Impaired nonspecific cellular immunity in experimental cholestasis.

The abilities of polymorphonuclear leukocytes (PMN) and pulmonary alveolar macrophages (PAM), to demonstrate chemotaxis, phagocytosis, and superoxide release after bile duct ligation in the rat were investigated to determine the effect of cholestasis on nonspecific cellular immune mechanisms. Chemotactic response to C5a and FMLP, phagocytosis of 14C labeled Staphylococcus aureus, and zymosan-induced superoxide release were evaluated 21 days after bile duct ligation (BDL), sham operation, or in normal controls. Serum total bilirubin level was elevated after BDL (p less than 0.01). Chemotactic ability was similar to each group. PMN phagocytic uptake of 14C labeled Staphylococcus aureus was depressed in BDL (p less than 0.05). BDL rats exhibited impaired PAM phagocytic indices and improved PMN superoxide release (p less than 0.03). PAM superoxide release was similar in each study group. Alterations in phagocytic function with cholestasis are important deficits in nonspecific cellular immunity that may contribute to the high incidence of infective complications associated with obstructive jaundice.     

Structural and functional alterations in the zona fasciculata of the rat adrenal cortex in obstructive jaundice

This study investigated the effect of experimentally-induced cholestasis in the rat on the structure and function of the zona fasciculata, the glucocorticoid secretory region of the adrenal cortex. Wistar-Furth rats (200-250g) were assigned to three groups: bile duct ligated (BDL), Sham operated (Sham) and unmodified normal control (NC). On day 14, serum bilirubin and liver histology were performed to confirm cholestasis in BDL animals together with basal 24 hour 17-hydroxycorti-costeroid excretion, adrenal histology and zona fasciculata ultrastructure in all experimental groups. Following this laparotomy, structural and functional studies were repeated on day 15 to evaluate the response of the gland to surgically induced stress. Basal 24 hr, 17-hydroxycorticoid steroid excretion was elevated in BDL animals (26.9 +/- 3.2 micrograms/24 hr) with respect to Sham (10.4 +/- 2.3) and NC groups (13.5 +/- 3.2) (p less than 0.05). Adrenal histology and ultrastructural studies demonstrated excessive accumulation of vesicles laden with glucocorticoid biogenic precursors. Following surgical stress 24 hr 17 OH corticosteroid excretion increased in all groups: BDL (31.0 +/- 3.0 micrograms/24 hr) vs Sham (15.6 +/- 1.8) and NC (14.5 +/- 2.4) Moderate alterations in zona fasciculata architecture were seen following surgery in all groups. Cholestasis induces overactivity of the zona fasciculata of the adrenal cortex, and may modify the normal metabolic responses to surgical and other stresses.     

Role of neutrophil derived oxidants and elastase in lipopolysaccharide-mediated renal injury

Gram-negative bacterial sepsis is frequently associated with acute renal failure but the specific effects of lipopolysaccharide (LPS) and other bacterial products on kidney function are not known. Since either LPS or formyl-methionyl-leucyl-phenylalanine (FMLP)--a chemotactic peptide from bacterial cell walls--activate neutrophils (PMN) to release a number of potentially toxic factors in vitro, we determined the effect of adding PMN with LPS and/or FMLP to isolated perfused rat kidneys. Isolated rat kidneys perfused with LPS alone or LPS and normal PMN had normal glomerular filtration rates (GFR) and tubular Na reabsorption (TNa). Kidneys perfused with FMLP alone or FMLP and normal PMN also had normal GFR and TNa. In contrast, addition of PMN with both FMLP and LPS caused progressive renal dysfunction. For example, after 60 minutes of perfusion, GFR was reduced from 610 +/- 31 to 147 +/- 17 microliters/min/g and TNa from 97 +/- 1 to 72 +/- 2%, both P less than 0.01. Perfusion with the O2 metabolite scavengers catalase or dimethylthiourea afforded no protection while perfusion with the neutrophil elastase inhibitor Eglin C conferred substantial, but not complete, protection: GFR 492 +/- 34 microliters/min/g; TNa 91 +/- 3%. However, perfusion with both Eglin C and catalase completely prevented the toxic effects of LPS and FMLP-treated PMN on renal function. We conclude that in isolated kidneys, 1) the toxic effects of LPS requires FMLP-treated PMN and that 2) LPS and FMLP treated PMN cause progressive renal injury which is mediated by both O2 metabolites and neutrophil elastase.     

Abnormal cytoplasmic pH regulation during activation in uremic neutrophils.

