocytes and cartilage. SAA and SAP deposition was examined using immunofluorescence and their levels were evaluated in the patients' plasma by ELISA. 2D electrophoresis was undertaken in AKU cells to evaluate the levels of proteins involved in amyloidogenesis. AKU osteoarticular tissues contained SAA-amyloid in 7/7 patients. Ochronotic pigment and amyloid co-localized in AKU osteoarticular tissues. SAA and SAP composition of the deposits assessed secondary type of amyloidosis. High levels of SAA and SAP were found in AKU patients' plasma. Systemic amyloidosis was assessed by Congo Red staining of patients' abdominal fat and salivary gland. AKU is the second pathology after Parkinson's disease where amyloid is associated with a form of melanin. Aberrant expression of proteins involved in amyloidogenesis has been found in AKU cells. Our findings on alkaptonuria as a novel type II AA amyloidosis open new important perspectives for its therapy, since methotrexate treatment proved to significantly reduce in vitro HGA-induced A-amyloid aggregates.

Spatial Variation of Surface Soil Available Phosphorous and Its Relation with Environmental Factors in the Chaohu Lake Watershed

The study presented in this paper attempts to evaluate the spatial pattern of soil available phosphorus, as well as the relation between soil available phosphorus and environment factors including elevation, slope, precipitation, percentage of cultivated land, percentage of forest land, percentage of construction land and NDVI using statistical methods and GIS spatial analysis techniques. The results showed that the Spline Tension method performed the best in the prediction of soil available phosphorus in the Chaohu Lake watershed. The spatial variation of surface soil available phosphorus was high in Chaohu Lake watershed and the upstream regions around Chaohu Lake, including the west of Chaohu lake (e.g., southwest of Feixi county, east of Shucheng county and north of Lujiang county) and to the north of Chaohu Lake (e.g., south of Hefei city, south of Feidong county, southwest of Juchao district), had the highest soil available phosphorus content. The mean and standard deviation of soil available phosphorus content gradually decreased as the elevation or slope increased. The cultivated land comprised 60.11% of the watershed and of that land 65.63% belonged to the medium to very high SAP level classes, and it played a major role in SAP availability within the watershed and a potential source of phosphorus to Chaohu Lake resulting in eutrophication. Among the land use types, paddy fields have some of the highest maximum values and variation of coefficients. Subwatershed scale soil available phosphorus was significantly affected by elevation, slope, precipitation, percentage of cultivated land and percentage of forest land and was decided by not only these environmental factors but also some other factors such as artificial phosphorus fertilizer application.

Spectrophotometric analysis of the expression of secreted aspartyl proteinases from Candida in leukoplakia and oral squamous cell carcinoma

Candida species are a normal commensal of the oral cavity in healthy individuals, but can become an opportunistic pathogen when the oral ecosystem is unbalanced. Several virulence attributes have been identified in candidal infection, among which are the hydrolases, including the secreted aspartyl proteinases (Saps). This study evaluated and compared the in vitro level of Saps from Candida albicans in nonsmokers, smokers, and patients with leukoplakia and oral squamous cell carcinoma (OSCC). Candida cell count (CCC) at 48 h was also assessed. The Sap level was measured by spectrophotometry in 38 clinical isolates of C. albicans obtained from the oral cavity of the four different groups. Culturing was done in yeast carbon base-bovine serum albumin. Speciation of Candida was performed by using a Candida identification kit, and CCC was measured by hemocytometer. Sap levels and CCC were higher in individuals with leukoplakia and OSCC than in nonsmokers or smokers (P = 0.001); however, there was no significant difference in Sap levels or CCC between smokers and nonsmokers (P = 0.529). Further, an intragroup correlation between CCC and Sap level was also observed. The higher level of Saps from C. albicans in individuals with leukoplakia and OSCC suggests that this pathogen plays a role in disease development and could aid in identifying the pathogenic commensal.

