A new efficient trial design for assessing reliability of ankle-brachial index measures by three different observer groups

BACKGROUND: The usual method of assessing the variability of a measure such as the ankle brachial index (ABI) as a function of different observer groups is to obtain repeated measurements. Because the number of possible observer-subject combinations is impractically large, only a few small studies on inter- and intraobserver variability of ABI measures have been carried out to date. The present study proposes a new and efficient study design. This paper describes the study methodology. METHODS: Using a partially balanced incomplete block design, six angiologists, six primary-care physicians and six trained medical office assistants performed two ABI measurements each on six individuals from a group of 36 unselected subjects aged 65–70 years. Each test subject is measured by one observer from each of the three observer groups, and each observer measures exactly six of the 36 subjects in the group. Each possible combination of two observers occurs exactly once per patient and is not repeated on a second subject. The study involved four groups of 36 subjects (144), plus standbys. RESULTS: The 192 volunteers present at the study day were similar in terms of demographic characteristics and vascular risk factors: mean age 68.6 ± 1.7; mean BMI 29.1 ± 4.6; mean waist-hip ratio 0.92 ± 0.09; active smokers 12%; hypertension 60.9%; hypercholesterolemia 53.4%; diabetic 17.2%. A complete set of ABI measurements (three observers performing two Doppler measurements each) was obtained from 108 subjects. From all other subjects at least one ABI measurement was obtained. The mean ABI was 1.08 (± 0.13), 15 (7.9%) volunteers had an ABI <0.9, and none had an ABI >1.4, i.e. a ratio that may be associated with increased stiffening of the arterial walls. CONCLUSION: This is the first large-scale study investigating the components of variability and thus reliability in ABI measurements. The advantage of the new study design introduced here is that only one sixth of the number of theoretically possible measurements is required to obtain information about measurement errors. Bland-Altman plots show that there are only small differences and no systematic bias between the observers from three occupational groups with different training backgrounds

The nuclear receptor PPARγ selectively inhibits Th17 differentiation in a T cell–intrinsic fashion and suppresses CNS autoimmunity

T helper cells secreting interleukin (IL)-17 (Th17 cells) play a crucial role in autoimmune diseases like multiple sclerosis (MS). Th17 differentiation, which is induced by a combination of transforming growth factor (TGF)-β/IL-6 or IL-21, requires expression of the transcription factor retinoic acid receptor–related orphan receptor γt (RORγt). We identify the nuclear receptor peroxisome proliferator–activated receptor γ (PPARγ) as a key negative regulator of human and mouse Th17 differentiation. PPARγ activation in CD4+ T cells selectively suppressed Th17 differentiation, but not differentiation into Th1, Th2, or regulatory T cells. Control of Th17 differentiation by PPARγ involved inhibition of TGF-β/IL-6–induced expression of RORγt in T cells. Pharmacologic activation of PPARγ prevented removal of the silencing mediator for retinoid and thyroid hormone receptors corepressor from the RORγt promoter in T cells, thus interfering with RORγt transcription. Both T cell–specific PPARγ knockout and endogenous ligand activation revealed the physiological role of PPARγ for continuous T cell–intrinsic control of Th17 differentiation and development of autoimmunity. Importantly, human CD4+ T cells from healthy controls and MS patients were strongly susceptible to PPARγ-mediated suppression of Th17 differentiation. In summary, we report a PPARγ-mediated T cell–intrinsic molecular mechanism that selectively controls Th17 differentiation in mice and in humans and that is amenable to pharmacologic modulation. We therefore propose that PPARγ represents a promising molecular target for specific immunointervention in Th17-mediated autoimmune diseases such as MS

Nitric Oxide-Mediated Regulation of Gamma Interferon-Induced Bacteriostasis: Inhibition and Degradation of Human Indoleamine 2,3-Dioxygenase

Tryptophan depletion resulting from indoleamine 2,3-dioxygenase (IDO) activity within the kynurenine pathway is one of the most prominent gamma interferon (IFN-γ)-inducible antimicrobial effector mechanisms in human cells. On the other hand, nitric oxide (NO) produced by the inducible isoform of NO synthase (iNOS) serves a more immunoregulatory role in human cells and thereby interacts with tryptophan depletion in a number of ways. We investigated the effects of NO on IDO gene transcription, protein synthesis, and enzyme activity as well as on IDO-mediated bacteriostasis in the human epithelial cell line RT4. IFN-γ-stimulated RT4 cells were able to inhibit the growth of Staphylococcus aureus in an IDO-mediated fashion, and this bacteriostatic effect was abolished by endogenously produced NO. These findings were supported by experiments which showed that IDO activity in extracts of IFN-γ-stimulated cells is inhibited by the chemical NO donors diethylenetriamine diazeniumdiolate, S-nitroso-l-cysteine, and S-nitroso-N-acetyl-d,l-penicillamine. Furthermore, we found that both endogenous and exogenous NO strongly reduced the level of IDO protein content in RT4 cells. This effect was not due to a decrease in IDO gene transcription or mRNA stability. By using inhibitors of proteasomal proteolytic activity, we showed that NO production led to an accelerated degradation of IDO protein in the proteasome. This is the first report, to our knowledge, that demonstrates that the IDO is degraded by the proteasome and that NO has an effect on IDO protein stability