Neutrophils isolated from ESRF patients demonstrated abnormal cytoplasmic pH changes after FMLP stimulation; the initial cytoplasmic acidification was absent (P less than 0.001 compared to controls) and the degree of alkalinization enhanced (P less than 0.05 compared to controls). This effect was not due to the absence of any of the factors associated with acidification in normal PMN since superoxide production was enhanced (P less than 0.05 compared to controls) and intracellular calcium release was normal. Our observations are not explicable by alterations in the function of the Na:H antiport since the kinetics of antiport activation by cytoplasmic pH were not different in uremic and control cells. Other factors must therefore be important in generating the abnormal pH response to chemotactic factors in uremic PMN. Cells from CAPD patients had some degree of initial acidification (P less than 0.001 compared to controls and P less than 0.05 compared to ESRF) and enhanced alkalinization (P less than 0.05 compared to controls). Preincubation of normal PMN in four-hour dwell PDE reproduced the responses of uremic PMN with absent acidification, enhanced alkalinization and enhanced superoxide generation after FMLP stimulation (P less than 0.05 compared to controls). Changes in the control of cytoplasmic pH in stimulated PMN may influence PMN function, and our observations may be relevant to the susceptibility of uremic patients to infection.     

TNF-alpha--accelerated apoptosis abrogates ANCA-mediated neutrophil respiratory burst by a caspase-dependent mechanism

BACKGROUND: Tumor necrosis factor (TNF)-alpha rapidly primes neutrophils (PMN) for an anti-neutrophil cytoplasmic antibody (ANCA)-induced respiratory burst and is thus proinflammatory. TNF-alpha also progressively accelerates apoptosis. We investigated the effect of TNF-alpha-mediated apoptosis on ANCA antigen expression and on ANCA-induced superoxide generation in human PMN. METHODS: PMN were brought to apoptosis by 10 ng/mL of TNF-alpha or a combination of TNF-alpha and 2.5 microg/mL cycloheximide, a protein synthesis inhibitor, or cycloheximide alone for three hours. Apoptosis and ANCA antigen expression were assessed by fluorescence-activated cell sorting (FACS) and microscopy. Superoxide was determined with the ferricytochrome C assay. RESULTS: TNF-alpha with cycloheximide for three hours caused apoptosis in 87% PMN compared to 2% in untreated controls (N=18; P < 0.01). Accelerated apoptosis was associated with an increase in ANCA-antigen expression for both proteinase 3 and myeloperoxidase (P < 0.05). Nevertheless, apoptosis was paralleled by a decreased proteinase 3 and myeloperoxidase ANCA-induced respiratory burst (P < 0.05). Furthermore, superoxide release in response to immune complexes, phorbol ester (PMA), and bacterial peptide (FMLP) was significantly decreased. Blocking caspase-3 activity prevented apoptosis in TNF-alpha with cycloheximide-treated cells (83% to 2%) and prevented compromised respiratory burst in response to ANCA. Caspase-3 inhibition abrogated apoptosis-mediated ANCA antigen up-regulation (PR3 141.6 +/- 34.1 MFI to 33.9 +/- 7.8; MPO 48.3 +/- 12.9 MFI to 11.9 +/- 3.2, N=6, P < 0.05). CONCLUSIONS: TNF-alpha-accelerated apoptosis was associated with increased ANCA antigen expression but with down-regulated respiratory burst activity in response to ANCA. Specific inhibition of apoptosis by caspase-3 blockade prevented the increase in ANCA-antigen expression and preserved the capability of generating superoxide, thereby establishing a causative role for apoptosis. We suggest that TNF-alpha exhibits dual actions by both priming and terminating ANCA-mediated activation of human PMN.     

Surface receptor mobilization in complement-mediated neutrophil activation: characterization and effects of methylprednisolone

Complement-mediated neutrophil activation (CMNA) is an important host defense mechanism that is essential for effective neutrophil (PMN) proinflammatory activity. It has also been implicated as a pathogenic mechanism contributing toward the development of adult respiratory distress syndrome. Utilizing zymosan-activated serum pretreatment as an in vitro model for CMNA, we characterized the effects of CMNA on PMN superoxide (SO) production and N-formyl-methionyl-leucyl-phenylalanine (FMLP) receptor status. CMNA was associated with a 132 +/- 38% increase in FMLP-induced SO generation and a 110 +/- 30% increase in FMLP receptor expression. Methylprednisolone pretreatment prevented both the FMLP receptor mobilization and the SO priming effects of CMNA. These data support a concept that FMLP receptor mobilization is an important element in the PMN activation process. In addition, blocking this phenomenon may have clinical significance in attempts to modulate the potential damaging effects of the increased PMN oxidative metabolism associated with CMNA.     