The activating receptors 2B4 and NTB-A, but not CRACC are subject to ligand-induced down-regulation on human natural killer cells

Activation of natural killer cells can be mediated by different receptors. Stimulation of the receptors 2B4, NTB-A and CRACC, members of the SLAM-related receptor family, induces cytotoxicity and cytokine production. The surface expression of 2B4 and other activating natural killer cell receptors is down-modulated after receptor engagement, which results in a weaker response to consecutive stimulation. We tested whether this regulatory mechanism applies to all SLAM-related receptors expressed by primary human natural killer cells. After co-culture with target cells expressing the respective ligands different effects on receptor surface expression were observed. While 2B4 ex-pression was strongly reduced, NTB-A showed less prominent down-modulation and the expression level of CRACC remained unchanged. The expression levels of the receptor-proximal signaling molecules SAP, EAT-2 and FynT did not change after receptor engagement. Co-culture with target cells expressing the ligands for NTB-A or CRACC had no impact on subsequent NTB-A or CRACC-mediated NK cell activation.

The activating receptors 2B4 and NTB-A, but not CRACC are subject to ligand-induced down-regulation on human natural killer cells

AbstractActivation of natural killer cells can be mediated by different receptors. Stimulation of the receptors 2B4, NTB-A and CRACC, members of the SLAM-related receptor family, induces cytotoxicity and cytokine production. The surface expression of 2B4 and other activating natural killer cell receptors is down-modulated after receptor engagement, which results in a weaker response to consecutive stimulation. We tested whether this regulatory mechanism applies to all SLAM-related receptors expressed by primary human natural killer cells. After co-culture with target cells expressing the respective ligands different effects on receptor surface expression were observed. While 2B4 ex-pression was strongly reduced, NTB-A showed less prominent down-modulation and the expression level of CRACC remained unchanged. The expression levels of the receptor-proximal signaling molecules SAP, EAT-2 and FynT did not change after receptor engagement. Co-culture with target cells expressing the ligands for NTB-A or CRACC had no impact on subsequent NTB-A or CRACC-mediated NK cell activation.

The proapoptotic function of SAP provides a clue to the clinical picture of X-linked lymphoproliferative disease

Deletion or mutation of the SAP gene is associated with the X-linked lymphoproliferative disease (XLP) that is characterized by extreme sensitivity to Epstein-Barr virus (EBV). Primary infection of the affected individuals leads to serious, sometimes fatal infectious mononucleosis (IM) and proneness to lymphoma. Our present results revealed a proapoptotic function of SAP by which it contributes to the maintenance of T-cell homeostasis and to the elimination of potentially dangerous DNA-damaged cells. Therefore, the loss of this function could be responsible for the uncontrolled T-cell proliferation in fatal IM and for the generation of lymphomas. We show now the role of SAP in apoptosis in T and B lymphocyte-derived lines. Among the clones of T-ALL line, the ones with higher SAP levels succumbed more promptly to activation induced cell death (AICD). Importantly, introduction of SAP expression into lymphoblastoid cell lines (LCL) established from XLP patients led to elevated apoptotic response to DNA damage. Similar results were obtained in the osteosarcoma line, Saos-2. We have shown that the anti-apoptotic protein VCP (valosin-containing protein) binds to SAP, suggesting that it could be instrumental in the enhanced apoptotic response modulated by SAP.

Endotoxin-like effects in acute phase response to Trypanosoma brucei brucei infection are not due to gastrointestinal leakage

Trypanosomosis is mainly an immunological and inflammatory response mediated by increased levels of pro-inflammatory cytokines. Evidence suggests that pathological changes produced during infection with trypanosomes could be initiated by nonspecific endotoxin-like substances in trypanosomes and/or Gram-negative secondary bacterial infection. Studies in trypanosome-infected rats indicate damage to the gastrointestinal tract (GIT) accompanied by increased leakage of the GIT mucosa. The current study was carried out to determine the in vivo response to endotoxin-like substances of Trypanosomabrucei brucei. To this purpose we neutralized the entrance of endotoxin through the GIT using polymyxin-B treatment and monitored the plasma concentration of the acute phase proteins SAP and Hp. The results in this study, where infection was performed in the presence of oral antibiotic that is not absorbed from GIT and which binds to and inactivates endotoxin, show that the elevated plasma levels of endotoxin-like activity and the resulting acute phase response indicated by an increase in levels of Hp and SAP, are due to trypanosome infection. Results obtained in the present study indicate that GIT is not the major source of elevated plasma endotoxin-like activity levels and the observed acute phase response was due to an increase in the levels of acute phase proteins SAP and haptoglobin.Therefore trypanosomes are responsible for the elevated plasma endotoxin-like activity levels and the subsequent systemic acute phase response in the host.

