Cytokine Gene Polymorphisms Associating With Severe Acute Graft-Versus-Host Disease in HLA-Identical Sibling Transplants

It is now well known that the initial phase of graft-versus-hostdisease (GVHD) involves cytokine release during preconditioning of therecipient of an allogeneic bone marrow transplant (BMT). Tumor necrosisfactor (TNF), in particular, has been implicated in pathologicaldamage and is released pretransplant due to irradiation and cytotoxicpreconditioning regimens. Interleukin-10 (IL-10), a naturalimmunosuppressant of TNF , may be involved in downregulation ofthese responses, which may be an individual patient-specific effect. Inthis study, we determined the genotype for polymorphisms associatedwith TNF and IL-10 in 80 potential allo-BMT recipients andcorrelated the genotype with the severity of GVHD in 49 patients forwhom clinical data relating to GVHD was available. The widely studiedTNF 308 polymorphism does not show any significantassociations, but the d3 homozygous allele of the TNFd microsatelliteis preferentially associated with grade III/IV GVHD (7 of 11 patients)compared with its occurrence in 8 of 38 patients with grade 0/II GVHD(P = .006). Alleles of the IL-10 1064 promoterregion microsatellite polymorphism that possess greater numbers ofdinucleotide (CA) repeats also significantly associate with more severeGVHD. This region has been demonstrated to be important in theregulation of the IL-10 promoter. Eighteen of 38 patients with grade0-II GVHD possessed alleles with greater numbers (12 or more) ofdinucleotide repeats, compared with 9 of 11 cases with grade III-IVGVHD (P < .02). Of the 38 patients with grade 0-II GVHD, 3 of38 had a both TNFd3/d3 and IL-10 (12-15) genotype,compared with 6 of 11 patients with grade III-IV GVHD (P < .001). There was no association of either the TNFd or IL-10 microsatellite polymorphisms with mortality (P = .43 and .51, respectively). Our results suggest that patient cytokine gene polymorphism genotypes may influence GVHD outcome by affecting cytokineactivation during the pretransplant conditioning regimens, and theseresults are the first to suggest a genetic predisposition to thisimportant transplant-related complication.     

Serum levels of interleukin-10 in patients with diffuse large cell lymphoma: lack of correlation with prognosis [see comments]

Interleukin-10 (IL-10), also known as cytokine synthesis inhibitory factor, has multiple effects on lymphoid development. In addition, it has been previously reported that serum levels of IL-10 correlate with failure-free and overall survival in patients with non-Hodgkin's lymphoma. In this study, we used a sensitive enzyme-linked immunosorbent assay specific for human IL-10 (lower limit of sensitivity, 5 pg/mL) to measure serum levels in 52 newly diagnosed patients with diffuse large cell lymphoma and at least one adverse prognostic feature who were subsequently treated in a uniform way. Lymphoma patients had significantly higher serum levels of IL-10 (median, 7.98 pg/mL; range, < or = 5 to 27,143 pg/mL) than healthy volunteers (N = 50; median, < or = 5 pg/mL; range, < or = 5 to 19.21 pg/mL) (P = .0000012). Individuals with B symptoms had significantly higher serum levels of IL-10 than those without them (P = .03), but there was no correlation between IL-10 levels and any of the other prognostic variables analyzed, including age, lactic dehydrogenase, beta 2-microglobulin levels, performance status, bulky disease, Ann Arbor stage, or International Index score. More importantly, we found no correlation between IL-10 levels and the achievement of complete remission, nor with failure-free survival or overall survival. We conclude that in a uniform population of untreated patients with diffuse large cell lymphoma, serum levels of IL-10 do not appear to have any prognostic value.     

Erythropoietin stimulates the tyrosine phosphorylation of Shc and its association with Grb2 and a 145-Kd tyrosine phosphorylated protein

Although the erythropoietin receptor (EpR) lacks a tyrosine kinase consensus sequence within its proline-rich intracellular domain, addition of its ligand to Ep-responsive cells stimulates the rapid and transient tyrosine phosphorylation of a number of cellular proteins. The characterization of these phosphorylatable substrates, which include 5 major phosphoproteins with molecular masses of approximately 145, 130, 97, 72, and 56 Kd is an essential step in understanding the signal transduction pathways used by Ep. Recently, we and others have shown that the major 72-Kd tyrosine phosphorylated protein is the EpR itself. We now report, using both murine DA-3 and human MO7E cell lines engineered to express high levels of biologically responsive EpRs (and designated DA-ER and MO7-ER, respectively), that the major 56-Kd tyrosine phosphorylated protein is the recently identified SH2- containing protein, p52shc. Interestingly, in Ep-stimulated cells, anti- Shc antibodies coprecipitate the major 145-Kd tyrosine phosphorylated protein in both DA-ER and MO7-ER cells. Tyrosine phosphorylation of both proteins is detectable within 30 seconds of incubation with Ep at 37 degrees C, reaches a maximum between 2 and 5 minutes, and declines by 30 minutes. In addition, tyrosine phosphorylated Shc appears capable of associating with the activated EpR, but this could only be shown in MO7-ER cells. Lastly, as has been shown previously with the tyrosine kinase containing receptors for epidermal growth factor, platelet derived growth factor, and insulin, activation of the EpR leads to the association of p52shc with the 25-Kd polypeptide, Grb2. Taken together, our data suggest that the previously reported increases in rasGTP observed with Ep result, in part, from the tyrosine phosphorylation of Shc and its association with Grb2 and/or a tyrosine phosphorylated 145- Kd protein.     