Echocardiographic characterization of left ventricular diastolic properties in patients presenting for the maze procedure.

The aim of this study is to characterize and compare the left ventricular (LV) diastolic filling patterns in patients with paroxysmal (PAF) versus chronic atrial fibrillation (CAF) undergoing the maze procedure and to examine their relation with the hemodynamic status. Fifty patients with PAF and 22 with CAF were studied. Hemodynamic measurements and transesophageal echocardiography (TEE) were performed after the induction of anesthesia but before surgical incision, at stable conditions. Transmitral (TMF) and pulmonary venous flow (PVF) velocities were recorded with the pulsed Doppler method. Statistical analysis between the two groups (PAF and CAF) was performed using Student's t-test and chi-squared test, with P less than .05 statistically significant. Compared with patients in the PAF group, those in the CAF group had: (1) higher pulmonary capillary wedge pressure (14 +/- 5 v 12 +/- 4 mm Hg; P < .05), (2) lower left ventricular fraction of area change (43% +/- 6% v 52% +/- 9%; P < .01), (3) slower PVF systolic wave velocity (23 +/- 10 v 35 +/- 15 cm/s; P < .05), and (4) lower ratio of PVF systolic to diastolic wave velocity (0.75 +/- 0.3 v 1.2 +/- 0.4; P < .05). In the present study, LV filling patterns of abnormal relaxation were found in all our patients who underwent the maze procedure for CAF or PAF. Although the cause of LV filling abnormalities is not apparent, the data suggest LV diastolic dysfunction is prevalent in these patients.     

Neutrophil priming and apoptosis in anti-neutrophil cytoplasmic autoantibody-associated vasculitis

BACKGROUND: Interactions between anti-neutrophil cytoplasmic autoantibody (ANCA) and primed neutrophils (PMNs) may be central to the pathogenesis of primary small vessel vasculitis. PMNs from patients are primed, expressing proteinase 3 (PR3) on the cell surface, which permits interaction with ANCA. In vitro ANCA activates primed PMN to degranulate and generate a respiratory burst. Resultant reactive oxygen species are important in triggering apoptosis, but the fate of PMN in ANCA-associated vasculitis is unknown. Failure to remove apoptotic PMN in a nonphlogistic manner may sustain the inflammatory response. METHODS: PMNs from patients or controls were isolated, and the basal production of superoxide was measured by the superoxide dismutase-inhibitable reduction of ferricytochrome C. ANCA antigen expression on apoptotic PMN was assessed at 0, 12, and 18 hours by flow cytometry using dual staining with FITC-conjugated annexin V and PE-conjugated anti-murine IgG against monoclonal ANCA. Apoptosis was also assessed by morphology. In further studies, apoptotic PMNs were opsonized with monoclonal anti-myeloperoxidase (MPO) or anti-proteinase-3 (PR3) or irrelevant isotype-matched IgG (N IgG) and phagocytosis by macrophages was measured using interaction assays. Cytokines interleukin-8 (IL-8) and interleukin-1 were measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: Proteinase-3 expression (active 63.04 +/- 5.6% of total number of cells, remission 51.47 +/- 7.9% of total number of cells, control 17.7 +/- 4.7% of total number of cells, P < 0.05) and basal superoxide production (active 6.9 +/- 0.8 nmol/L x 10(6) cells, remission 5.15 +/- 0.4 nmol/L/10(6) cells, control 3.63 +/- 0.3 nmol/L/10(6) cells, P < 0.001) were significantly greater with freshly isolated PMN from patients than controls. PR3 expression and superoxide generation were positively correlated. PMN from patients with active disease became apoptotic at a greater rate than those of controls (at 18 hours, patients 72.3 +/- 3.9% apoptosis, controls 53.2 +/- 2.7% apoptosis, P < 0.05). PR3 and MPO expression were significantly greater on PMN isolated from patients at 12 and 18 hours. Opsonization of apoptotic PMN with ANCA significantly enhanced recognition and phagocytosis by scavenger macrophages (anti-MPO 88.95 +/- 6.27, anti-PR3 93.98 +/- 4.90, N IgG 44.89 +/- 3.44, P < 0.01) with increased secretion of IL-1 (anti-PR3 34.73 +/- 6.8 pg/mL, anti-MPO 42.01 +/- 12.3 pg/mL, N IgG 8.04 +/- 6.3 pg/mL, P < 0.05) and IL-8 (anti-PR3 8.97 +/- 0.93 ng/mL, anti-MPO 8.45 +/- 1.46 ng/mL, N IgG 0.96 +/- 0.15 ng/mL, P < 0.01). CONCLUSION: In vivo circulating PMNs are primed as assessed by PR3 expression and basal superoxide production, thereby enhancing their inflammatory potential. These PMNs undergo apoptosis more readily, at which times they express PR3 and MPO on their surface. These antigens may then provide targets for ANCA. Opsonization of apoptotic PMN will enhance clearance by macrophages but will also trigger the release of pro-inflammatory cytokines that may contribute to chronic inflammation.     