Motility and fertilizing capacity of boar semen stored in raffia palm (Raffia hookeri) sap extender at 15°C

Two separate experiments were conducted to test the ability of new semen extenders containing various levels of tropical raffia palm sap in sustaining the motility and fertilizing capacity of boar spermatozoa during storage at 15°C. Highly significant variations (P < 0.01) occurred in the ability of extenders to sustain progressive motility from days 1 to 4 of storage. One of the new extenders, raffia palm extender 2 (RPE–2) was superior to other trial extenders including the control (coconut milk extender, CME) in sustaining progressive motility of boar sperm from 24 - 72 h of storage; average motility scores at 24 and 72 h were 73.0 and 50.0% for RPE-2, 65.0 and 41.0% for CME (control), 55.0 and 36.0% for RPE-1 and 45.0 and 6.5% for RPE-0. Sperm tail vibratory movement was observed in sperm stored in RPE-1 and RPE-0 from day 2 of storage. Sperm in RPE-0 and RPE-1 lost motility completely at 96 h while motility of sperm in RPE-2 showed a sharp drop in motility values from 72 – 96 h relative to control. Average conception rates in 24 gilts inseminated per treatment with semen stored for 24, 48 and 72 h respectively were 83.3, 66.6 and 16.6% for RPE-2, 50, 16.6 and 0.0 percent for RPE-1, 16.6, 0.0 and 0.0% for RPE-0 and 75.0, 50.0 and 26.6% for CME (control). Average number of piglets born using semen stored for 24, 48 and 72 h respectively were 8.0, 8.0 and 6.5 for RPE-2 and 8.2, 8.2 and 6.0 for the control. RPE-2 supported better fetal survival rate than other trial extenders and control. Highly significant variations (P < 0.01) occurred among extenders in conception rate and number of piglets born. These results portray RPE-2 as a reliable short-term liquid semen storage medium for swine artificial insemination in the humid tropics.

The microinjection of AMPA receptor antagonist into the accumbens shell failed to change food intake, but reduced fear-motivated behaviour in free-feeding female rats

This study investigated the effect of the AMPA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 2.5 and 5.0 nmol/side) microinjected into the core and shell sub-regions of the accumbens (Acb) nucleus, on food intake and the level of anxiety in female rats. Bilateral microinjections of CNQX (5.0 nmol/side) into the Acb shell (AP, +1.08 to +2.04 ), but not into the Acb core, induced an anxiolytic-like effect in relation to rats microinjected with vehicle, since the animals exhibited low level of SAP in the feeding test. The anxiolytic-like effect induced by 5.0 nmol CNQX microinjection into the Acb shell may not be ascribed to changes in the motor activity of the animals, because the frequency of locomotion, rearing and grooming remained unchanged after the drug microinjection. However, neither Acb shell nor Acb core CNQX microinjections were able to change the animals food intake along 1 h feeding behaviour evaluation. Food intake remained unchanged 24 h after the drug microinjections either into the Acb shell or into the Acb core. The data suggest that AMPA receptor blockade in the Acb nucleus may differentially change the ingestive and defensive behaviours in female rats.

Bleeding sap and leaves of silver birch (Betula pendula) as bioindicators of metal contaminated soils

Bleeding sap of Betula pendula contained high levels of manganese on soils near ore outcrops and on acid soils, whereas high zinc levels were restricted to metal-mining and metal-smelting areas. The relationships between metal contents of bleeding sap and those of the soils were dependent on the kind of chemical soil extraction. Copper and zinc sap levels had a significantly positive correlation with the aqua regia soil extract, iron and manganese with the CaCl 2 -extract, and cadmium with the NH 4 -acetate soil extract. Metal contents of mature leaves were highest in autumn and all, except cadmium, were related in a similar metal-specific way to the same chemical soil extracts as the bleeding sap. Lead levels of sap and leaves had no consistent correlation with type of soil extract. The timing of collection and the advantages of the analysis of bleeding sap versus mature leaves are discussed.

