Collaborative meta-analysis of individual participant data from observational studies of Lp-PLA2 and cardiovascular diseases.

BACKGROUND: A large number of observational epidemiological studies have reported generally positive associations between circulating mass and activity levels of lipoprotein-associated phospholipase A2 (Lp-PLA2) and the risk of cardiovascular diseases. Few studies have been large enough to provide reliable estimates in different circumstances, such as in different subgroups (e.g., by age group, sex, or smoking status) or at different Lp-PLA2 levels. Moreover, most published studies have related disease risk only to baseline values of Lp-PLA2 markers (which can lead to substantial underestimation of any risk relationships because of within-person variability over time) and have used different approaches to adjustment for possible confounding factors. OBJECTIVES: By combination of data from individual participants from all relevant observational studies in a systematic 'meta-analysis', with correction for regression dilution (using available data on serial measurements of Lp-PLA2), the Lp-PLA2 Studies Collaboration will aim to characterize more precisely than has previously been possible the strength and shape of the age and sex-specific associations of plasma Lp-PLA2 with coronary heart disease (and, where data are sufficient, with other vascular diseases, such as ischaemic stroke). It will also help to determine to what extent such associations are independent of possible confounding factors and to explore potential sources of heterogeneity among studies, such as those related to assay methods and study design. It is anticipated that the present collaboration will serve as a framework to investigate related questions on Lp-PLA2 and cardiovascular outcomes. METHODS: A central database is being established containing data on circulating Lp-PLA2 values, sex and other potential confounding factors, age at baseline Lp-PLA2 measurement, age at event or at last follow-up, major vascular morbidity and cause-specific mortality. Information about any repeat measurements of Lp-PLA2 and potential confounding factors has been sought to allow adjustment for possible confounding and correction for regression dilution. The analyses will involve age-specific regression models. Synthesis of the available observational studies of Lp-PLA2 will yield information on a total of about 15 000 cardiovascular disease endpoints.

Pain and negative mood during rehabilitation after anterior cruciate ligament reconstruction: a daily process analysis.

Daily diary methods were used to examine changes in pain and negative mood over the first 6 weeks of rehabilitation after surgical reconstruction of the anterior cruciate ligament (ACL). Participants (58 men and 33 women) completed measures of personal factors (i.e., age, athletic identity, neuroticism, optimism) before surgery and indices of daily pain, negative mood, and stress for 42 days after surgery. Multilevel modeling revealed that, as would be expected, daily pain ratings decreased significantly over the course of the study and that the rate of decline in pain ratings decreased over time. Age and daily negative mood were positively associated with daily pain ratings. Daily negative mood also decreased significantly over the course of the study and was positively associated with neuroticism, daily pain, and daily stress. Athletic identity and optimism interacted with time since surgery in predicting daily negative mood such that participants with high levels of athletic identity and low levels of optimism reported greater decreases in daily negative mood over time. Overall, the findings reveal a pattern of improved psychological functioning over the early stages of post-operative ACL rehabilitation.

Insulin-treated Zucker diabetic fatty rats retain the hypertriglyceridemia associated with obesity.

Lipoprotein and apolipoprotein changes were evaluated in 10-week-old Zucker diabetic fatty (ZDF) male rats following 12 weeks of insulin treatment, which normalized blood glucose and maintained weight gaining characteristic of nondiabetic Zucker fatty rats. Compared with untreated ZDF rats (saline-injected), insulin treatment resulted in increased very-low-density lipoprotein (VLDL; d < 1.006 g/mL) and decreased alpha lipoprotein on agarose gel electrophoresis. These findings were consistent with an observed increase in VLDL triglyceride and cholesterol, and decreased high-density lipoprotein (HDL) cholesterol with insulin treatment in isolated lipoproteins. B100 levels were unchanged by insulin treatment, but B48 levels were significantly increased in the VLDL fraction. Insulin treatment depressed apolipoprotein (apo) A-I levels in HDL, but had little effect on total apo E, apo A-IV, or apo C, although apo C was redistributed to the VLDL fraction. These results suggest that insulin treatment of ZDF rats normalizes hyperglycemia and prevents age-related changes in lipoprotein parameters associated with development of insulinopenic diabetes. Insulin therapy in ZDF rats thereby sustains the hyperlipidemic lipoprotein pattern associated with hyperinsulinemia and obesity.

