The potentiation of human C1-inhibitor by dextran sulphate is transient in vivo: studies in a rat model

C1-inhibitor (C1-Inh) is an important regulator of inflammatory reactions because it is a potent inhibitor of the contact and complement system. C1-Inh application in inflammatory disease is, however, restricted because of the high doses required. The glycosaminoglycan-like molecule dextran sulphate (DXS) enhances C1-Inh function in vitro. Hence, we investigated whether co-administration with dextran sulphate reduces the amount of C1-Inh required, through enhancement in vivo. C1-Inh potentiation was measured in a newly developed C1s-inactivation assay that is based on activation of C4 by purified C1s. Activated C4 in rat plasma was quantified with a newly developed ELISA. Human C1-Inh (2.5 microM) inhibited C1s in rat plasma 55-fold faster in the presence of dextran sulphate (15 kDa, 5 microM). To study the stability of the complex in vivo, rats were given a mixture of C1-Inh (10 mg/kg) and dextran sulphate (3 mg/kg). C1-Inh activity during 5 h was analyzed ex vivo with the C1s inactivation assay. The noncovalent C1-Inh-dextran sulphate complex resulted in a transient enhancement of the inhibitory capacity of C1-Inh, lasting for 60-90 min. Dextran sulphate did not affect plasma clearance of C1-Inh. We conclude that the enhanced inhibitory capacity of C1-Inh complexed to dextran sulphate is transient in vivo. Hence, co-administration of these compounds seems a feasible approach to achieve short-term inhibition of complement in vivo.     

The prevalence of latex allergy in operating theatre staff

Latex allergy has become increasingly common amongst health care workers. The prevalence of latex allergy in 102 theatre personnel at Princess Alexandra Hospital was determined by the results of a standardized questionnaire and a latex specific IgE radioallergosorbent test (RAST). Volunteers had their forced expiratory volume in 1 second/forced vital capacity (FEV1/FVC) measured at the beginning and end of their working shifts. Only one of 102 volunteers had a positive latex specific IgE RAST and he also experienced both local and systemic symptoms with latex exposure and a deterioration in daily FEV1/FVC. The 14 volunteers using asthma medications had significantly higher incidence of operating theatre symptoms (10/14), local reactions (8/14), and atopy (14/14). There was no clinically significant daily change in FEV1 in the total population or any specific group. The 1% (1/102) prevalence of latex allergy and sensitization in theatre staff demonstrates a much lower incidence than previously reported (12.5%, 15.8%).     

Early inhibition of activated fibrinolysis predicts microbial infection, shock and mortality in febrile medical patients

To evaluate the contribution of an imbalance between coagulation activation and fibinolysis activation and inhibition to morbidity and mortality in sepsis, we determined in medical hospitalized patients at inclusion (day 0) for fever (temperature above 38.0 degrees C axillary or 38.3 degrees C rectally), and daily thereafter for two days, circulating thrombin-antithrombin III (TAT) complexes, plasmin-alpha2-antiplasmin (PAP) complexes (day 0 only), tissue-type plasminogen activator (t-PA), plasminogen activator inhibitor-1 (PAI-1) and interleukin (IL)-6, the latter as a marker of the inflammatory host response. Study variables were 1) positive microbiological results for specimens from local sites associated with a clinical infection, positive blood cultures (including parasitemia) or both, within 7 days after inclusion, 2) development of shock, i.e. systolic blood pressure <90 mmHg or a reduction of 40 mmHg from baseline within 7 days after inclusion, and 3) death related to febrile illness within 28 days after inclusion. The peak plasma levels of TAT complexes were elevated in 44% and the PAP complexes in all patients. The t-PA and PAI-1 levels were elevated in 74 and 94% of patients, respectively. Values for TAT and PAP did not differ among subgroups, while peak t-PA and IL-6 levels were higher in patients with positive microbiological results, developing shock or ultimately dying than in those without the complications (p<0.005). Peak PAI-1 levels were elevated in patients developing shock and ultimate death versus those with an uncomplicated course (p <0.05). Peak IL-6 related to PAI-1 and t-PA levels, which interrelated. Patients with elevated TAT levels had increased plasma levels of IL-6, PAP, PAI-1 and t-PA versus those with normal TAT (p <0.05). Our data indicate that inhibition of activated fibrinolysis, which may partly depend on both cytokinemia and activation of coagulation, predicts microbial infection, septic shock and mortality of febrile medical patients. This suggests an early pathogenic role of inhibition of activated fibrinolysis in the downhill course of serious microbial infection.     