Use of Diluted Tomato Petiole Sap for Potassium Measurement with the Cardy Electrode Meter

Tomato petiole sap from potassium (K) rate experiments conducted on prairie loam and river coarse sand soils from 1998 to 2002 was used to measure K concentration via the battery‐operated portable Cardy flat membrane electrode meter and inductive coupled plasma (ICP) laboratory instrumentation. In 1998, for undiluted sap levels less than 3000 mg kg−1 the linear Cardy–ICP correlation was r=0.94, but when K was more than 3000 mg kg−1, r was only 0.53. Diluting the sap with water (1∶1) improved the linear range to 6000 mg K L−1 (r=0.87, n=190). Petiole sap K, measured by either Cardy or ICP, was highly correlated to whole‐leaf K concentration all years, r=0.8 to 0.91. Even through the slope of the regression lines were similar for the years, the intercepts were significantly different (P<0.01). The Cardy electrode can be reliably used for tomato petiole K determination provided the sap is diluted and the usual handling precautions are taken to prevent petiole moisture loss and maintain stability in electrode readings.

Orthostatic Hypotension and Autonomic Pathways after Spinal Cord Injury

Individuals with spinal cord injury (SCI) are prone to orthostatic hypotension (OH). We aimed to develop a simple bedside test to evaluate autonomic control following chronic SCI, and to identify those most at risk of OH and cardiovascular dysfunction. We studied 14 subjects with cervical SCI, 11 with thoracic SCI, and 17 able-bodied controls. We continuously recorded heart rate (HR; ECG) and beat-to-beat systolic (SAP), diastolic (DAP) and mean (MAP) arterial pressures (Finometer) while supine, and following the passive assumption of an upright seated position. Stroke volume (SV), cardiac output (CO), and total peripheral resistance (TPR) were calculated. Plasma catecholamines were determined. Motor and sensory loss was assessed using the American Spinal Injury Association (ASIA) impairment scale. Autonomic pathways were assessed from sympathetic skin responses (SSR). Cervical SCI subjects had lower supine HR, SAP, and noradrenaline levels than thoracic SCI and controls (p < 0.05), and lower DAP and MAP than controls (p < 0.05). When upright, HR increased in all groups (p < 0.05); SAP, DAP, and MAP increased (p < 0.01) in thoracic SCI and controls, but not in cervical SCI. Cervical SCI had larger postural falls in SV (p < 0.05) and CO, with smaller increases in TPR than the other two groups. Upright catecholamine levels were lower in cervical SCI (p < 0.05) than thoracic SCI and controls. Completeness of SCI assessed by ASIA scale did not necessarily correlate with autonomic completeness assessed by SSR. Cardiovascular control during orthostasis was impaired and OH was common in cervical SCI, but not thoracic SCI. SSR may identify those at greatest risk of orthostatic hypotension and impaired cardiovascular control. We advocate that assessments of autonomic function be included in the neurological evaluation of SCI, in addition to the ASIA assessment.

Spinal Versus Epidural Anesthesia for Cesarean Delivery in Severe Preeclampsia: A Prospective Randomized, Multicenter Study