Glucose-stimulated insulin secretion suppresses hepatic triglyceride-rich lipoprotein and apoB production.

The current study assessed in vivo the effect of insulin on triglyceride-rich lipoprotein (TRL) production by rat liver. Hepatic triglyceride and apolipoprotein B (apoB) production were measured in anesthetized, fasted rats injected intravenously with Triton WR-1339 (400 mg/kg). After intravascular catabolism was blocked by detergent treatment, glucose (500 mg/kg) was injected to elicit insulin secretion, and serum triglyceride and apoB accumulation were monitored over the next 3 h. In glucose-injected rats, triglyceride secretion averaged 22.5 +/- 2.1 microg.ml(-1).min(-1), which was significantly less by 30% than that observed in saline-injected rats, which averaged 32.1 +/- 1.4 microg.ml(-1).min(-1). ApoB secretion was also significantly reduced by 66% in glucose-injected rats. ApoB immunoblotting indicated that both B100 and B48 production were significantly reduced after glucose injection. Results support the conclusion that insulin acts in vivo to suppress hepatic very low density lipoprotein (VLDL) triglyceride and apoB secretion and strengthen the concept of a regulatory role for insulin in VLDL metabolism postprandially.

Effect of dietary fat on the development of non-insulin dependent diabetes mellitus in obese Zucker diabetic fatty male and female rats.

The obese Zucker diabetic fatty male rat (ZDF/Gmi¿trade mark omitted¿-fa) has become a widely used animal model of NIDDM, in contrast to the obese ZDF females that rarely develop NIDDM. However, preliminary observations suggest that obese ZDF females may become diabetic on high-fat diets. Therefore, we studied the effect of dietary fat on development of NIDDM, dyslipidemia, and alterations in organ-specific serum panels in obese ZDF males and females. Results indicated different effects of dietary fat-content on development of diabetes in males and females. Males, even on low fat-content diets, developed diabetes but the process was accelerated as a function of dietary fat-content, whereas only the highest fat-content diet induced development of NIDDM in obese ZDF females. Additionally, triglyceride/apolipoprotein B ratios demonstrated gender-specific differences in the nature of circulating lipoprotein particles independent of diabetic state with values for females approximately twice those of males indicating more highly triglyceride-enriched lipoprotein particles in females. We conclude that the obese ZDF female rat has the potential to become an important animal model of NIDDM especially in women where few models currently exist.

Characterization of paprika (Capsicum annuum) extract in orange juices by liquid chromatography of carotenoid profiles.

The carotenoid pigment profiles of authentic pure orange juices from Spain and Florida and an industrial paprika (Capsicum annuum) extract used for food coloring were obtained using reversed-phase liquid chromatography with a C18 packed column and an acetone/methanol/water eluent system. The procedure involving the carotenoid extraction is described. Both retention times and spectral properties using photodiode array detection for characterization of the major carotenoids at 430 and 519 nm are given. The influence of external addition of tangerine juice and/or paprika extract on orange juice color is described using the U.S. Department of Agriculture scale and adulterated orange juice. The procedure for quantitation of externally added paprika extract to orange juice is investigated, and the limit of quantitation, coefficient of variation, and recoveries are determined.

Differentiation of several geographical origins in single-strength Valencia orange juices using quantitative comparison of carotenoid profiles.