Neutrophil phagocytosis during endotoxin-induced lung injury

Depressed neutrophil (PMN) phagocytosis in patients with ARDS may contribute to the known increased incidence of pulmonary sepsis. To evaluate changes in phagocytosis, circulating PMNs from normal rats were compared to circulating and alveolar PMNs (obtained by bronchoalveolar lavage, BAL) from rats after 72 hr of endotoxin infusion (LPS-Rx)-induced acute lung injury. Since phagocytosis correlates with adherence, PMN adherence to coverslips and to a standard nylon wool column was also measured. PMN adherence to nylon wool was 65% for control, 77% for circulating LPS-Rx, and 20% for BAL PMNs. As a measure of phagocytosis the PMNs were incubated for 30 min with opsonized fluorescent (FITC) tagged yeast. Total PMN with yeast were 95.4 +/- 2.1% for control; 96.4 +/- 1.8% for circulating LPS-Rx; and 78.7 +/- 7.8% (P less than 0.05 compared to control) for BAL PMNs. Total numbers of yeast particles per 100 PMN are 270 +/- 64 for control, 300 +/- 42 for circulating LPS-Rx, and 170 +/- 45 (P less than 0.05 compared to control) for BAL PMN. Conclusions: (1) Intraalveolar (BAL) PMNs have decreased adherence; (2) nonadherent PMNs have decreased uptake of yeast; (3) BAL PMNs, overall, have a significantly decreased uptake of yeast; (4) this depression in BAL PMN phagocytosis may partially explain the known decreased rate of bacterial clearance in injured lungs and the increased risk of pulmonary sepsis with adult respiratory distress syndrome.     

The effect of tumor necrosis factor-alpha and interferon-gamma on neutrophil function

Tumor necrosis factor-alpha (TNF) and interferon-gamma (IFN-gamma) have been shown to regulate cell-mediated immunity and act as effective modifiers of immune function; however, their influence on neutrophil (PMN) function is not well defined. This study investigated the effect of these cytokines on PMN phagocytosis, respiratory burst, and complement receptor (C3b) expression. Human citrated whole blood was incubated with either phosphate-buffered saline (control), 20 micrograms Escherichia coli lipopolysaccharide (LPS), TNF (1, 10, or 100 units), or IFN-gamma (1, 10, or 100 units). Synergy was also assessed between TNF and IFN-gamma. Phagocytosis and respiratory burst were assayed by sequential incubation of blood with dichlorofluorescein diacetate followed by labeled Staphylococcus aureus. C3b receptor expression was assayed by labeled anti-CR3 monoclonal antibody. Measurements were expressed as the mean channel fluorescence of 2000 PMNs counted by flow cytometry. IFN-gamma alone at all doses had no effect on any of the parameters measured. TNF 1 unit/ml increased phagocytosis (666 +/- 47 vs 542 +/- 19), respiratory burst (326 +/- 33 vs 258 +/- 17), and C3b (374 +/- 42 vs 157 +/- 14; all P less than 0.05) over those of control. TNF also demonstrated dose-dependent PMN activation. The combination of TNF + IFN-gamma increased both respiratory burst and C3b compared to either agent alone. These data indicate that TNF enhances PMN function and cytokine interaction may be important in PMN activation.     