Epidural anesthesia currently is the preferred method for cesarean delivery in women with severe preeclampsia (defined in this study as a systolic arterial blood pressure [SAP] of 160 mm Hg or higher or a diastolic pressure [DAP] of at least 100 mm Hg in addition to proteinuria of 100 mg/dL or more). Although spinal anesthesia can be instituted more rapidly, has fewer complications, and is more cost-effective, there is concern that rapid and profound hypotension might further compromise the newborn infant. In this prospective, randomized, multicenter study, 47 women not in labor received epidural anesthesia with 18 to 23 mL of 2% lidocaine and 1:400,000 epinephrine followed by 3 mg of morphine after delivery. Another 53 nonlaboring women with severe preeclampsia received 2.2 mL of 0.5% hyperbaric bupivacaine plus 0.2 mg of morphine for spinal anesthesia. The 2 groups were demographically similar and comparable with regard to the volume of intravenous fluid given, treatment with magnesium sulfate or hydralazine, and estimated blood loss. Anesthesia lasted longer in the epidural group despite similar surgical duration times. Median sensory blocked levels were comparable in the epidural and spinal groups. The mean lowest levels of SAP, DAP, and mean arterial pressure (MAP) from induction to delivery were consistently lower in the spinal group. Mean differences were 14, 9, and 10 mm Hg, respectively. After delivery, the 2 groups had similar mean lowest arterial pressures. Significant hypotension, a SAP of 100 mm Hg or less, was approximately twice as frequent in the spinal group, but significant hypotension lasted no longer than 1 minute in both groups. Women given spinal anesthesia received more ephedrine, but hypotension was easily treated. In neither of the premature infants with an adverse outcome was the mother hypotensive. Birth weights, Apgar scores, and the need for admission to a neonatal intensive care unit all were similar in the spinal and epidural groups. These findings support the use of spinal anesthesia for cesarean delivery in women with severe preeclampsia.

Investigation of Monthly Variation in Some Plant-Nutrient Contents of Guttation Fluid Samples Taken from Dieffenbachia Plants

ABSTRACT Monthly variation in some of the plant-nutrient contents were investigated using the guttation fluid samples taken from dieffenbachia (Dieffenbachiae anoena) plants. Guttation-fluid samples were taken between September 1, 2001 and March 31, 2002. Amounts of potassium (K), calcium (Ca), magnesium (Mg), sodium (Na), iron (Fe), zinc (Zn), manganese (Mn), and copper (Cu) were analyzed by atomic absorption spectrophotometer. According to the results of the experiment, plant-nutrient composition of the fluids ranged in the following order during the experimental period: Ca (62.55%–82.91%) > Mg (11.05%–16.52%) > Na (2.70%–13.76%) > K (0.03%–20.53%) > Mn (0.26%–1.10%) > Zn (0.10%–0.31%) > Cu (0.05%–0.16%) > Fe (0.00%–0.55%). It was also observed that macro-cation contents of the samples decreased during winter and increased during the autumn and spring. Meanwhile, percentage microelement content of the fluid samples increased during the winter months, as macro-cations decreased. This result might have been due to the fact that plants had evidently developed a defense mechanism against cold damage during the winter season: by not emitting macro-cations out of their leaves, they were able to increase the pH level of xylem sap.

Use Diluted Tomato Petiole Sap for K Measurement with the Cardy Meter

Undiluted tomato petiole sap from a variety by K rate experiment (48 treatment rep combinations) was used to measure K concentration via the battery operated portable Cardy meter and ICP laboratory instrumentation. Three sample 1998 dates, 16 July., 21 Aug., and 8 Sept., resulted in K sap readings by ICP of 3917, 2612, and 2297 ppm, respectively. At sap levels below 3000 ppm the linear Cardy:ICP correlation was r = 0.04, but above 3000 ppm only 0.53. From 3500 to 5000 ppm K the Cardy meter under estimated actual sap K by 200 to 900 ppm. For the years 1999 and 2001, tomato petiole sap at each sample date (4) was diluted 1:1 with deionized water. The linear regression equation describing the relationship between ICP and Cardy meter measurements was: Cardy K ppm = 0.733 * ICP + 685 (r = 0.92, n = 190). The Cardy meter error over the 2000 to 5000 ppm K range was 8 to 12%. Petiole sap K, measured by either Cardy or ICP, was highly correlated to whole leaf K concentration both years. But even though the slope of the regression lines was similar the intercepts were significantly different (P≤.01). The significant 0.32% K difference in whole leaf between years precluded the development of a common regression line to predict whole leaf K from Cardy petiole sap determinations. The Cardy meter can be reliably used for tomato petiole K determination provided the sap is diluted and the usual handling precautions are taken to prevent petiole moisture loss.