Pure Valencia orange (Citrus sinensis) juices (64 samples) from Spain, Israel, Belize, Cuba, and Florida, harvested during two seasons (1996-1997 and 1997-1998), were analyzed for their carotenoid profiles. The detection of saponified carotenoid pigments has been achieved and quantitated using a photodiode array detection monitored at 350, 430, and 486 nm. Carotenoid pigments commonly found in the orange variety Valencia have been separated on a polymeric C-30 column using a ternary gradient as eluent. Pure Valencia juices from oranges grown in Mediterranean regions (Israel and Spain) have a high carotenoid content, expressed in beta-carotene (5-18 and 14-35 mg L(-)(1), respectively), compared to those grown in tropical and subtropical regions (Cuba, Belize, and Florida) (4-10, 2-8, and 5-10 mg L(-)(1), respectively). Quantitative results allowed the differentiation of Valencia variety geographical origins, in particular, the Mediterranean area from tropical and subtropical areas, using multidimensional analyses of carotenoid contents.

Differentiation of several geographical origins in single strength Valencia orange juices using quantitative comparison of carotenoid profiles

Pure Valencia orange (Citrus sinensis) juices (64 samples) from Spain, Israel, Belize, Cuba, and Florida, harvested during two seasons (1996-1997 and 1997-1998), were analyzed for their carotenoid profiles. The detection of saponified carotenoid pigments has been achieved and quantified using a photodiode array detection monitored at 350, 430, and 486 nm. Carotenoid pigments commonly found in the orange variety Valencia have been separated on a polymeric C-30 column using a ternary gradient as eluent. Pure Valencia juices from oranges grown in the Mediterranean regions (Israel and Spain) have a high carotenoid content, expressed in beta-carotene (5-18 and 14-35 mg L(-)(1), respectively), compared to those grown in tropical and subtropical regions (Cuba, Belize, and Florida) (4-10, 2-8, and 5-10 mg L(-)(1), respectively). Quantitative results allowed the differentiation of Valencia variety geographical origins, in particular, the Mediterranean area from tropical and subtropical areas, using multidimensinal analyses of carotenoid contents. (AU)

Lipoprotein alterations in 10- and 20-week-old Zucker diabetic fatty rats: hyperinsulinemic versus insulinopenic hyperglycemia.

Lipoprotein and apolipoprotein parameters were studied in the male Zucker diabetic fatty (ZDF) rat at 10 and 20 weeks of age, corresponding to hyperinsulinemic and insulinopenic type 2 diabetes mellitus, respectively. At both ages, ZDF rats had elevated serum triglycerides, free fatty acids, and corticosterone, whereas 20-week ZDF rats had reduced thyroid hormones. At 10 weeks, the hyperlipidemia was confined to elevations in pre-beta triglyceride-rich (d < 1.006 g/mL) lipoproteins. By 20 weeks, all lipoprotein density fractions were increased compared with lean rats, with substantial increases in both low-density lipoprotein (LDL) and high-density lipoprotein (HDL) cholesterol. In ZDF rats, there was a progressive increase in apolipoprotein B (apo B) from 1.9 times control at 10 weeks to three times control at 20 weeks. The increase in apo B was accompanied by a shift of apo B, particularly B100, from very-low-density lipoprotein (VLDL) into denser lipoproteins corresponding to intermediate-density lipoproteins plus LDLs (1.006 < d < 1.063 g/mL). In Zucker and 10-week ZDF rats, in the presence of hyperinsulinemia, the increase in serum apo B was predominantly apo B48 present in VLDL. By 20 weeks, when ZDF rats are insulinopenic, the mass ratio of B48:B100 shifted from 2.7 to 0.7. The shift was associated with a decrease in hepatic-edited apo B mRNA. Apo E increased in lean rats between 10 and 20 weeks of age. Although apo E also increased in ZDF rats, the increase by 20 weeks was less than that of lean rats. The molar ratio of apo E to B in VLDL was decreased in ZDF rats. In lean rats, greater than 50% of apo E was present in HDL, in contrast to ZDF rats, where less than 20% of apo E was present in HDL. VLDL apo E shifted to denser fractions by 20 weeks of age, similar to apo B. The apo C level was more than double compared with the level in lean rats and was redistributed from the HDL fraction to lipoprotein fractions containing apo B. Both apo A-I and apo A-IV levels more than doubled between 10 and 20 weeks in ZDF rats. The ZDF rat model may be useful in comparative studies of lipoproteins during diabetic progression from hyperinsulinemia to insulinopenia.