Glomerular complement regulation is overwhelmed in passive Heymann nephritis

BACKGROUND: An injection of anti-Fx1A antibodies in rats leads to passive Heymann nephritis (PHN), a model of membranous nephropathy. Fx1A is a crude extract of renal cortex that contains megalin as a principal component. However, when rats are given anti-megalin antibodies, abnormal proteinuria does not occur. Because of the established complement dependence of PHN, we hypothesized that antibodies neutralizing complement regulatory proteins in the rat glomerulus also were required to induce PHN. Two likely targets are Crry and CD59, proteins abundant on the rat podocyte and contained within Fx1A that inhibit the C3 convertase and C5b-9 assembly, respectively. METHODS: Rats were injected with anti-megalin monoclonal antibodies, followed by anti-Crry and/or anti-CD59 F(ab')(2) antibodies five days later. In a second group of experiments, rats were injected with anti-Fx1A or anti-Fx1A immunodepleted of reactivity against Crry and/or CD59. RESULTS: In the setting of podocyte-associated anti-megalin monoclonal antibodies, simultaneous neutralization of Crry and CD59 function led to the development of significant proteinuria (11.0 +/- 2.1 mg/day, P < 0.001 vs. all other groups). In contrast, animals that had neither or only one of these complement regulators inhibited had normal urinary protein excretion (< or =6 mg/day). In animals given anti-Fx1A depleted of anti-Crry and/or anti-CD59, all groups developed typical PHN, characterized by heavy proteinuria and extensive glomerular deposition of C3 and C5b-9. CONCLUSION: Crry and CD59 play an important role in restraining complement-mediated injury following subepithelial immune complex deposition; however, in PHN, their regulatory capacity is overwhelmed.     

Human granzyme B mediates cartilage proteoglycan degradation and is expressed at the invasive front of the synovium in rheumatoid arthritis

OBJECTIVE: To investigate the cartilage-degrading capacity of granzyme B and the presence of granzyme B-positive cells at sites of erosion in the rheumatoid synovium. METHODS: Granzyme B was added to [(3)H]proline/[(35)S]sulphate-labelled cartilage matrices and to cartilage explants. Proteoglycan degradation was assessed by the release of (35)S and glycosaminoglycans into the medium and collagen degradation was assessed by the release of (3)H and hydroxyproline and by measuring the fraction of denatured collagen. Granzyme B expression was studied at the invasive front of the synovium by immunohistochemistry. RESULTS: Granzyme B induced loss of both newly synthesized, radiolabelled proteoglycans in cartilage matrices and resident proteoglycans of the cartilage explants. No effect on collagen degradation was found. Granzyme B-positive cells were present throughout the synovium and at the invasive front. CONCLUSION: The presence of granzyme B-positive cells at the invasive front of the synovium together with its ability to degrade articular proteoglycans supports the view that granzyme B may contribute to joint destruction in rheumatoid arthritis.     

Heterogeneous distribution of AMPA glutamate receptor subunits at the photoreceptor synapses of rodent retina

In the retina the segregation of different aspects of visual information starts at the first synapse in signal transfer from the photoreceptors to the second-order neurons, via the neurotransmitter glutamate. We examined the distribution of the four AMPA glutamate receptor subunits GluR1-GluR4 at the photoreceptor synapses in mouse and rat retinae by light and immunoelectron microscopy and serial section reconstructions. On the dendrites of OFF-cone bipolar cells, which make flat, noninvaginating contacts postsynaptic at cone synaptic terminals, the subunits GluR1 and GluR2 were predominantly found. Horizontal cell processes postsynaptic at both rod and cone synaptic terminals preferentially expressed the subunits GluR2, GluR2/3 and GluR4. An intriguing finding was the presence of GluR2/3 and GluR4 subunits on dendrites of putative rod bipolar cells, which are thought to signal through the sign-inverting metabotropic glutamate receptor 6, mGluR6. Furthermore, at the rod terminals, horizontal cell processes and rod bipolar cell dendrites showed labelling for the AMPA receptor subunits at the ribbon synaptic site or perisynaptically at their site of invagination into the rod terminal. The wide distribution of AMPA receptor subunits at the photoreceptor synapses suggests that AMPA receptors play an important role in visual signal transfer from the photoreceptors to their postsynaptic partners.     

Placental lesions as predictors of cerebral palsy and abnormal neurocognitive function at school age in extremely low birth weight infants (<1 kg).