N-stress alters aspartate and asparagine levels of xylem sap in soybean

The influence of N-stress on the transport of amino acids in the xylem sap was studied for nodulated and non-nodulated soybean plants. The transfer of non-nodulated plants cultivated with nitrate or of nodulated plants dependent exclusively on N 2 fixation to hydroponics with N-free nutrient solution (N-stress), led to a pronounced increase in aspartate and fall in asparagine (Asn) in the xylem sap. Such a change was also observed during the ‘N-hunger’ phase of the growth cycle of nodulated plants, characterized by the transient yellowing of leaves and coinciding with the initial stages of nodule development. In this case, aspartate increased to around 68% of the xylem amino acids while Asn, normally the most abundant amino acid, fell to 10%. The changes were reversible on recovery of plants from N-stress. In the case of nodulated plants, the fall in xylem Asn during N-stress and its subsequent rise during recovery closely followed Asn synthetase activity in the nodule, while aspartate produced an inverse relationship. Aspartate was prominent in the phloem, independent of the treatment. The possibility that the phenomenon is related to the metabolism of aspartate recycled through the root system is discussed.

Osteoclast differentiation from circulating mononuclear precursors in Paget’s disease is hypersensitive to 1,25-dihydroxyvitamin D 3 and RANKL

A characteristic feature of Paget’s disease is an increase in the number of osteoclasts in bone. Osteoclasts are formed from mononuclear phagocyte precursors that circulate in the monocyte fraction of peripheral blood. These cells require the presence of RANK ligand (RANKL)-expressing osteoblastic cells and human macrophage colony-stimulating factor (M-CSF) to form osteoclasts in vitro. To determine the role of osteoclast differentiation from circulating precursors in Paget’s disease, we cultured monocytes from Paget’s patients and gender- and age-matched normal controls with no evidence of bone disease for up to 21 days in the presence of UMR 106 cells and various concentrations of M-CSF (1–25 ng/mL) and 1,25-dihydroxyvitamin D 3 [1,25(OH) 2D 3] (10 −10 to 10 −7 mol/L). Relative to controls, there was a significant increase in the extent of osteoclast differentiation from pagetic monocytes as assessed by expression of tartrate-resistant acid phosphatase (TRAP), vitronectin receptor (VNR), and lacunar bone resorption. Serial dilution experiments (2 × 10 5 to 2 × 10 2 cells/well) showed no difference in the concentration of osteoclast precursors in the peripheral blood. In Paget’s patients with high serum alkaline phosphatase (sAP) levels, increased sensitivity to the osteoclastogenic effect of 1,25(OH) 2D 3 was noted. Osteoclast differentiation did not occur when M-CSF was substituted by interleukin-6 (IL-6) and soluble IL-6 receptor (sIL-6R), and these factors did not stimulate osteoclast differentiation in the presence of M-CSF. In this in vitro coculture system, osteoclast formation was inhibited by osteoprotegerin in a dose-dependent manner. In the presence of RANKL (5–30 ng/mL) and M-CSF (25 ng/mL), osteoclast formation and bone resorption were significantly increased in cultures of monocytes from patients with high and low sAP levels as compared with normal controls. Our findings suggest that the increase in osteoclast numbers seen in Paget’s disease results not from an increase in the number of circulating precursors in peripheral blood but rather from an increased sensitivity of osteoclast precursors to the humoral factors, 1,25(OH) 2D 3 and RANKL, which regulate osteoclast formation.

Modified atmosphere and modified humidity packaging alleviates chilling injury symptoms in mango fruit

Storage of mango (Mangifera indica L. cvs. Tommy Atkins and Keitt) fruits at 12°C caused slight chilling injury (CI) symptoms on the fruit peel, expressed as red spots around the lenticels (lenticel spotting). A modified atmosphere (5% CO2 and 10% O2) was created in 4-kg film-lined cartons by using microperforated polyethylene (PE) or Xtend ® film (XF). For ‘Keitt’ fruit, a similar atmosphere was also applied using controlled atmosphere chambers. After 3 weeks of storage at 12°C plus 1 week at 20°C, both modified and controlled atmosphere treatments were effective in reducing CI. The most effective reduction was found in fruits that were packed in the XF film. A second advantage of using XF film was the reduction in the level of sap inside the package due to the lower relative humidity in the XF film (90%) compared with that of PE packaging ( 99%). © 2000 Elsevier Science B.V. All rights reserved.