Sulfhydryl reducing agents and shape regulation in human erythrocytes

Metabolic crenation of red cells is reversible; on addition of nutrients, echinocytes recover the normal discoid shape. When the shape recovery takes place in the presence of reducing agents such as dithiothreitol (DTT), morphological change continues until the cells are stomatocytic. The degree of stomatocytosis varies, depending on the cell morphology when the nutrients and reducing agent are added. DTT has minimal effect on the shape of normal discocytes, but in its presence, mildly echinocytic cells become slightly cupped and advanced- stage echinocytes become severely stomatocytic. DTT must be present continuously for development and retention of stomatocytosis; echinocytes preincubated with or metabolically depleted in DTT do not become stomatocytic when supplemented in the absence of DTT, and DTT- induced stomatocytes revert to discocytes when the reducing agent is removed. DTT has no effect on adenosine triphosphate synthesis or equilibrium cell glutathione levels, and the induced stomatocytosis is not inhibited by excluding oxygen from cells during depletion. Spectrin phosphorylation and phosphate turnover are not affected by DTT. The echinocyte-to-discocyte transformation coincides with phosphorylation of membrane inner monolayer lipids (diacylglycerol to phosphatidic acid and phosphatidylinositol to phosphatidylinositol-4,5-bisphosphate). Overphosphorylation of these phospholipids is not responsible for the exaggerated shape recovery seen with reducing agents; phosphorylation of inner monolayer lipids proceeds identically in the presence and absence of DTT.     

Association between autoimmune thyroid disease and Familial Alzheimers disease

OBJECTIVE: To determine the prevalence of autoimmune thyroid disease in Familial Alzheimer's Disease kindreds and to ascertain whether there is any evidence for genetic linkage between the two conditions. DESIGN: Retrospective study of Familial Alzheimer's Disease kindreds. PATIENTS: Seventy affected and unaffected family members from 12 kindreds. MEASUREMENTS: Anti-thyroglobulin and anti-microsomal autoantibody status was determined using an enzyme-linked immunosorbent assay. Thyrotrophin levels were determined by an immunoradiometric assay. RESULTS: Of the family members, 41.4% had evidence of autoimmune thyroid disease, with significant co-segregation between the presence of thyroid autoantibodies and the development of Alzheimer's disease (P less than 0.01). CONCLUSIONS: This study demonstrates a very high prevalence of autoimmune thyroid disease in Familial Alzheimer's Disease kindreds and suggests that a genetic factor contributing towards the development of autoimmune thyroid disease may be located on chromosome 21 within close proximity to the Familial Alzheimer's Disease gene.     

Detection of benzo[a]pyrene diol epoxide-DNA adducts in human placenta.

Human placenta is a readily available organ that responds to maternal environmental insult and has been previously used to investigate metabolism and bioactivation of procarcinogens, for example, benzo[a]pyrene. HPLC in combination with synchronous fluorescence spectroscopy was used to examine 28 placentas for the presence of benzo[a]pyrene diol epoxide-DNA adducts, and 10 of these were found to be positive. DNA samples from these placentas were subsequently pooled and subjected to partial enzymatic digestion to oligonucleotide fragments. Concentration of those DNA fragments containing benzo[a]pyrene diol epoxide-DNA adducts was achieved by immunoaffinity chromatography with polyclonal antibodies raised against these adducts. Column eluates were hydrolyzed under mild acid conditions and extracted with an organic solvent. The presence of benzo[a]pyrene-7,10/8,9-tetrahydrotetrol residues in the extracts was determined by HPLC and synchronous fluorescence spectroscopy and was confirmed by GC/MS. The results unequivocally confirm bioactivation and formation of DNA adducts from benzo[a]pyrene in human placenta in vivo and establish a methodological approach to direct measurement of carcinogen-DNA adducts that are formed as a result of human environmental exposure.     

Proteolytic inactivation of human factor VIII procoagulant protein by activated human protein C and its analogy with factor V

Purified human factor VIII procoagulant protein (VIII:C) was treated with purified human activated protein C (APC) and the loss of VIII:C activity correlated with proteolysis of the VIII:C polypeptides. APC proteolyzed all VIII:C polypeptides with mol wt = 92,000 or greater, but not the doublet at mol wt = 79-80,000. These results and our previous thrombin activation studies of purified VIII:C, are analogous with similar studies of factor V and form the basis for the following hypothesis: activated VIII:C consists of heavy and light chain polypeptides [mol wt = 92,000 and mol wt = 79-80,000 (or 71-72,000), respectively] which are similar in Mr to the heavy and light chains of activated factor V. Thrombin activates VIII:C and V by generating these polypeptide chains from larger precursors and APC inactivates both molecules by cleavage at a site located in the heavy chain region of activated VIII:C and V.     

Blood pressure-related cognitive decline: does age make a difference?

Systolic and diastolic blood pressures have been inversely related to cognitive performance in prospective and cross-sectional studies. However, in large, community-based samples, these findings have been limited to older adults. In this 20-year longitudinal study, we examined the relationship between baseline blood pressure and cognitive decline for 529 participants using 2 age groups (18 to 46 years and 47 to 83 years). Cognitive performance was measured over multiple examinations with the Wechsler Adult Intelligence Scale from which 4 scores were derived by factor analysis. A 2-stage growth curve method of analysis was used to model cognitive change. Results indicated that higher levels of baseline systolic blood pressure, diastolic blood pressure, mean arterial pressure, and blood pressure categories as defined by the Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure were significantly associated with decline in Visualization/Fluid abilities in both younger and older age groups. Young adults are as susceptible to blood pressure-related longitudinal decline in cognitive performance as are older adults.