Increased restenosis in diabetes mellitus after coronary interventions is due to exaggerated intimal hyperplasia. A serial intravascular ultrasound study

Hepatic hydrothorax is a rare complication of portal hypertension. Conservative therapy may be successful but refractory hepatic hydrothorax is not uncommon. Management of refractory hydrothorax is usually ineffective and can result in a worsened clinical status. Transjugular intrahepatic portosystemic shunts (TIPS) lower portal pressure and have been used in the treatment of refractory ascites. The aim of this study was to determine the efficacy of TIPS in the treatment of symptomatic refractory hepatic hydrothorax. A TIPS was placed in 24 consecutive cirrhotic patients with symptomatic refractory hepatic hydrothorax. Five patients (20.8%) were Child's/Pugh class B and 19 (79.2%) were class C. All had undergone multiple thoracenteses and were hypoalbuminemic. Mean follow-up was 7.2 months (range, 0.25-49 months). Fourteen (58.3%) of 24 patients had complete relief of symptoms after shunt placement and did not require further thoracentesis. Five (20.8%) additional patients required fewer thoracenteses. Five (20.8%) patients developed worsening liver function and died within 45 days. In eight (66.7%) of 12 patients with > or = 60 days of follow-up, the serum albumin increased by a mean of 1.2 g/dL (range, 0.1-2.2 g/dL). The Child's-Pugh score improved in 7 (58.3%) of these 12 patients and two patients improved from class C to class A. These two patients no longer require liver transplantation. This study shows that TIPS can be effective in the management of symptomatic, refractory hepatic hydrothorax. Clinical and laboratory improvement may be seen and liver transplantation may become unnecessary.     

Reconstitution of receptors and GTP-binding regulatory proteins (G proteins) in Sf9 cells. A direct evaluation of selectivity in receptor.G protein coupling

The selectivity in coupling of various receptors to GTP-binding regulatory proteins (G proteins) was examined directly by a novel assay entailing the use of proteins overexpressed in Spodoptera frugiperda (Sf9) cells. Activation of G proteins was monitored in membranes prepared from Sf9 cells co-expressing selected pairs of receptors and G proteins (i.e. alpha, beta1, and gamma2 subunits). Membranes were incubated with [35S]guanosine 5'-(3-O-thio)triphosphate (GTPgammaS) +/- an agonist, and the amount of radiolabel bound to the alpha subunit was quantitated following immunoprecipitation. When expressed without receptor (but with beta1gamma2), the G protein subunits alphaz, alpha12, and alpha13 did not bind appreciable levels of [35S]GTPgammaS, consistent with a minimal level of GDP/[35S]GTPgammaS exchange. In contrast, the subunits alphas and alphaq bound measurable levels of the nucleotide. Co-expression of the 5-hydroxytryptamine1A (5-HT1A) receptor promoted binding of [35S]GTPgammaS to alphaz but not to alpha12, alpha13, or alphas. Binding to alphaz was enhanced by inclusion of serotonin in the assay. Agonist activation of both thrombin and neurokinin-1 receptors promoted a modest increase in [35S]GTPgammaS binding to alphaz and more robust increases in binding to alphaq, alpha12, and alpha13. Binding of [35S]GTPgammaS to alphas was strongly enhanced only by the activated beta1-adrenergic receptor. Our data identify interactions of receptors and G proteins directly, without resort to measurements of effector activity, confirm the coupling of the 5-HT1A receptor to Gz and extend the list of receptors that interact with this unique G protein to the receptors for thrombin and substance P, imply constitutive activity for the 5-HT1A receptor, and demonstrate for the first time that the cloned receptors for thrombin and substance P activate G12 and G13.     

Truncated trkB receptors on nonneuronal cells inhibit BDNF-induced neurite outgrowth in vitro

The function of truncated trkB receptors during nervous system plasticity and regeneration is currently unknown. The extensive nonneuronal localization of truncated trkB-T1 receptors, coupled with their up-regulation by CNS glial cells in response to injury, has led to the speculation that these receptors may sequester BDNF and NT-4/5 to reduce their local availability and, thus, limit axonal sprouting. Conversely, trkB-T1 receptors could bind and present neurotrophins to injured axons and facilitate their regeneration in a manor analogous to that proposed for p75(NTR) receptors on Schwann cells. To address this issue, we used an in vitro coculture paradigm in which wild-type 3T3 NIH fibroblasts or two different 3T3 cell clones stably expressing trkB-T1 receptors served as monolayer substrates upon which to evaluate the effect of trkB-T1 receptors on nonneuronal cells to influence neurotrophin (NGF, BDNF, NT-3, and NT-4/5)-induced neurite outgrowth from retinoic acid (RA)-treated SY5Y neuroblastoma cells. In these experiments, BDNF and NT-4/5 produce a strong phosphorylation of trk receptors on the RA-SY5Y cells and induce differentiation of the SY5Y cells (as measured by the development of neurofilament-positive neuritic processes). This ability of the trkB ligands to stimulate neurite outgrowth is dose dependent since increasing concentrations of BDNF (5, 25, and 100 ng/ml) result in an increased percentage of SY5Y cells developing neurites and in progressively longer neurites from SY5Y cells on the control 3T3 monolayers. In these experiments, BDNF and NT-4/5 induce the strongest neurite outgrowth, followed by NT-3 and then NGF. When trkB-T1 receptors are present on the 3T3 cell substratum both BDNF- and NT-4/5-induced neurite extension from the SY5Y cells are strongly inhibited. In contrast, NGF-induced neurite growth is unaffected and NT-3-associated growth is somewhat reduced. These results suggest that the inhibitory effect of the trkB-T1 receptors on the nonneuronal cell substrates is selective for neurite outgrowth that is mediated via the trkB-kinase receptors on the neuroblastoma cells. This ability of trkB-T1 receptors on the nonneuronal substratum to inhibit BDNF-induced neurite outgrowth can be overcome by the addition of high concentrations of BDNF (1 microg/ml). Binding assays using 125I-BDNF suggest that this inhibitory effect could be mediated via binding and internalization of BDNF by the trkB-T1 receptors on the 3T3 cells. These results provide strong support for the hypothesis that the up-regulation of trkB-T1 receptors on astrocytes following CNS lesions enhances the sequestration of the trkB ligands, BDNF and NT- 4/5, at the site of reactive gliosis and, thus, contributes to the inhibition of CNS axonal regeneration from neurons expressing trkB-kinase receptors by removing their ligands from the extracellular environment.     