Biochemical pathway mediating the response of bone cells to capacitive coupling.

Rat calvarial bone cells or mouse MC3T3-E1 bone cells subjected to a capacitively coupled electric field of 20 mV/cm consistently showed significant increases in cellular proliferation as determined by deoxyribonucleic acid content. Verapamil, a membrane calcium channel blocker; W-7, a calmodulin antagonist; indocin, a prostaglandin synthesis inhibitor; or bromophenacyl bromide, a phospholipase A2 inhibitor, each at a concentration that did not interfere with cell proliferation in control cultures, inhibited proliferation in those cultures subjected to the electric field. In contrast, neomycin, an inhibitor of the inositol phosphate cascade, did not inhibit this electrically induced cellular proliferation. Prostaglandin E2 production also was increased significantly with electrical stimulation, and this increase was inhibited by verapamil or indocin but not by neomycin. Thus, the data suggest that the signal transduction mediating the proliferative response of cultured bone cells to a capacitively coupled field involved transmembrane calcium translocation via voltage gated calcium channels, activation of phospholipase A2, and a subsequent increase in prostaglandin E2. Increases in cytosolic calcium and activated calmodulin are implied. The inositol phosphate pathway, unlike its dominant role in signal transduction in mechanically stimulated bone cells, does not appear to play a role in signal transduction in the proliferative response of bone cells to electrical stimulation.

The search for the Holy Grail: a century of anterior cruciate ligament reconstruction.

A perspective on the history of the development of anterior cruciate ligament reconstruction is presented. The lack of critical analyses establishing the relative effectiveness of many previously described procedures is documented.

The biochemical pathway mediating the proliferative response of bone cells to a mechanical stimulus.

Calvarial bone cells of rats were subjected to either a cyclic biaxial strain of 0.17 per cent (1700 microstrain) or a hydrostatic pressure of 2.5, five, or ten pounds per square inch (17.2, 34.5, or sixty-nine kilopascals). The frequency was held constant at one hertz for both types of mechanical stimulation. When cultured bone cells that had been subjected to a cyclic biaxial strain for two hours were harvested twenty-two hours later, it was found that the level of prostaglandin E2 had increased significantly (p < 0.01) as had cellular proliferation (p < 0.01), as indicated by the incorporation of [3H]-thymidine. The addition to the medium of indomethacin, an inhibitor of prostaglandin synthesis, at a ten-micromolar concentration significantly inhibited (p < 0.01) the increase in prostaglandin E2 synthesis but had no effect on the strain-induced increase in cellular proliferation, as indicated by the incorporation of [3H]-thymidine. Twenty-four hours after exposure to the same cyclic biaxial strain for thirty seconds, other cultured bone cells showed a significant increase in the level of cytoskeletal calmodulin (p < 0.05) and in the DNA content (p < 0.05). N-(6-aminohexyl)-5-chloro-1-naphthalene-sulfonamide (W-7), a calmodulin antagonist, was added to the medium at a one-micromolar concentration, which had been shown to have no effect on the increase in the DNA content of control cells; W-7 completely blocked the increase in the level of cytoskeletal calmodulin and in the DNA content in the cells that were subjected to a cyclic biaxial strain. The bone cells subjected to a hydrostatic pressure showed a dose-dependent increase in the concentration of cytosolic Ca2+, as measured with Fura 2-AM, a fluorescent indicator of intracellular calcium. With a pressure of ten pounds per square inch (sixty-nine kilopascals), the increase in the concentration of cytosolic Ca2+ was nearly eight times greater than that at 2.5 pounds per square inch (17.2 kilopascals) (126 +/- 15.2 compared with 16 +/- 8.0 nanomolar, mean and standard deviation). The addition to the medium of neomycin, an inhibitor of the inositol phosphate cascade, at a ten-millimolar concentration completely blocked the increase in the concentration of cytosolic Ca2+ in these cells; this concentration of neomycin had been shown to have no effect on proliferation in control bone cells. There was also a dose-dependent relationship between the duration of the stimulus and the cellular proliferation. Remarkably, one cycle of pressure at ten pounds per square inch (sixty-nine kilopascals) and a frequency of approximately one hertz produced a 57 per cent increase in the incorporation of [3H]-thymidine at twenty-four hours (p < 0.001). From these findings, we hypothesized that the inositol phosphate cascade-cytosolic Ca(2+)-cytoskeletal calmodulin system plays a dominant role in the signal transduction of a mechanical stimulus into increased proliferation of bone cells, at least under the conditions reported here.

