An international collaborative study to establish the WHO 1st international standard for alpha-1-antitrypsin

Background and Objectives The aim was to establish the 1st International Standard (IS) for alpha‐1‐antitrypsin (AAT) to standardise potency assignment of therapeutic products, calibrated in moles and mg active AAT in line with product labelling practice. Assigning total protein and antigen values to the IS was also investigated. Materials and Methods The active concentration of four candidate AAT preparations was determined in an international collaborative study by inhibition of trypsin (calibrated by active‐site titration). Total protein and antigen content were determined for each candidate using local methods and in‐house standards, and a common AAT preparation. The total protein content of the IS was also determined by amino acid analysis. Potency determination of recombinant and transgenic materials against the IS was investigated in a follow‐up study. Results Data analysis for potency determination indicated no statistical difference between any of the candidates, or between the results for recombinant and plasma‐derived products. Total protein content of the IS determined by amino acid analysis was consistent with the potency value. The variability in the total protein and antigen results for the other candidates was reduced when the data were recalculated relative to the IS. Conclusions Candidate C (05/162) was established by the WHO Expert Committee on Biological Standardization (ECBS) in 2006 as the WHO 1st IS for AAT with a potency of 243 nmoles (12·4 mg) active inhibitor per ampoule. In 2008, ECBS approved the IS for potency determination of recombinant material and assigned a total protein and antigen value of 12·4 mg.     

Induction of LFA-1-dependent neutrophil rolling on ICAM-1 by engagement of E-selectin

OBJECTIVE: To study rolling of mouse neutrophils on E-selectin and ICAM-1 in an ex vivo flow chamber system. METHODS: The authors developed a small autoperfused flow chamber (20 x 200-microm cross section) that allows direct visualization of cells with and without fluorescent labeling and does not require recirculation of blood. RESULTS: Neutrophils rolled on E-selectin alone, but were unable to interact with immobilized ICAM-1. When ICAM-1 was co-immobilized with E-selectin, the number of cells that rolled was doubled, but no significant firm adhesion was observed. This phenomenon was specific for E-selectin, and no enhancement of rolling was observed when P-selectin was immobilized with ICAM-1. The increased neutrophil rolling seen on E-selectin and ICAM-1 substrates required beta2 integrins. Treating mice with antibodies to the beta2 integrins LFA-1 and Mac-1 showed that LFA-1 was primarily responsible for mediating rolling on ICAM-1 in this model. Increased rolling on E-selectin and ICAM-1 was significantly reduced following administration of a specific p38 mitogen-activated protein kinase (MAPK) inhibitor. CONCLUSION: The data show that neutrophil rolling on E-selectin leads to partial activation of LFA-1, enabling LFA-1-dependent rolling on ICAM-1. This mechanism is likely to amplify and accelerate neutrophil recruitment in inflammation.     

The relationship between ApoE, TNFA, IL1a, IL1b and IL12b genes and HIV-1-associated dementia

Objectives

Host genetic factors implicated in AIDS dementia complex (ADC) were studied.

Methods

DNA from ADC patients (n=56), unselected HIV‐seropositive patients (n=112, 171, 185 and 204) and HIV‐seronegative controls (n=204, 60, 60, 96 and 624) were typed for polymorphic loci in genes encoding tumour necrosis factor (TNF)‐α, interleukin (IL)‐1α, IL‐1β, IL‐12 and Apolipoprotein E (ApoE). Diagnosis of ADC was based on neurological symptoms, signs and neuroimaging findings with other causes of dementia excluded. Patients selected had ADC stage ≥1 and CD4 counts of <500 cells/μL.

Results

Allele 2 of TNFA‐308 was more common in ADC patients compared to HIV‐positive or HIV‐negative controls (P=0.005, 0.024). No other differences between ADC patients and control groups were significant. Meta‐analyses confirmed these results.

Conclusions

This study suggests that TNFA‐308 allele 2 or an allele in linkage disequilibrium with this locus influences ADC.     