Extremely low birth weight (ELBW) infants (<1 kg) have high rates of neurodisability. Although previous studies have implicated placental lesions in adverse short-term neurologic outcomes in this population, none have assessed their effects in these children once they reach school age. We conducted a secondary analysis of placental pathology in a cohort study of inborn singleton ELBW infants born between 1992 and 1995 and evaluated for cerebral palsy (CP) and abnormal neurocognitive testing at 8 years of age (N = 129). The neurocognitive tests were the Kaufman Assessment Battery for Children (K-ABC) and 6 subtests of the NEPSY: A Developmental Neuropsychological Assessment. We found that placental lesions associated with maternal vascular underperfusion (increased syncytial knots and acute atherosis) were risk factors for CP, while villous edema was associated with low scores on both neurocognitive tests. Histologic chorioamnionitis (HCA) was not predictive of outcome in the population as a whole, but a severe fetal vascular response was associated with a lower NEPSY score in the subpopulation with HCA (N = 69). Placentas with increased syncytial knots, villous edema, and those with neither finding constituted nonoverlapping subgroups with distinct pathologic and perinatal characteristics. Among infants with villous edema (N = 25), those with neurologic impairment had lower gestational ages and more severe degrees of HCA. However, by logistic regression these other factors were not independent risk factors for abnormal neurocognitive testing, and only HCA with a severe fetal vascular response decreased the association of villous edema with low test scores for NEPSY, but not K-ABC.     

Activation of clotting factors XI and IX in patients with acute myocardial infarction

In acute coronary events, plaque rupture and the subsequent formation of the catalytic tissue factor-factor VIIa complex is considered to initiate coagulation. It is unknown whether clotting factors XI and IX are activated in acute coronary events. Therefore, we prospectively investigated the activation of clotting factors XI and IX as well as activation of the contact system and the common pathway in 50 patients with acute myocardial infarction (AMI), in 50 patients with unstable angina pectoris (UAP), and in 50 patients with stable angina pectoris (SAP). Factor XIa-C1 inhibitor complexes, which reflect acute activation of factor XI, were detected in 24% of the patients with AMI, 8% of the patients with UAP, and 4% of the patients with SAP (P<0.05), whereas factor XIa-alpha(1)-antitrypsin complexes, which reflect chronic activation, were observed equally in all 3 study groups. Factor IX peptide levels were significantly higher in the patients with AMI and UAP compared with the patients with SAP (P<0.01). No differences regarding markers of the common pathway were demonstrated. Fibrinopeptide A levels were elevated in patients with AMI compared with patients with UAP and those with SAP (P<0.01). Factor XIIa- or kallikrein-C1 inhibitor complexes were not increased. In conclusion, this is the first demonstration of the activation of clotting factors XI and IX in patients with acute coronary syndromes. Because these clotting factors are considered to be important for continuous thrombin generation and clot stability, their activation might have clinical and therapeutic consequences.     

Physical and psychological morbidity after axillary lymph node dissection for breast cancer.

PURPOSE: Alternatives to axillary lymph node dissection (ALND) are being developed amid controversy surrounding the therapeutic benefit and overall utility of this routine surgical procedure. Although potential negative side effects associated with ALND are known, we set out to examine whether these side effects contribute significantly to patient reports of quality of life and mental health. PATIENTS AND METHODS: We surveyed 222 women who had received an ALND as part of breast cancer surgery. All women underwent a physical therapy assessment of range of arm/shoulder motion and completed the Modified Post-operative Pain Questionnaire, the Pain Disability Index, the McGill Pain Questionnaire (short form), the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire, and the Mental Health Inventory. RESULTS: Seventy-two percent of the women experienced arm/shoulder pain, weakness, or numbness in the week before the interview, and range of motion of the affected arm/shoulder was impaired in 73% of the women. Severity of pain was reported to be low to moderate, and younger patients experienced greater pain than older patients. Pain severity correlated positively with the number of lymph nodes removed and receipt of chemotherapy and was not significantly related to length of time since surgery or receipt of radiation therapy. Generally high levels of cancer-specific quality of life and mental health were reported. Quality of life was significantly predicted by the McGill Pain Questionnaire, and mental health was significantly predicted by the Pain Disability Index and the physical therapy assessment. CONCLUSION: Surgery-related symptoms after ALND persist for a majority of women with breast cancer and are not significantly related to time since surgery or receipt of radiation therapy. These symptoms and associated disability are significantly predictive of cancer-specific quality of life and mental health.