Failure of anterior cruciate ligament reconstruction: the biologic basis.

The replacement tissue used for anterior cruciate ligament reconstruction undergoes extensive biologic remodeling and incorporation after implantation. Successful biologic incorporation of the graft is dependent on a number of factors including graft placement, tensioning, and the nature of the tissue (allograft versus autograft). Failure of an anterior cruciate ligament reconstruction may occur on the basis of either technical, mechanical, or biological factors. Biologic factors include cellular repopulation, matrix remodeling, the ultimate small diameter collagen fibril orientation, the final cross sectional area of the graft, a favorable vascularization, and not overloading the graft during the remodeling process. The fully incorporated graft never duplicates the native anterior cruciate ligament but works as a check reign that makes the knee more functional.

Biologic incorporation of allograft anterior cruciate ligament replacements.

Soft tissue allografts allow the orthopaedic surgeon to reconstruct ligaments without having to harvest additional tissue from the patient, which can eliminate donor tissue site morbidity and reduce surgical time. There is still much to be learned about the biologic aspects of the remodeling and incorporation of allografts in comparison with autografts. The interaction of cells, matrix, and biomolecules, such as growth factors, plays an important role that can potentially modulate, enhance, or impede the healing response in allografts. The authors have shown that, in the short term, allografts used in anterior cruciate ligament reconstruction are not as rapidly remodeled and incorporated into host tissue as are autografts. The long-term implications of this slower allograft incorporation in anterior cruciate ligament reconstruction are still unknown. The cells that repopulate allografts and autografts favor production of smaller diameter collagen fibrils, which in sufficient numbers can provide significant strength. Use of allografts raises other issues and potential disadvantages, including scarcity, immunogenicity, the potential for disease transmission, and cost-effectiveness in anterior cruciate ligament reconstruction.

Liver regrowth and apolipoprotein B secretion by rat hepatocytes following partial hepatectomy.

Apolipoprotein (apo) B is an essential component for the assembly and secretion of lipoproteins. The current report examines apo B production using primary cultures of hepatocytes derived from rats 3 to 21 days after partial hepatectomy (PH) to determine the effects of liver regrowth on apo B. Studies indicate that hepatocytes stimulated by PH have a two-thirds reduction in net apo B production 3 to 7 days after surgery, which coincides with the period of maximum rate of liver regrowth. Both higher (apo BH)- and lower-molecular-weight (apo BL) apo B are synthesized and secreted after PH, indicating the presence of edited apo B mRNA in hepatocytes. Hepatocytes derived from PH rats are more sensitive to insulin inhibition of apo B secretion compared with controls, suggesting an enhanced effect of insulin on newly replicated hepatocytes. Epidermal growth factor (EGF), a key regulator of liver regrowth following PH, potentiates the inhibitory action of insulin on apo B secretion in control hepatocytes and those derived from rats 2 to 3 weeks after PH. However, the potentiating effect of EGF on insulin inhibition of apo B is not discernible in hepatocytes 3 to 7 days after PH. The short-term in vitro hormonal effects occurring even with decreased apo B production suggest that this pathway remains available following PH to balance lipoprotein secretion with lipid and energy requirements necessary for liver regeneration.

Combined serum amylase and lipase determinations for diagnosis of suspected acute pancreatitis.