Altered peripheral amino acid uptake in obstructive jaundice

To characterize amino acid metabolism in obstructive jaundice, the amino acid uptake in tissues of bile duct-ligated (BDL) rats was determined. Fischer 344 rats underwent either bile duct ligation or sham laparotomy and were pair fed for 72 hr. Amino acid uptake was determined in peripheral skeletal muscle (quadriceps femoris, soleus, and rectus abdominis), liver, blood, and other tissues by accumulation of alpha-[14C]-aminoisobutyric acid following intracardiac injection. Although total hepatic amino acid uptake was unaltered in the BDL animals compared with the sham-operated controls, amino acid uptake in peripheral skeletal muscle was significantly decreased in all muscle groups studied in the BDL rats. The relative concentration (percentage dose per gram normalized to the animal mass) for quadriceps femoris was 0.16 +/- 0.02 for BDL and 0.32 +/- 0.04 for sham-operated rats, P less than 0.005. Muscle protein was lower in BDL animals when compared with sham-operated rats (P less than 0.05). Total trunk blood amino acid levels were not significantly different in the two groups; however, there was a decreased serum level of branched-chain amino acids in the BDL group, P less than 0.05. No differences in plasma glucose or serum insulin were found in the two groups; lactate levels were lower in the BDL group, and plasma triglyceride levels were three times higher in the BDL animals. These data suggest that obstructive jaundice in the rat is associated with organ-specific metabolic abnormalities consistent with impaired peripheral amino acid uptake.     

Impairment of phagocyte oxidative metabolism in hemodialyzed patients with iron overload

In order to study the influence of iron overload on the polymorphonuclear leucocyte (PMN) metabolism of patients on chronic hemodialysis, generation of superoxide anion (O2-) by PMN in whole blood was compared in two groups of hemodialyzed patients: group A consisted of twenty-one individuals with serum ferritin levels above 1000 ng/ml and group B of nineteen individuals with serum ferritin levels below 1000 ng/ml. Whereas basal production of O2- was similar in the two groups (6.3 +/- 4.6 vs 11.5 +/- 8.3 nmoles O2- 10(6) granulocytes-1 15 min-1) (mean +/- s.e.m.), PMN response to opsonized zymosan was significantly lower in group A as compared with group B (86.5 +/- 6.3 vs 120.4 +/- 8.2 nmoles O2- 10(6) granulocytes-1 15 min-1) (p less than 0.01). Superoxide anion generation induced by the dialysis procedure was reduced in eight patients from group A (89.2 +/- 32.1) as compared with eight patients from group B (374.3 +/- 100.0 nmoles O2- 10(6) granulocytes-1 15 min-1) (p less than 0.05). These data suggest that iron overload may be involved in the impairment of neutrophil phagocytosis in patients on chronic hemodialysis.     

A potential basis for suppressed inflammatory cell function in pediatric cholestatic hosts

Infective mortality is common in children who have hepatic failure. We have demonstrated that experimental hepatic failure (EHF) profoundly suppresses T cell function in vivo. To determine the basis for immune suppression in EHF we postulated that this phenomenon is attributable to alterations in accessory macrophage (Ma) function, T cell subsets, interleukin-2 (IL-2) production, or serum inhibition. Wistar Furth rats (200 g) were randomized to EHF (n = 23), Sham (n = 23), and normal control (NC) (n = 23) groups. On day 21, splenocytes and sera were harvested and immune assays performed in vitro. Following are the results (mean +/- SEM; Student's t test). Serum bilirubin was elevated in EHF versus Sham and NC groups (P less than .01). EHF splenic macrophages suppressed PHA when added to microcultures at 10(5) concentration (-140 +/- 550 v 12,263 +/- 2,492 [Sham] and 21,413 +/- 1,702 [NC] P less than .01). This effect was not evident when macrophages were added back to microcultures at 10(3) and 10(4) concentrations, suggesting a dose-dependent inhibitory effect. T helper: suppressor ratios did not differ in EHF (1.3 +/- 0.2) compared with Sham (1.4 +/- 0.2) and NC groups (1.2 +/- 0.1). IL-2 production was similar in EHF, Sham, and NC animals (112,141 +/- 5,232 versus 106,691 +/- 1,419 and 120,759 +/- 3,249 counts per minute). T cell inhibitory activity was not demonstrable in EHF sera. These data show that splenic macrophages can inhibit T cell function in vitro. This phenomenon may be paramount in predisposing children with liver disease to infection.     

Thromboxane synthetase inhibition decreases polymorphonuclear leukocyte activation following hindlimb ischemia.

