Cryptosporidium parvum Infection of Human Intestinal Xenografts in SCID Mice Induces Production of Human Tumor Necrosis Factor Alpha and Interleukin-8

The protozoan parasite Cryptosporidium parvum invades intestinal epithelial cells and can cause life-threatening diarrhea in immunocompromised individuals. Despite the clinical importance of this organism, much remains to be learned about the pathogenesis of C. parvum-induced diarrhea. To explore the role of the intestinal inflammatory response in C. parvum disease, using C. parvum oocysts we infected human intestinal xenografts in severe combined immunodeficient (SCID) mice. Seven days after infection, we found levels of human tumor necrosis factor alpha and interleukin-8 in C. parvum-infected human intestinal xenografts that were significantly higher than those seen in uninfected control xenografts. These results demonstrate that human intestinal cells produce proinflammatory cytokines in response to C. parvum infection and establish SCID-HU-INT mice as a model system to study the interactions of C. parvum with the human intestine.     

Genetic control of the antibody response to poly(L Tyr, L Glu)-poly(DL Ala)--poly(L Lys) in mice: analysis of (low responder x low responder)F1 hybrids

The antibody response to the synthetic polypeptide poly (L Tyr, L Glu)-poly (DL Ala)--poly (L Lys) designated (T,G)-A--L, was investigated in inbred, congenic, F1 and F2 hybrid strains of mice. The antibody response was analysed at both low (10 microgram) and high (50 microgram) immunizing doses of (T,G)-A--L. Antibodies were measured using both a modified Farr assay and a plate binding assay. At low immunizing doses it was found that all of the congenic and non-congenic (low responder x low responder) F1 hybrids were low responders. However, the quantitative antibody response of one non-congenic (low responder x low responder) F2 hybrid segregated in a 1:1 ratio of high responders to low responders, suggesting some form of complementation of (T,G)-A--L Ir genes. At high immunizing doses it was found that congenic and non-congenic (low responder x low responder) F1 hybrids were all high responders, indicating a complementation of Ir genes to (T,G)-A--L. This complementation was confirmed using two different routes of immunization, namely footpad and intraperitoneal. Furthermore the quantitative antibody responses of (low responder x low responder) F2 hybrids segregate in a 1:1 ratio of high responders to low responders. The class of antibodies produced to (T,G)-A--L in (low responder x low responder) F1 hybrids was determined by gel filtration on Sephadex G-200, and found to be predominantly IgG, with lesser amounts of IgM.     

Transgenic and Natural Mouse Models of Proteolipid Protein (PLP)-Related Dysmyelination and Demyelination

The X chromosome-linked PLP/DM-20 gene is the CNS myelin gene most frequently associated with mutations, resulting in dysmyelination in several species including man (Pelizaeus-Merzbacher disease, X-linked Spastic Paraplegia). The pathology of most PLP gene mutations is characterized by hypomyelination, glial cell proliferation, increased numbers of microglia, and premature oligodendrocyte death. In most mutants, residual myelin structures have an abnormal ultrastructure and periodicity. Surprisingly, transgenic mice which carry extra copies of the wild type PLP gene show dysmyelination, demonstrating that the PLP gene is dosage sensitive. Pathological changes of transgenic mice vary from the phenotype of natural mutants. Specifically, many Golgi saccules of oligodendrocytes are vacuolated and the cytoplasm contains autophagic vacuoles hinting at a perturbation in protein trafficking. In fact, upon transgenic overexpression PLP becomes a prominent peripheral myelin protein, whereas in normal Schwann cells PLP is restricted from entering the myelin compartment. Surprisingly, transgenic animals which overexpress PLP/DM-20 at a low level appear normal during early development, but later spontaneously demyelinate. The mechanisms underlying this demyelination phenotype is unknown but an immune-mediated process has been suggested. All attempts to correct the phenotype of natural PLP mutants, such as jimpy mice, with a wild type transgene have had little effects, indicating a dominant-negative effect of the mutant gene product. On the other hand, mice with a targeted disruption of the PLP/DM-20 gene have suprisingly minor clinical signs. This suggests that the lethal phenotype associated with the majority of PLP gene mutations is a complex combination of loss and gain-of-function effects of a mutant myelin protein.     

Direct modulatory effect of C-reactive protein on primary human monocyte adhesion to human endothelial cells

C-reactive protein (CRP) is the prototypic acute phase serum protein in humans. The effects of CRP on primary human monocyte adhesion molecule expression and interaction with the endothelium have not been studied. Herein, we describe an investigation into the phenotypic and functional consequences of CRP binding to peripheral blood monocytes ex vivo. Peripheral whole blood was collected from healthy, non-smoking males. Mononuclear cells (MNC) and monocytes were isolated by differential centrifugation using lymphoprep and Dynal negative isolation kit, respectively. Cells were exposed to CRP from 0 to 250 micro g/ml for 0-60 min at 37 degrees C and analysed for (a) CD11b, PECAM-1 (CD31) and CD32 expression by flow cytometry and (b) adhesion to LPS (1 micro g/ml; 0-24 h) treated human umbilical vein endothelial cells (HUVEC). CD14+ monocyte expression of CD11b increased significantly up to twofold when exposed to CRP, compared to controls. There was no significant difference in CD32 expression, whereas CD31 expression decreased after exposure to CRP. CRP treatment of monocytes inhibited their adhesion to early LPS-activated HUVEC (0-5 h). However, the adhesion of CRP-treated monocytes to HUVEC was significantly greater to late activation antigens on HUVEC (24 h, LPS) compared to controls. We have shown that CRP can affect monocyte activation ex vivo and induce phenotypic changes that result in an altered recruitment to endothelial cells. This study provides the first evidence for a further role for C-reactive protein in both monocyte activation and adhesion, which may be of importance during an inflammatory event.     