Regulation of T-lymphocyte trafficking by ICAM-1, MAdCAM-1, and CCR7 in microcirculation of appendicular and intestinal lymphoid tissues

OBJECTIVE: Although the appendix is recognized as an inductive site of intestinal inflammation, lymphocyte migration to lymphoid tissues of the appendix has not been characterized. The authors investigated if there are specific features in T-lymphocyte adhesion to microvessels of the appendix compared to mouse Peyer's patches (PPs). METHODS: T-lymphocyte interaction with postcapillary venules (PCVs) of lymph follicles of the appendix and PPs was observed using an intravital microscope. Antibodies against ICAM-1, MAdCAM-1, or anti-L-selectin were administered prior to lymphocyte administration, and in some experiments CCR7 on T-lymphocytes was desensitized by excess CCL21. RESULTS: The number of adhered T-lymphocytes reached the maximum value earlier in PCVs of PPs than in those of the appendix. T-lymphocyte adherence was significantly inhibited by anti-MAdCAM-1 at either the appendix or PPs, but adherence in the appendix was also significantly inhibited by anti-ICAM-1, suggesting a dependency on ICAM-1 in the appendix. Histologically, there was a significant ICAM-1 expression in the appendix. Desensitization of CCR7 suppressed T-cell adhesion in PCVs of the appendix and PPs to the same extent. CONCLUSION: ICAM-1 appeared to be more important in T-lymphocyte sticking in PCVs of the appendix compared with intestinal PPs, while MAdCAM-1 and CCR7 were shown to play important roles in T-lymphocyte adherence in all sites.     

Associations between interleukin-1 (IL-1) gene variations or IL-1 receptor antagonist levels and the development of type 2 diabetes

Abstract. Luotola K, Pietilä A, Zeller T, Moilanen L, Kähönen M, Nieminen MS, Kesäniemi YA, Blankenberg S, Jula A, Perola M, Salomaa V (Helsinki University Hospital, Helsinki; National Institute for Health and Welfare, Helsinki, Finland; University Medical Center Mainz, Johannes Gutenberg University Mainz, Mainz, Germany; University Hospital of Kuopio, Kuopio; Tampere University Hospital and Medical School, University of Tampere, Tampere; University of Oulu and Clinical Research Center, Oulu University Hospital, Oulu; National Institute for Health and Welfare, Turku; and Institute for Molecular Medicine Finland, Helsinki, Finland). Associations between interleukin‐1 (IL‐1) gene variations or IL‐1 receptor antagonist levels and the development of type 2 diabetes. J Intern Med 2010; 269 : 322–332. Objectives. To examine whether interleukin‐1 receptor antagonist (IL‐1Ra) is a predictor for clinically incident diabetes in subjects with metabolic syndrome (MetS) and whether its predictive power is independent of C‐reactive protein (CRP), an established marker of inflammation. We further examined whether genetic variants at the interleukin‐1 (IL‐1) locus would predict clinically incident diabetes. Design. Two observational prospective cohort studies. Setting. Two separate cohorts, Health 2000 and FINRISK 1997, followed up for an average of 7.1 and 10.8 years, respectively. Subjects. Random population samples consisting of 5511 subjects aged 30–74 years in Health 2000 and 7374 subjects aged 25–74 years in FINRISK 1997. Results. During follow‐up, 141 cases of clinically incident diabetes were observed amongst subjects with MetS at baseline in Health 2000 and 248 cases in FINRISK 97. After adjustment for multiple traditional risk factors of diabetes, including age and body mass index, IL‐1Ra was a significant (P < 0.01) predictor of incident diabetes amongst men in both cohorts and amongst women in FINRISK 1997. Further adjustment for CRP reduced the hazard ratios only slightly. Genetic analyses produced nominally significant associations for three single‐nucleotide polymorphisms: rs3213448 in IL‐1 receptor antagonist (IL1RN), rs1143634 in IL‐1 beta (IL1B) and rs1800587 in IL‐1 alpha (IL1A). The two latter variants had an interaction with gender (P = 0.023 and 0.002, respectively) suggesting the presence of gender‐specific associations with the risk of clinically incident diabetes. Conclusions. IL‐1Ra predicted the progression of MetS to clinically incident diabetes independently of CRP and other risk factors. Genetic variation in the IL‐1 locus may have gender‐specific associations with the risk of type 2 diabetes.     