Ischemia of the lower extremity has been shown to cause pulmonary leukostasis and increased pulmonary artery pressure. Thromboxane (TX) has been implicated as a mediator in this process. The effect of OKY-046, a TX synthetase inhibitor, on polymorphonuclear leukocyte (PMN) production of superoxide anion (O2-) as determined by ferricytochrome reduction was examined. Fourteen dogs were subjected to 6 hours of bilateral gracilis muscle ischemia followed by 1 hour of reperfusion. O2- production from resting PMNs and PMNs stimulated with opsonized zymosan (OZ, 0.1 mg/ml) was measured prior to ischemia or drug treatment (baseline), and following reperfusion in both treated (n = 7) and control groups (n = 7). Serum TX levels were measured using a radioimmunoassay. Following reperfusion, TX levels in the treated group were decreased as compared with the control group (18 +/- 2 pg/ml vs. 72 +/- 26 pg/ml, P less than 0.05). Superoxide production by both resting and stimulated PMNs was also decreased in the treated group; from 0.98 +/- 0.16 nmol to 0.43 +/- 0.12 nmol O2- in the resting state (P less than 0.05) and from 13.3 +/- 1.5 nmol to 9.0 +/- 1.1 nmol O2- after stimulation (P less than 0.005). O2- production was increased in the control group following reperfusion as compared with baseline samples, and this increase was attenuated by treatment with OKY-046. TX synthetase inhibition decreases activation of PMNs following hindlimb ischemia.     

Vascular reactivity in reversible experimental obstructive jaundice

We studied the effect of jaundice on in vitro vascular reactivity to cumulative doses of norepinephrine (NE) by measuring the maximal response (Rmax) and the concentration of NE required to cause a 50% response (ED50) of isolated vascular smooth muscle. For this we prepared helically cut strips of thoracic aorta from bile duct ligated (BDL) rats at 1, 3, 6, 14, and 28 days postligation and compared them with those of nonoperated and sham-operated controls. From 1 to 6 days post-BDL, changes in liver blood chemistry and liver histology indicated cholestasis with necrosis. By 14 days, the tests for liver function and histology indicated a return to normal liver function and histology. In nonoperated controls, mean Rmax increased significantly from 883 +/- 67 mg of tension to 1220 +/- 68 mg of tension (P less than 0.0025) from 0 to 28 days, whereas ED50 remained unchanged. In sham-operated controls and BDL rats, an age-dependent increase in Rmax was also observed. However, in the sham groups, ED50 tended to decrease compared with nonoperated controls, indicating a surgically induced "sensitization" phenomenon of the vascular smooth muscle. In contrast, this was not seen in BDL rats since in these groups, the ED50 remained unchanged and significantly higher than in the sham groups, in both the jaundiced (1-6 days) and nonjaundiced (14-28 days) period. Furthermore, these changes occurred in the absence of any alteration in portal pressure. These changes may be important in understanding the mechanism of hypotension and shock in postoperative patients with obstructive jaundice even after the jaundice has been relieved.     

α-硫辛酸对阻塞性黄疸大鼠肝细胞线粒体内能量代谢保护作用及机制

目的 探讨α-硫辛酸对阻塞性黄疸大鼠肝细胞线粒体内能量代谢的保护作用及其机制. 方法 将72只SD雄性大鼠随机分为对照组(SO组),胆总管结扎+0.9%氯化钠溶液组(BDL+NS组),胆总管结扎α-硫辛酸组(BDL+LA组).分别于术后7 d、14 d和2l d检测肝细胞线粒体内MDA、SOD、ATP、ADP和AMP含量并计算总腺苷酸(TAN)和能荷(EC). 结果 BDL+NS组各时间点肝细胞线粒体内MDA、ADP、AMP含量明显升高,而SOD、ATP、EC含量却明显降低.胆总管结扎7 d、14 d时,肝细胞线粒体内MDA含量BDL+LA组比BDL+NS组低,差异有统计学意义(P＜0.01);2l d时,AMP、MDA含量BDL+NS组和BDL+LA中都更进一步升高,但两组比较差异无统计学意义(P＞0.05).胆总管结扎7 d、14 d时,肝细胞线粒体内SOD含量、ATP含量BDL+LA组比BDL+NS组下降程度有显著性差异(7 d,P＜0.01;14 d,P＜0.05);21 d时,两组肝细胞线粒体内SOD含量都进一步下降,但差异无统计学意义(p＞0.05). 结论 α-硫辛酸在阻塞性黄疸早、中期有保护线粒体能量代谢作用,减轻了阻塞性黄疸时肝的损伤.     