Induction of biologically active IL-1 beta-converting enzyme and mature IL-1 beta in human keratinocytes by inflammatory and immunologic stimuli

IL-1beta, a major mediator of inflammatory and immunologic skin disease, undergoes post-translational site-specific cleavage by a novel cysteine protease termed IL-1beta-converting enzyme (ICE). Although in human skin keratinocytes produce significant amounts of the 31-kDa IL-1beta precursor protein, they fail under nonpathologic conditions to convert it to the 17.5-kDa bioactive form. In this study, we examined whether haptens and inflammatory agents might serve as stimuli for ICE activity in human keratinocytes, and, if so, whether ICE activity might precipitate enzymatic processing of IL-1beta to its 17.5-kDa form. Baseline levels of ICE mRNA were detected in keratinocyte cultures devoid of Langerhans cells and were up-regulated by nontoxic concentrations of the reactive hapten urushiol and by the irritant chemicals sodium lauryl sulfate and PMA. Although untreated keratinocytes expressed the 31-kDa form of the protein, 17.5-kDa IL-1beta was easily detected in keratinocytes and keratinocyte supernatants treated with either urushiol or the irritant chemicals. Enzymatic conversion from the 31-kDa to the 17.5-kDa form of IL-1beta was blocked by addition of a highly specific aldehyde inhibitor that contained a tetrapeptide recognition sequence specific for ICE, but not by an aldehyde inhibitor of a related ICE-like cysteine protease. Induction of IL-1beta-converting enzyme by immunologic and inflammatory stimuli may be one of the key regulatory elements in the pathogenesis of allergic and irritant contact hypersensitivity.     

A prospective study of trephined bone grafts of the tibial shaft and iliac crest

BACKGROUND: Our purpose was to analyze the expression of Fas antigen on CD4+ lymphocytes in the aqueous humor (AH) and cerebrospinal fluid (CSF) of patients with Vogt-Koyanagi-Harada disease (VKH). METHODS: Using three-color flow cytometry, we assessed T-lymphocyte subsets stained with fluorescence-conjugated anti-CD3, CD4, CD8, CD29, CD45RA, CD45RO, HLA-DR, and Fas monoclonal antibodies in AH, CSF and peripheral blood (PB) from 8 patients with active VKH. RESULTS: CD3+ T cells constituted the majority of lymphocytes in AH and CSF, in contrast to with PB. The percentages of CD4+ lymphocytes in uveitic AH and CSF were significantly higher than that in PB (P < 0.01). Activated CD4+ and CD8+ cells were significantly more frequent in AH than in CSF and PB (P < 0.01). Although the percentages of CD45RA+ cells within CD4+ cells in AH and CSF were extremely low compared with those in PB, the proportions of CD29+ and CD45RO+ (memory) cells within CD4+ were much higher than those in PB (P < 0.01). Fas antigen was also highly expressed on such CD4+ cells in AH, as in other uveitis patients and on such cells in CSF. Moreover, the percentages of Fas+ and memory cells in AH were significantly higher than those in CSF. CONCLUSIONS: The majority of CD4+ lymphocytes in AH and CSF from patients with active VKH were activated memory cells, on which Fas antigen was also highly expressed. Although this Fas expression may not be an apoptosis-related phenomenon, accumulation of Fas+ memory T lymphocytes in AH and CSF probably reflects the immunopathologic mechanism of VKH.     

The effects of knee reconstruction on combined anterior cruciate ligament and anterolateral capsular deficiencies

We tested the effect of intraarticular reconstructions of the anterior cruciate ligament alone and in combination with extraarticular reconstructions in 10 cadaveric knees. These knees had anterior cruciate ligament deficiency alone or in combination with anterolateral capsuloligamentous deficiencies. In the knees with combined injury, intraarticular reconstruction returned anterior stability to levels not significantly different from levels found for the knees deficient in the anterior cruciate ligament alone and treated with this procedure. After intraarticular reconstruction, rotational stability of the knee with combined injuries failed to return to the levels seen in the knee with isolated anterior cruciate ligament deficiencies that underwent the same treatment. When a tenodesis with either 0 N or 22 N of tension was added to the intraarticular reconstruction in the knee with combined injuries, we found that excessive internal rotation significantly decreased at all angles of flexion, except at full extension with 0 N of tension. In addition, the extraarticular reconstruction with 22 N of tension in the tenodesis overconstrained the knee in internal rotation between 30 degrees and 90 degrees of knee flexion. The tenodesis with 0 N of tension overconstrained the knee at only 60 degrees and 90 degrees of flexion. These results suggest extraarticular reconstruction as an adjunct to the intraarticular operation for the knee with anterior cruciate ligament and anterolateral structural injuries. The results also suggest that the surgeon can affect anterior and rotational laxity by adjusting the tension in the tenodesis.