Effect of ICAM‐1 and LFA‐1 in hyperleukocytic acute myeloid leukaemia

Acute hyperleukocytic leukemia [AHL; WBC count >100 × 10⁹/l] is associated with a life‐threatening complication. The mechanisms of hyperleukocytosis in acute myeloid leukaemia (AML) remain unclear. However, the interaction of intercellular adhesion molecule‐1 (ICAM‐1) and lymphocyte function‐associated antigen‐1 (LFA‐1) plays an important role in the adhesion and migration of normal leukocytes and AML cells. Therefore, effects of ICAM‐1 and LFA‐1 were studied in hyperleukocytic AML. The adhesion of hyperleukocytic AML blasts and human umbilical vein endothelial cells (HUVECs) was significantly increased compared with that of blasts from non‐hyperleukocytic AML (WBC < 100 × 10⁹/l). The adhesion of normal neutrophils and HUVECs treated with hyperleukocytic AML blast supernatant was increased significantly. Finally, we determined the ICAM‐1 on the surface of HUVECs treated with the supernatant of hyperleukocytic AML blasts and LFA‐1 on hyperleukocytic AML blasts by flow cytometry. It showed that the ICAM‐1 expression on the surface of the HUVECs treated with hyperleukocytic AML blast supernatant for 24 h could be increased, and the expression of LFA‐1 on hyperleukocytic AML was also increased significantly. Our data show that hyperleukocytic AML blasts stimulate the endothelium to secrete more ICAM‐1 and promote their own adhesion to vascular endothelium, suggesting that ICAM‐1 and LFA‐1 may have a role in hyperleukocytic AML.     

Glucagon-like peptide 1(GLP-1) in biology and pathology

Post-translational proteolytic processing of the preproglucagon gene in the gut results in the formation of glucagon-like peptide 1 (GLP-1). Owing to its glucose-dependent insulinotropic effect, this hormone was postulated to primarily act as an incretin, i.e. to augment insulin secretion after oral glucose or meal ingestion. In addition, GLP-1 decelerates gastric emptying and suppresses glucagon secretion. Under physiological conditions, GLP-1 acts as a part of the 'ileal brake', meaning that is slows the transition of nutrients into the distal gut. Animal studies suggest a role for GLP-1 in the development and growth of the endocrine pancreas. In light of its multiple actions throughout the body, different therapeutic applications of GLP-1 are possible. Promising results have been obtained with GLP-1 in the treatment of type 2 diabetes, but its potential to reduce appetite and food intake may also allow its use for the treatment of obesity. While rapid in vivo degradation of GLP-1 has yet prevented its broad clinical use, different pharmacological approaches aiming to extend the in vivo half-life of GLP-1 or to inhibit its inactivation are currently being evaluated. Therefore, antidiabetic treatment based on GLP-1 may become available within the next years. This review will summarize the biological effects of GLP-1, characterize its role in human biology and pathology, and discuss potential clinical applications as well as current clinical studies.     

Influenza A H1N1 in HIV-infected adults

Objectives

HIV‐infected adults are considered to be at higher risk for influenza A H1N1 complications but data supporting this belief are lacking. We aimed to compare epidemiological data, clinical characteristics, and outcomes of influenza A H1N1 infection between HIV‐infected and ‐uninfected adults.

Methods

From 26 April to 6 December 2009, each adult presenting with acute respiratory illness at the emergency department of our institution was considered for an influenza A H1N1 diagnosis by specific multiplex real‐time polymerase chain reaction. For every HIV‐infected adult diagnosed, three consecutive adults not known to be HIV‐infected diagnosed in the same calendar week were randomly chosen as controls.