Comparison of the real-time PCR method and the Gen-Probe amplified Mycobacterium tuberculosis direct test for detection of Mycobacterium tuberculosis in pulmonary and nonpulmonary specimens

Real-time PCR was compared to Amplified Mycobacterium tuberculosis Direct Test (AMTDII) for 100 clinical specimens. The overall sensitivities of the real-time PCR method and AMTDII were similar for respiratory and nonrespiratory specimens. However, real-time PCR seemed to be less susceptible to amplification inhibitors than AMTDII.     

Bladder cancer immunogenicity: expression of CD80 and CD86 is insufficient to allow primary CD4+ T cell activation in vitro

Transitional cell carcinomas (TCC) of the urinary bladder are known to express proteins which can yield potentially immunogenic peptide epitopes for expression in the context of cell surface class I or class II MHC antigens. However, additional costimulatory ligands must also be expressed before such a cell might directly induce full activation and proliferation of resting, antigen-specific T lymphocytes. Intravesical therapy might be used to manipulate T cell costimulation in order to promote specific rejection of TCC cells. This in vitro study examined the potential of such a strategy by transfection of the prototypical TCC line J82 with the important costimulatory molecules CD80 (B7-1) and CD86 (B7-2). Untransfected J82 cells expressed class I and II MHC antigens, a range of cell adhesion molecules, though did not induce T cell proliferation in a robust, allogeneic co-culture system. Transfected J82 cells expressed CD80 or CD86 at levels comparable to an antigen-presenting B cell line. Furthermore, functional surface expression of CD80 and CD86 was demonstrated in a mitogen-dependent assay of costimulation. However, neither CD80+ nor CD86+ transfectant J82 cells could induce significant proliferation of antigen-specific CD4+ T cells. Further analysis showed that bystander J82 cells could inhibit independent T cell activation in an effect dependent on direct cell contact. This inhibitory effect was associated with increased cell death in the responding lymphocyte population and is concordant with surface expression of CD95L by the J82 cell line.     

Interpretation of gas-blood ratio inP O 2polarography

Summary The differences inP _{O 2}readings between gas and blood were studied with a Clark-type electrode in the range of 38.5 to 713 mm HgP _{O 2}.The tonometered blood samples were taken in two different ways. The results showed that the gas-blood ratior _b(equilibrating gasP _{O 2}reading/equilibrated bloodP _{O 2}reading) depended not only on the sampling method but also on theP _{O 2}range: it varied from 1.005 to 1.032 for aP _{O 2}of 96.5 mm Hg, and from 1.040 to 1.081 for aP _{O 2}of 713 mm Hg according to the sampling procedure.A theoretical analysis demonstrated that the variation ofr _bwith the bloodP _{O 2}can be attributed to the influence of the degree of oxygen saturation of the hemoglobin on theP _{O 2}gradient existing in the blood diffusion boundary layer adhering to the electrode membrane.This work was supported by grants from the High Authority of the European Coal and Steel Community and from the Fonds de la Recherche Scientifique Médicale, Belgium.     

Response of left ventricular diastolic filling to graded exercise relative to the lactate threshold

Summary During incremental exercise, the left ventricular ejection fraction increases up to the intensity of the anaerobic threshold and tends to level off at higher exercise intensities. Since there is a correlation between the response of peak filling rate and ejection fraction to exercise, this study was conducted to determine whether the response of left ventricular diastolic function is similar to the response of systolic function relative to lactate threshold. Twelve healthy men performed two exercise tests on a cycle ergometer. In the first test, lactate threshold and maximal power output were determined. In the second exercise test, gated radionuclide ventriculography was performed at rest, at the lactate threshold intensity, and at peak exercise to measure ejection fraction and peak filling rate. Ejection fraction increased significantly from rest [mean (SD): 62 (5)%] to lactate threshold [76 (7) %] and did not change significantly from lactate threshold to peak exercise [77 (7)%]. Likewise, peak filling rate (normalized for stroke counts) increased from resting [6.1 (0.9)V _s · s^–1] to lactate threshold [9.4 (1.8)V _s · s^–1] and did not change significantly from lactate threshold to peak exercise [9.6 (2.9)V _s · s^–1]. There was no correlation between the change in peak filling rate and the change in ejection fraction from rest to lactate threshold. Thus, during incremental exercise, left ventricular diastolic function responds qualitatively similar to systolic function.     

Transition from a botanical to a molecular classification in tree pollen allergy: implications for diagnosis and therapy

Tree pollens are among the most important allergen sources. Allergic cross-reactivity to pollens of trees from various plant orders has so far been classified according to botanical relationships. In this context, cross-reactivities to pollens of trees of the Fagales order (birch, alder, hazel, hornbeam, oak, chestnut), fruits and vegetables, between pollens of the Scrophulariales (olive, ash, plantain, privet, lilac) and pollens of the Coniferales (cedar, cypress, pine) are well established. The application of molecular biology methods for allergen characterization has revealed the molecular nature of many important tree pollen allergens. We review the spectrum of tree pollen allergens and propose a classification of tree pollen and related allergies based on major allergen molecules instead of botanical relationships among the allergenic sources. This molecular classification suggests the major birch pollen allergen, Bet v 1 as a marker for Fagales pollen and related plant food allergies, the major olive pollen allergen, Ole e 1, as a possible marker for Scrophulariales pollen allergy and the cedar allergens, Cry j 1 and Cry j 2, as potential markers for allergy to Coniferales pollens. We exemplify for Fagales pollen allergy and Bet v 1 that major marker allergens are diagnostic tools to determine the disease-eliciting allergen source. Information obtained by diagnostic testing with marker allergens will be important for the appropriate selection of patients for allergen-specific forms of therapy.