Intraoperative bupivacaine during outpatient hernia repair in children: a randomized double blind trial

Postoperative pain is a major problem following surgery in the ambulatory child. A study was undertaken to test the effect of intraoperative bupivacaine on postoperative pain in children undergoing outpatient hernia repair. Ninety-nine children aged 1 to 7 years underwent outpatient inguinal herniorrhaphy under general anesthesia. Each was randomly assigned to receive bupivacaine (group 1) or saline (group 2), infiltrating the ilioinguinal and iliohypogastric nerves. Drug administration and patient evaluation were double-blinded. The groups were similar with respect to age, sex, side of procedure, and length of operation. In the immediate postoperative period, 17 group 1 patients required analgesics compared with 39 in group 2 (P less than .01); total codeine dosage was lower in group 1 (4.0 +/- 7.1 mg v 11.8 +/- 10.5 mg, P less than .05). Activity level 45 minutes after surgery (using a standardized scale) was greater in group 1 (P less than .05). Acetaminophen requirements at home were lower in group 1 on the day of surgery (3.1 +/- 4.3 mL v 5.7 +/- 7.4 mL, P less than .05) and over the following 48 hours (1.5 +/- 3.4 mL v 4.9 +/- 10.7 mL, P less than .05). Activity level at home on the day of surgery did not differ significantly between groups, but activity level over the following 48 hours was higher in group 1 (P less than .05). The two groups were similar with respect to all other parameters. We conclude that intraoperative bupivacaine decreases post-operative pain and analgesic use, and promotes early ambulation in children undergoing hernia repair.     

Modulation of pulmonary permeability in vivo with agents that affect the cytoskeleton

In vitro studies show that the cytoskeleton of the microvascular barrier moderates polymorphonuclear leukocyte (PMN) diapedesis and the transport of macromolecules. This in vivo study tests indirectly whether the cytoskeleton of the pulmonary microvasculature responds similarly to agents known to reorganize the cytoskeleton to alter diapedesis and permeability in vitro. One of four agents, saline solution, cytochalasin B (which promotes actin microfilament disassembly), leukotriene (LT) B4 (PMN chemoattractant), or phalloidin (which promotes and stabilizes F-actin polymerization) was introduced into the bronchus of the anterior segment of the left lung of anesthetized rats (n = 152) through a tracheostomy and fine-bore cannula. Twenty minutes later, saline solution, cytochalasin B, LTB4, or phalloidin was similarly introduced. PMN accumulations (x 10(4) cells/ml), protein concentration in bronchoalveolar lavage fluid, and lung wet/dry weight ratios were measured 3 hours later. When the initial and secondary treatments were saline solution, PMN accumulations were 3 +/- 1 cells/ml and the protein level was 274 +/- 48 micrograms/ml. Secondary treatment of the saline-treated group with cytochalasin B or LTB4 led to increases in PMN to 18 +/- 2 and 9 +/- 2 cells/ml and protein to 960 +/- 50 and 840 +/- 40 micrograms/ml (p less than 0.05); secondary treatment with phalloidin was similar to that with saline solution. With saline solution used for both treatments, the wet/dry ratio was 3.4 +/- 0.2. Primary saline solution and secondary treatments with either cytochalasin B or LTB4 led to a similar increase in wet/dry ratios to 3.9 +/- 0.1 (p less than 0.05), whereas phalloidin was without effect (3.5 +/- 0.3). Initial cytochalasin B treatment followed by LTB4 led to increased PMN diapedesis (43 +/- 6 cells/ml and wet/dry ratio 4.3 +/- 0.2) (p less than 0.05). Secondary phalloidin treatment attenuated the cytochalasin B effect with counts of 12 +/- 2 PMN/ml, protein levels of 460 +/- 30 micrograms/ml, and a lower wet/dry ratio of 3.7 +/- 0.1 (p less than 0.05). Even more striking, phalloidin as initial treatment further reduced the cytochalasin B effect to 7 +/- 1 PMN/ml, whereas protein level was 490 +/- 60 micrograms/ml and wet/dry ratio was 3.5 +/- 0.1 (p less than 0.05). Further, phalloidin, given initially, attenuated the effect of secondary treatment with LTB4, resulting in fewer cells (4 +/- 2 PMN/ml), a lower wet/dry ratio (3.4 +/- 0.1), and protein level of 650 +/- 20 micrograms/ml (p less than 0.05).(ABSTRACT TRUNCATED AT 400 WORDS)     