Results

Among 2106 adults tested, 623 (30%) had influenza A H1N1 infection confirmed. Fifty‐six (9%) were HIV‐positive and were compared with 168 HIV‐negative controls. Relative to HIV‐negative controls, HIV‐positive patients were older, more frequently male, and more frequently smokers (P≤0.02). In the HIV‐positive group, prior or current AIDS‐defining events were reported for 30% of patients, 9% and 30% had CD4 counts of <200 and 200–500 cells/μL, respectively, and 95% had HIV‐1 RNA <50 copies/mL. Pneumonia (9%vs. 25%, respectively, in the HIV‐positive and HIV‐negative groups; P=0.01) and respiratory failure (9%vs. 21%, respectively; P=0.04) were less common in the HIV‐positive group. Oseltamivir (95%vs. 71% in the HIV‐positive and HIV‐negative groups, respectively; P=0.003) was administered more often in HIV‐positive patients. Three patients (all HIV‐negative) died. In the HIV‐positive group, CD4 cell count and plasma HIV‐1 RNA did not differ before and 4–6 weeks after influenza A H1N1 diagnosis (P>0.05).

Conclusions

HIV infection did not increase the severity of influenza A H1N1 infection, and influenza A H1N1 infection did not have a major effect on HIV infection.     

Do plasminogen activator inhibitor (PAI-1) or tissue plasminogen activator PAI-1 complexes predict complications in Type 1 diabetes: the Pittsburgh Epidemiology of Diabetes Complications Study.

AIMS: To examine the predictive power of plasminogen activator inhibitor-1 (PAI-1) and the complexes it forms with tissue plasminogen activator (tPA-PAI-1) for the two major Type 1 diabetes (T1D) complications (coronary artery disease (CAD) and overt nephropathy) in the context of standard risk factors. METHODS: Observational prospective study of 454 participants with childhood onset (< 17 years) T1D, aged 18+ years at baseline. PAI-1 and tPA-PAI-1 were determined using ELISA methodology. Follow-up (6 years) was limited to 382 individuals for CAD and 294 individuals for overt nephropathy, after excluding baseline cases. Total, HDL and LDL-cholesterol, triglycerides, HbA1, blood pressure, body mass index (BMI), waist-hip ratio (WHR), leucocyte count, Beck depression score and fibrinogen were also examined. RESULTS: The 56 incident cases of CAD had marginally lower PAI-1 and higher tPA-PAI-1 levels compared with those free of CAD. However, marginally higher PAI-1 and significantly higher tPA-PAI-1 (P = 0.04) levels were seen in those who developed nephropathy. After controlling for age, both PAI-1 and tPA-PAI-1 showed significant negative correlations with HDL-cholesterol, and positive correlations with triglycerides, WHR, HbA1 and fibrinogen. tPA-PAI-1 was also positively correlated with total and LDL-cholesterol. In multivariate analyses, neither PAI-1 nor tPA-PAI-1 was an independent predictor of CAD or overt nephropathy. CONCLUSIONS: These results suggest little association between PAI-1 and later CAD in patients with T1D. However, tPA-PAI-1 complexes may be involved in the pathogenesis of overt nephropathy.     

PD‐1/PD‐L1 pathway and T‐cell exhaustion in chronic hepatitis virus infection

Summary. Dysfunctional virus‐specific T cells are a hallmark of many chronic viral infections. Recent studies have implicated the inhibitory PD‐1/PD‐L1 pathway with the functional impairment of T cells. In this respect, we will review the latest research on PD‐1/PD‐L1 pathway and T‐cell exhaustion in the context of human chronic hepatitis B and C virus infections. We will also discuss the therapeutic potential of PD‐1 blockade and how it may be enhanced through the modulation of other co‐stimulatory/inhibitory pathways.