Serum amylase and lipase measurements are often used to diagnose acute pancreatitis. This study addresses the question of whether it is advantageous to order serum amylase and lipase tests simultaneously. We evaluated performance of the two tests separately and in combination through a retrospective study of patients for whom both amylase and lipase determinations were ordered. Initial analysis of test performance was conducted with a uniformly applied criterion based on determination of optimal sensitivity-specificity pairs. Individual tests and combinations of tests, including the "AND" and "OR" rules and discriminant functions, were examined. Only the discriminant approach demonstrated better performance than the lipase test alone. This finding was subsequently confirmed by logistic regression analysis. We conclude that ordering both tests simultaneously can be advantageous in diagnosing acute pancreatitis when a bivariate approach is used; however, this must be weighed against the difficulties associated with clinical implementation of such approaches.

Cellular heterogeneity in binding and uptake of low-density lipoprotein in primary rat hepatocytes.

Hepatocellular heterogeneity of biochemical function is well established for many aspects of liver metabolism. This study addresses the question of cellular heterogeneity in the catabolism of low-density lipoprotein by rat hepatocytes. Low-density lipoprotein binding (4 degrees C) and uptake (37 degrees C) by rat hepatocytes were studied by use of human low-density lipoprotein labeled with a highly fluorescent lipophilic probe, N,N-dipentadecylaminostyrylpyridinium iodide, recently developed by us. Single-cell suspensions derived from rat hepatocytes in primary culture and from liver perfusion were studied with flow cytometry with and an approximation algorithm for data analysis. These studies show subpopulations of cells negative and positive for the specific binding and uptake of low-density lipoprotein. Dissociation constants for low-density lipoprotein binding and uptake were determined for the total population (18 micrograms/ml, binding; 12 micrograms/ml, uptake) and found to be in good agreement with previously reported values. Additionally, the dissociation constant for binding for the positive subpopulation was determined and found to be 3 micrograms/ml. This lower value is more typical of the values seen in other cell types. These findings are strongly suggestive of functional heterogeneity in the hepatic catabolism of low-density lipoprotein.

Arthroscopically assisted proximal quadricepsplasty for patellar instability.

Arthroscopy has come to play a major role in the evaluation and treatment of patellofemoral instability. In fact, most patients sustaining a traumatic dislocation or recurrent subluxations are subjected to, at a minimum, an arthroscopic evaluation to assess intraarticular damage and patellar tracking.

Immunolocalization, quantitation and cellular heterogeneity of apolipoprotein B in rat hepatocytes.

Hepatocyte autofluorescence represents a major problem in immunofluorescence studies with fluorescein conjugates because of significant spectral overlap. We describe a method for immunostaining hepatocytes with R-phycoerythrin (a fluorochrome with minimal overlap with autofluorescence) with paraformaldehyde fixation and Triton X-100 permeabilization for better antibody penetration. This method produced both perinuclear (presumed Golgi apparatus) and dispersed, reticular staining (presumed endoplasmic reticulum) in rat hepatocytes in culture stained with a monoclonal antibody to rat apolipoprotein B. Treatment with brefeldin A resulted in loss of apolipoprotein B perinuclear staining and increased reticular immunofluorescence consistent with known properties of brefeldin A (inhibition of protein transport within the secretory pathway by dissolution of Golgi bodies). This suggests that apolipoprotein B epitopes are present in both Golgi bodies and endoplasmic reticulum. To demonstrate the utility of the technique for quantitative studies, static cell cytofluorometry of brefeldin A-treated cells was performed, demonstrating increases in specific immunofluorescence of apolipoprotein B corresponding closely to results estimated by monoclonal antibody radioimmunoassays of cellular homogenates. The technique was then used with flow cytometry of single-cell suspensions of control rat hepatocytes derived from immunostained primary cultures to reveal cell-to-cell heterogeneity of apolipoprotein B epitope expression manifested as apolipoprotein B-negative and positive populations. Results for brefeldin A-treated cells revealed even clearer delineation of heterogeneity as indicated by frank bimodality of the populations, along with not only higher mean apolipoprotein B levels but also a significantly higher proportion of apolipoprotein B-positive cells than in the control.