Reversibility of leukocyte dysfunction in rats with obstructive jaundice

BACKGROUND: The role of leukocytes in obstructive jaundice is obscure and the effect of relieving cholestasis on leukocyte function is unclear. We postulated that cholestasis affects systemic polymorphonuclear leukocyte function by deranging phagocytosis and hydrogen peroxide release and the leukocyte dysfunction is reversible by internal and external biliary drainage. MATERIALS AND METHODS: Sixty male Sprague Dawley rats were randomly assigned to four groups: obstructive jaundice (OJ), sham operation (SH), OJ with internal drainage (ID), and OJ with external drainage (ED). The phagocytic functions of neutrophils and monocytes in whole blood were measured with flow cytometry using fluorescent microspheres. Intracellular hydrogen peroxide production by leukocytes was assessed with flow cytometry using dihydrorhodamine-123 as probes. RESULTS: Leukocyte count and percentage of monocytes in rats with OJ was significantly increased compared with SH rats (P < 0.001). These elevations could be reversed by both ID and ED method (P < 0.001). The phagocytic function of neutrophils and monocytes was significantly depressed in OJ rats compared with that in SH rats (P < 0.001). After relief of the OJ, the suppressed phagocytic function of neutrophils and monocytes was completely improved in ID rats (ID versus OJ, P < 0.001), but only partially reversed in ED rats. The hydrogen peroxide production by monocytes and lymphocytes was significantly increased in OJ rats (P < 0.05). ID reversed the increased hydrogen peroxide generation (P < 0.05), but ED only partially did. CONCLUSIONS: In our rodent model of biliary obstruction, deranged phagocytosis, and hydrogen peroxide generation by leukocytes was found. Internal drainage is superior to external drainage for reversal of the distorted leukocyte function.     

Effect of glyburide on glycemic control, insulin requirement, and glucose metabolism in insulin-treated diabetic patients

Glycemic control and glucose metabolism were examined in 5 patients with insulin-dependent diabetes mellitus (IDDM) and 8 insulin-treated non-insulin-dependent diabetes mellitus (NIDDM) patients before and after 2 mo of therapy with glyburide (20 mg/day). Glycemic control was assessed by daily insulin requirement, 24-h plasma glucose profile, glucosuria, and glycosylated hemoglobin. Insulin secretion was evaluated by glucagon stimulation of C-peptide secretion, and insulin sensitivity was determined by a two-step euglycemic insulin clamp (1 and 10 mU X kg-1. X min-1) performed with indirect calorimetry and [3-3H]glucose. In the IDDM patients, the addition of glyburide produced no change in daily insulin dose (54 +/- 8 vs. 53 +/- 7 U/day), mean 24-h glucose level (177 +/- 20 vs. 174 +/- 29 mg/dl), glucosuria (20 +/- 6 vs. 35 +/- 12 g/day) or glycosylated hemoglobin (10.1 +/- 1.0 vs. 9.5 +/- 0.7%). Furthermore, there was no improvement in basal hepatic glucose production (2.1 +/- 0.2 vs. 2.4 +/- 0.1 mg X kg-1 X min-1), suppression of hepatic glucose production by low- and high-dose insulin infusion, or in any measure of total, oxidative, or nonoxidative glucose metabolism in the basal state or during insulin infusion. C-peptide levels were undetectable (less than 0.01 pmol/ml) in the basal state and after glucagon infusion and remained undetectable after glyburide therapy. In contrast to the IDDM patients, the insulin-treated NIDDM subjects exhibited significant reductions in daily insulin requirement (72 +/- 6 vs. 58 +/- 9 U/day), mean 24-h plasma glucose concentration (153 +/- 10 vs. 131 +/- 5 mg/dl), glucosuria (14 +/- 5 vs. 4 +/- 1 g/day), and glycosylated hemoglobin (10.3 +/- 0.7 vs. 8.0 +/- 0.4%) after glyburide treatment (all P less than or equal to .05). However, there was no change in basal hepatic glucose production (1.7 +/- 0.1 vs. 1.7 +/- 0.1 mg X kg-1 X min-1), suppression of hepatic glucose production by insulin, or insulin sensitivity during the two-step insulin-clamp study. Both basal (0.14 +/- 0.05 vs. 0.32 +/- 0.05 pmol/ml, P less than .05) and glucagon-stimulated (0.24 +/- 0.07 vs. 0.44 +/- 0.09 pmol/ml) C-peptide levels rose after 2 mo of glyburide therapy and both were correlated with the decrease in insulin requirement (basal: r = .65, P = .08; glucagon stimulated: r = .93, P less than .001).(ABSTRACT TRUNCATED AT 400 WORDS)